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Functional impact of microRNA regulation in models of extreme stress adaptation

Functional impact of microRNA regulation in models of extreme stress adaptation doi:10.1093/jmcb/mjx053 Journal of Molecular Cell Biology (2018), 10(2), 93–101 j 93 Published online January 9, 2018 Review Functional impact of microRNA regulation in models of extreme stress adaptation Kyle K. Biggar and Kenneth B. Storey Institute of Biochemistry & Department of Biology, Carleton University, Ottawa, ON K1S 5B6, Canada * Correspondence to: Kenneth B. Storey, E-mail: kenneth_storey@carleton.ca Edited by Zefeng Wang When confronted with severe environmental stress, some animals are able to undergo a substantial reorganization of their cellular environment that enables long-term survival. One molecular mechanism of adaptation that has received considerable attention in recent years has been the action of reversible transcriptome regulation by microRNA. The implementation of new computational and high-throughput experimental approaches has started to uncover the vital contributions of microRNA towards stress adaptation. Indeed, recent studies have suggested that microRNA may have a major regulatory influence over a number of cellular processes that are essential to prolonged environmental stress survival. To date, a number of studies have highlighted the role of microRNA in the regulation of a metabolically depressed state, documenting stress-responsive microRNA expression during mammalian hibernation, frog and insect freeze tolerance, and turtle and marine snail anoxia tolerance. These studies collectively indicate a conserved principle of microRNA stress response across phylogeny. As we are on the verge of dissecting the role of microRNA in environmental stress adaptation, this review summarizes recent research advances and the hallmark expression patterns that facilitate stress survival. Keywords: hypometabolism, metabolism, temperature, metabolic rate depression, stress response Introduction readily reversible, and capable of eliciting selective control over A select few animals have developed an incredible ability to both essential and non-essential cellular processes. This review survive and overcome severe environmental challenges that will discuss new advances in the roles of non-coding RNA in pro- include dehydration, oxygen deprivation (i.e. anoxia), as well as moting animal survival in extreme environments, as well as the temperature changes and even freezing of body fluids (Storey transition into (and survival within) the hypometabolic state. The and Storey, 2004). These fascinating feats and the study of how function of small non-coding RNA, namely microRNA (miRNA), will these resilient animals are able to survive such extreme stresses be a primary focus given the extensive amount of miRNA research has captivated researchers for many years. In particular, the past that has taken place in this field over the past 10 years (Morin few decades research in comparative stress biology has focused et al., 2007; Biggar and Storey, 2015a, b; Frigault et al., 2017). on elucidating the molecular survival mechanisms at play, those Mechanisms that help to coordinate stress survival must be easily working to reorganize the cellular landscape for life in a new inducible and reversible so that they can be initiated once a stress is environmental extreme (Storey and Storey, 2004; Storey, 2015). detected. The contribution of readily reversible protein modifica- These mechanisms work to reprioritize cellular processes, such tions, such as Ser/Thr/Tyr phosphorylation, has been extensively that these animals can conserve vital energy stores, with some studied as a dynamic mechanism to quicklymodifyprotein function entering a state of dormancy that is characterized by the depres- and achieve hypometabolism. These studies established that the sion of metabolic rate to 10%–30% of basal levels (or even basic principles involved in regulating stress-responsive signal trans- more), a state commonly referred to as hypometabolism (Guppy duction events included the coordinated reversible phosphorylation and Withers, 1999). Given the relatively rapid onset of many of many proteins, setting this modification as one of the hallmarks of environmental stresses, molecular mechanisms that help to gov- the central mechanisms of metabolic reorganization for many years ern the transition into a hypometabolic state must be rapid, (Cowan and Storey, 2003). Indeed, reversible post-translational modifications have been shown to affect the function of numerous cellular proteins and to serve as a common response element in Received August 13, 2017. Revised November 20, 2017. Accepted November 30, the survival of many environmental challenges and metabolic stresses (Brooks and Storey, 1995; Cowan et al., 2000; Biggar © The Author (2018). Published by Oxford University Press on behalf of Journal of and Storey, 2012; Wu and Storey, 2013). Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 94 j Biggar and Storey Mechanisms of reversible control over protein translation mRNA content, but rather achieved through post-translational Beginning in the early 1990s, it was established that an over- modifications made to translational machinery. all suppression of the energetically costly cellular process of Despite a possible global shutdown of protein translation protein translation was an essential part of hypometabolism pathways, there still remains the question of how the proteins (Brooks and Storey, 1993; Land et al., 1993; Frerichs et al., that are essential for survival continue to be created in the 1998; Fraser et al., 2001; Larade and Storey, 2002; Hittel and hypometabolic state. There must be mechanisms at play that Storey, 2002). Indeed, a depression of translational rate neces- allow for essential mRNA to be translated, while non-essential sarily contributes to energy rationing, thereby helping to reserve mRNA are blocked from translation. This requirement highlights ATP turnover for essential survival-related cellular processes. the regulatory influence of miRNA in the translational control of For example, the rate of protein translation in oxygen-deprived specific mRNA. Perhaps such regulatory mechanisms would turtles was shown to decrease below measurable levels follow- allow gene-specific control over protein translation during peri- ing 3 h of anoxic exposure (at 23°C) as compared with normoxic ods of cell stress. Indeed, this area of research has gained con- values at the same temperature (Fraser et al., 2001). Similarly, siderable interest in recent years within the field of comparative in the brain of torpid thirteen-lined ground squirrels, the rate of biochemistry. protein translation was documented to be only 0.04% of euther- mic rates (Frerichs et al., 1998) and, when corrected for differ- Reversible control of protein translation by microRNA ences in body temperature between euthermic and torpid state, MiRNA are short, non-coding RNA capable of rapidly and rates in torpor were still approximately only one-third of the reversibly regulating the expression of targeted proteins within euthermic values. Together, these results demonstrated that a cell and have been found in plants, animals, bacteria, as well temperature influences alone do not explain reductions in trans- as some viruses. It is well documented that these 18–24 nt tran- lational rates and that the depression of protein translation scripts are able to bind, with full or partial complementarity, to likely involves other molecular mechanisms. Indeed, the sup- specific mRNA targets, resulting in either the inhibition of trans- pression of protein translation in the hypometabolic state is a lation or degradation of the target mRNA (Bartel, 2004). As a multifaceted event, one that is likely achieved through a com- reflection of their regulatory potential, miRNA have been pre- bination of different mechanisms that include reductions in dicted to be involved in all biological process in some aspect gene transcription, inactivation of the translational machinery, throughout the animal kingdom (Friedman et al., 2009). and/or through mRNA-specific interference by miRNA. The biogenesis of a mature miRNA begins with the initial tran- As a result of the high energetic costs of protein synthesis scription of a primary miRNA (pri-miRNA). The pri-miRNA tran- and mRNA transcription (accounting for greater than 25%of script is much larger than the final mature miRNA product, basal ATP turnover in many instances), it would not be surpris- ranging from hundreds to thousands of nucleotides in length ing to see that a strong stress-responsive suppression of mRNA (Krol et al., 2010). Following transcription, the pri-miRNA is then transcription would accompany the observed stress-responsive cleaved to form a hairpin RNA structure called a precursor decrease in translational rate (Hochachka et al., 1996; Podrabsky miRNA (pre-miRNA) and is exported from the nucleus (Yi et al., and Hand, 2000). In this regard, previously studies have shown 2003; Zeng et al., 2005; Krol et al., 2010). Once in the cyto- that reversible control over translational machinery may be a plasm, the pre-miRNA is further processed into a mature miRNA powerful driving force behind global translational silencing during duplex by Dicer, an riboendonuclease protein. From the mature hypometabolism. For example, research on protein translation in miRNA duplex, it is suggested that only one strand will function hibernating thirteen-lined ground squirrels has documented a as a mature miRNA, leaving the passenger strand to be dynamic stress-responsive increase in the relative phosphorylation degraded (Krol et al., 2010). Although this is generally thought of the translational initiation factor eIF2α (Hittel and Storey, to be the case, the idea of passenger miRNA degradation has 2002). The phosphorylation of eIF2α is a well-established marker been challenged in light of recent research suggesting that the of both translational arrest and the disruption of active polyribo- abundance, possible function, and physiological relevance of somes (Yamasaki and Anderson, 2008). Indeed, the same study of passenger miRNA has been underestimated (Cipolla, 2014). hibernating ground squirrels also demonstrated a hibernation- Following mature miRNA processing, the mature miRNA is then responsive disaggregation of polyribosomes (Hittel and Storey, loaded into a protein complex referred to as the RNA-induced 2002). Both of these results indicate that a translationally silent silencing complex (RISC). Interestingly, the miRNA–RISC and state is imposed by a stress-responsive inactivation of the transla- bound mRNA complex have also been found to aggregate within tional machinery. Furthermore, neither the total cellular polyade- stress granules and p-bodies in a stress-responsive manner, nylated RNA content nor the mRNA transcript levels of many such as in response to DNA damage or hypoxia stresses, and genes were found to decrease in hibernation or anoxia models of are eventually either degraded or directed into translation hypometabolism (Epperson and Martin, 2002; Storey and Storey, once the stress is lifted (Pothof et al., 2009; Wu et al., 2011). 2004; Rouble et al., 2014). Therefore, it is reasonable to conclude Indeed, the stress-responsive formation of reversible stress that the suppression of global protein translation rates during granules have been shown to occur in response to mammalian hypometabolism may not be a direct result of altered cellular hibernation (Tessier et al., 2014). Take together, this reversible Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 95 mRNA storage system represents a promising mechanism for as part of winter survival by freeze-tolerant (Littorina littorea, reversible translational arrest in models of hypometabolism. Eurosta solidaginis) and freeze-avoidant (Epiblema scudderiana) species (Biggar et al., 2012; Courteau et al., 2012; Lyons et al., MicroRNA discovery in non-model organisms 2013, 2015a, b, 2016). In general, miRNA and their target recognition sites have Given the inherent difficulty of studying poorly conserved been subject to exquisite conservation across phylogeny. The miRNA and an interest in discovering possible specifies-specific conservation of miRNA by >90% among vertebrates is reflective miRNA, computation-based methods have recently been devel- of the important regulatory function they impose in normal cell oped specifically to identify (i) highly conserved, (ii) poorly con- biology. This high level of sequence homology between species served, and (iii) species-specific miRNA from organisms with has made it possible to study miRNA expression patterns in newly sequenced genomes. The widely used miRDeep2 platform many different species with relative ease. Indeed, multiple stud- accomplishes this by using both an available genome sequence ies have been able to explore the dynamic expression of miRNA and small RNA sequencing information to map mature miRNA in response to a variety of environmental stresses in a diverse sequences back to a reference genomic location and to then array of animals for which little or no genomic sequence infor- evaluate the associated pre-miRNA structure and identify high- mation is available. To date, stress-responsive miRNA expres- confidence miRNA (Friedlander et al., 2012). Given the import- sion has been studied in models of hypometabolism that ance miRNA sequence on its function, it is essential to validate include ground squirrels, bats, frogs, turtles, and many inverte- all computationally predicted candidate miRNAs experimentally, brate species (Table 1 with references). Indeed the dynamic including PCR-based methods, such that their expression and change in stress-responsive miRNA expression may be the result sequence of their mature forms can be verified. Recently, small of multiple regulatory steps, including transcriptional modula- RNA sequencing and miRDeep2 have been used to identify tion, microprocessor or miRNA biogenesis co-factor regulation, novel miRNA regulating gene expression during hibernation in changes in RNA/protein complex localization, and modifications thirteen-lined ground squirrels, Ictidomys tridecemlineatus (Luu of miRNA ends. Although the exact mechanisms controlling et al., 2016). The study identified and experimentally validated stress-induced miRNA expression are not yet known, some of 17 novel miRNA and characterized their relative expression in these steps may play pivotal roles in cellular homeostasis in the liver, skeletal muscle, and heart tissues over the torpor–arousal context of the stress response. cycle. Interestingly, these squirrel-specific miRNA were predicted Much of the initial hypometabolism-focused miRNA research to target mRNA enriched in biological processes that are known was carried out on vertebrate and invertebrate species with lim- to be dynamically regulated during hibernation, including lipid ited genomic sequence information available (Morin et al., metabolism, ion-transport ATPases, and various cellular signal- 2007; Biggar et al., 2009, 2012; Courteau et al., 2012; Lyons ing cascades (Luu et al., 2016). This study provides an example et al., 2013). Given this restriction, these studies were limited to of how computational identification of new miRNA and analysis the analysis of highly conserved miRNA using methods that had of their cellular function is of growing interest in comparative been developed to aid in the amplification, sequencing, and val- research on environmental stress adaptation. Indeed, such idation of these conserved miRNA (Biggar et al., 2011, 2014). knowledge may be crucial to developing a deeper understand- However, while vertebrate miRNA are highly conserved, low con- ing of the roles that miRNA may play in the hypometabolic servation between vertebrate and invertebrate miRNA slowed stress response. The major restriction of this particular platform the initial progress of miRNA research in invertebrate species. is the requirement for small RNA sequencing information. In the Despite this challenge, several studies have explored the regula- absence of this information, another platform, called SMIRP, tion of various relatively conserved invertebrate miRNA, particularly has been developed recently to identify species-specific miRNA Table 1 Studies exploring the role of stress-responsive miRNA regulation in animal models of hypometabolism. Stress Species References Hibernation Ictidomys tridecemlineatus Morin et al. (2007), Maistrovski et al. (2012), Lang-Ouellette and Morin (2014), Wu et al. (2014a), Luu et al. (2016), Wu et al. (2016), Frigault et al. (2016) Spermophilus parryii Liu et al. (2010) Myotis lucifigus Biggar and Storey (2014a, b), Kornfeld et al. (2012), Maistrovski et al. (2012) Dromiciops gliroides Hadj-Moussa et al. (2016) Freezing Rana sylvatica Biggar et al. (2009), Bansal et al. (2016) Littorina littorea Biggar et al. (2012) Eurosta solidaginis Courteau et al. (2012), Lyons et al. (2013), Lyons et al. (2015a, b), Lyons et al. (2016) Chrysemys picta Shaffer et al. (2013), Biggar and Storey (2015a, b) Freeze avoidance Epiblema scudderiana Lyons et al. (2015a, b) Oxygen restriction Littorina littorea Biggar et al. (2012) Trachemys scripta elegans Biggar and Storey (2011), Zhang et al. (2013) Ictidomys tridecemlineatus Lee et al. (2012) Aestivation Xenopus laevis Wu et al. (2013), Luu and Storey (2015) Apostichopus japonicus Chen et al. (2013), Chen and Storey (2014) Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 96 j Biggar and Storey de novo directly from an available genome (Peace et al., 2015). ground squirrels, documenting the torpor-responsive expression The SMIRP platform identifies both known and novel species- of 117 miRNA across four stages of the torpor–arousal cycle specific miRNA by scanning the genome for pre-miRNA-like (Wu et al., 2016). A follow-up study identified 17 additional novel hairpins and evaluating select features of these hairpins (i.e. torpor-responsive miRNA, currently thought to be unique to the % nucleotide content, RNA folding measures, and other topo- thirteen-lined ground squirrel (Luu et al., 2016). Another study uti- logical descriptors) against miRNA that have been previously lized advances in parallel RNA sequencing to study the expression identified from related species (Peace et al., 2015; Schaap of >200 ground squirrel miRNA, including 18 novel miRNA, in the et al., 2016; Adema et al., 2017). liver of the hibernating Arctic ground squirrel, Spermophilus par- Recently, the SMIRP platform was used to identify species- ryii (Liu et al., 2010). To date, the regulatory influence of miRNA specific miRNA from the schistosomiasis-transmitting freshwater expression has been most extensively explored in the context of Ramshorn snail (Biomphalara glabrata)(Adema et al., 2017). mammalian hibernation (Table 1). These studies, and those of a Identification of these miRNA provided information on miRNA variety of other animals, have demonstrated the importance of evolution, conservation, and suggested influence on transla- studying these small regulatory molecules as a conserved dynamic tional regulation that may lead to possible mechanisms for response to environmental stress and adaptation across phyl- population control in this snail species. Researchers from this ogeny. Given the growing implication of miRNA in the regulation of study performed an in silico prediction of miRNA using SMIRP environmental stress tolerance, studies have now begun to dig within the B. glabrata genome scaffolds. Uniquely, SMIRP lever- deeper into miRNA function, asking specific research questions, aged a novel approach to miRNA classifier construction in which such as what miRNA are stress responsive? what mRNA are stress- models are built dynamically and are targeted toward B. glabra- responsive miRNA targeting? what are the global functional impli- ta. This differs from other methods (e.g. precursor miRNA pre- cations of miRNA? diction methods) that attempt to produce generalized models that are applicable to a large number of species. From this MicroRNA-influenced control over cellular pathways and approach, 202 pre-miRNA (95 known and 107 novels) and asso- processes ciated mature miRNA from B. glabrata were identified (Adema It is well established that a single miRNA sequence can exert et al., 2017). No homologous sequences to the identified novel regulatory effects on numerous different targets. Similarly, a single B. glabrata precursor miRNA were found in either the sea slug mRNA is likely the regulatory target of multiple miRNA, with differ- (Aplysia californica) or the sea snail (Lottia gigantea) annotated ent miRNA binding at varied locations within the 3′UTR. This com- miRNA, or present within their available genomes. Interestingly, plex regulatory system creates a model of enormous regulatory this study also identified the possible biological context of novel potential that cannot be ignored when studying miRNA function. B. glabrata miRNA, predicting mRNA targets from the 3′UTR of However, given the limited genomic resources for many experi- available B. glabrata transcripts (John et al., 2005). A significant mental models of hypometabolism, initial explorations into miRNA proportion of the identified target genes of these novel miRNA regulatory potential have been appropriately limited in scope. included multi-miRNA gene regulation of proteins involved in cel- These studies primarily characterized miRNA and target expression lular processes such as secretory mucal proteins (mucin-21-like) patterns in asinglemiRNA,singlemRNAtargetformat (Morin (Gabrial et al., 2011), matricellular proteins (thrombospondin-3b- et al., 2007; Biggar et al, 2009, 2012). One good example from like) (Marxen and Becker, 2011) and shell formation proteins (den- early miRNA research on models of hypometabolism is the stress- tin sialophosphoprotein-like) (Volk et al., 2014)(Figure 1). These responsive characterization of miR-21. This miRNA has been stud- newly identified miRNA greatly enrich the repertoire of known mol- ied in multiple animals in the context of regulating anti-apoptotic lusk miRNA, providing insights into mollusk miRNA function, as well genes in response to environmental stress (Morin et al., 2007; as their evolution and biogenesis. Such species-specific miRNA can Biggar et al., 2009; Biggar and Storey, 2011, 2012; Wu et al., also provide possible biocontrol targets for B. glabrata population 2014b). Following the development of bioinformatics methods that control, or may even play a role in the control of aestivation in this allowed for the widespread study of multiple miRNA from a single species (Britton et al., 2014). set of RNA samples (Biggar et al., 2014), recent studies began to move away from single miRNA:target-based candidate characteriza- MicroRNA and adaptation to extreme environments tion. The scope of current studies is growing larger as we begin to A growing number of studies have begun to highlight the con- learn more about miRNA target selection and begin to apply new served response of miRNA in the regulation of the hypometabolic computational-based methods to explore the regulatory impact of a state. Beginning in 2007, the first study of this type explored the greater set of stress-responsive miRNA on the complete system of expression of several well-conserved miRNA in response to sea- cellular processes (Luu et al., 2016; Wu et al., 2016). sonal hibernation in the thirteen-lined ground squirrel (Morin In 2016, an expansive study looking at the torpor-responsive et al., 2007). Although only exploring the expression of a handful expression of 117 conserved miRNA in hibernating thirteen-lined of miRNA, this was a keystone paper in introducing the possible ground squirrels over four stages of the torpor–arousal cycle influence of miRNA as a mechanism to reversibly control mRNA (euthermia, early torpor, late torpor, and interbout arousal) (Wu translation in models of hypometabolism. More recently, several et al., 2016). Moving away from candidate miRNA expression other studies have explored miRNA expression in hibernation of analysis, this study found significant differential expression of a Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 97 Figure 1 Prediction of species-specific miRNA function from the freshwater Ramshorn snail, Biomphalaria glabrata. Novel miRNA were iden- tified from the B. glabrata genome (BgalB1; vectorbase.org) using SMIRP. Targets of novel snail miRNA were then identified using miRDeep2 from the available transcript assembly (BgalB1.5; vectorbase.org). Target genes of novel miRNA included multi-miRNA gene regu- lation of proteins involved in cellular processes such as secretory mucal proteins (mucin-21-like), matricellular proteins (thrombospondin- 3b-like), and shell formation (dentin sialophosphoprotein-like). number of miRNA in both a tissue and torpor stage-specific manner, clearly demonstrating that miRNA likely play an active role in mammalian hibernation and dynamic metabolic regulation. Although miRNA expression profiles were largely tissue-specific for the three organs studied (liver, heart, skeletal muscle), gene ontology (GO) annotation analysis revealed that the putative tar- gets of the upregulated miRNA were commonly enriched in cellular processes involved in the suppression of pro-growth. For example, in liver tissue, upregulated miRNA targeted genes enriched in cel- lular processes such as growth factor receptor signaling pathways, regulation of nuclear division, and glycolysis during the early tor- por stage (Wu et al., 2016). To further expand upon this analysis, we worked to elucidate the organization of the miRNA-targeted cellular system using the targets of the significantly upregulated miRNA from the ET stage of liver tissue. By mapping the collective targets of torpor- responsive miRNA with their known protein interactions, we were able to obtain an intuitive representation of miRNA-regulated Figure 2 Predicted functional interaction map of stress-responsive processes within a functional network. To accomplish this, we miRNA targets from the hibernating thirteen-lined ground squirrel. A used the spatial analysis of functional enrichment (SAFE) tool protein interaction map of all predicted targets of miRNA found to within the Cytoscape software (v3.4.0) (Figure 2). For each be significantly overexpressed during the early torpor (ET) stage of miRNA target within the network, SAFE defined the local neigh- ground squirrel hibernation. Network was constructed using known borhood of targets by identifying those that look to be clustered human protein interactions from the STRING database (https:// within the network. GO analysis was then carried to determine string-db.org/), contained 532 miRNA targets (i.e. nodes) and 1433 enriched miRNA-regulated cellular processes (Baryshnikova, known interactions (i.e. edges), and was originally constructed 2016). Similar to the original study, we identified an enrichment using Cytoscape (v3.4.0). Spatial analysis of functional enrichment (SAFE)-based construction of a functional map of the network by in targeting cellular signaling pathways, including growth factor combining all region-specific human GO terms into 12 functional signaling and carboxylic acid catabolism. We also identified a domains based on the similarity of their enrichment landscapes significant enrichment in the regulation of lipid biosynthesis, −4 (P values < 2 × 10 , Fisher’s exact test). Different colors represent hormone response signaling, and RNA processing (Figure 2). different functional domains. Each domain is labeled with a tag list, Lipid metabolism is known to be under tight regulatory control composed of the five words that occur most frequently within the during hibernation as triglycerides are the primary energy source names of the associated GO terms. for the hibernating mammal and are known to influence the Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 98 j Biggar and Storey length of torpor bouts and metabolic rate (Florant, 1998; Dark, and TIAR function during stress suggested that enhanced protein 2005; Wu et al., 2013). Since the entry into and arousal from aggregation was not present during torpor. Our identification of torpor is a short process that requires rapid changes in many miRNA-targeted regulation of RNA processing agrees with this study metabolic processes, this particular result is not surprising. as it identifies posttranscriptional regulatory mechanisms at play in Furthermore, with regards to the predicted enrichment of RNA reducing translational rates and/or mRNA processing. As demon- processing in torpid thirteen-lined ground squirrels, a previous strated by our deeper network-based functional analysis into torpor- study has characterized the roles of three RNA binding proteins: responsive miRNA function (Figure 2), a global analysis of miRNA T-cell intracellular antigen 1 (TIA-1), TIA-1 related (TIAR), and targets can help to provide deeper, meaningful, insight into the poly(A)-binding protein (PABP-1)(Tessier et al., 2014). TIA-1 role of miRNA in helping to coordinate the torpor–arousal cycle. was identified as a major component of sub-nuclear structures with up to a 7-fold increase in relative protein levels found in Temperature influence over microRNA–target interaction the nucleus during hibernation. Additionally, analysis of the Given the state of miRNA research within the field of com- formation of reversible aggregates that are associated with TIA-1 parative biochemistry and the functional insight that it has Figure 3 Temperature-associated regulation of miRNA function. (A) The average G/C content of complete miRNA sequences as well as their seed regions plotted against the physiological temperature of each organism. (B) For each of the organisms, miRNAs were divided into two subsets that contain (i) miRNAs that are specific to a taxonomic group and (ii) the rest miRNAs that are shared by other taxonomic groups. Each pair of bars shows the difference in G/C content between these two subsets for miRNAs (dark gray) and for seeds (light gray). Error bars show standard error of the difference between two means; (*) P < 0.05 and (**) P < 0.0005.(C) Depiction of proposed temperature influence in RNA binding thermodynamics. It is hypothesized that low temperature could act to stabilize miRNA:target interactions that were once unfavorable or unstable at higher temperatures. Data for A and B were derived from Carmel et al. (2012). Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 99 provided to date, it is becoming increasingly clear that these Overall, the increasing realization that T will likely have a small regulatory RNA are an important component of environ- dramatic impact on miRNA function, presents the possibility that mental stress survival and the hypometabolic response. Given distinct temperature-induced miRNA targeting programs may be this interest, there has also been an increasing attempt to at play and help to facilitate cellular function at various tem- explore the targets that these stress-responsive miRNA regulate peratures. In this way, there may exist distinct cold-influenced (Figure 3). In recent years, there has also been a growing inter- miRNA targeting programs that facilitate unique hypometabolic est in the possibility of temperature influencing the regulatory survival of extreme stresses. Given that the strict seed pairing function of miRNA (Figure 3). This has been previously dis- requirement of miRNA would still be required, it is possible that cussed both in terms of miRNA base-content and its relationship the relatively lower G/C content within the seed region of organ- to T (Figure 3A and B), as well as in the context of low-temperature isms living at lower temperatures may also have a role in aiding influence over miRNA:mRNA binding thermodynamics (Figure 3C) this temperature-sensitive process (Figure 3). Thus, it is interesting (Carmel et al., 2012; Biggar and Storey, 2014a, b, 2015a, b, 2017). to propose that miRNA could play a specific temperature-sensitive Given the relationship between miRNA seed binding potential role in helping various species tocopewithtemperature-related (i.e. G/C content) and temperature, the question as to whether stress. there are other mechanisms by which temperature can influence miRNA function should be asked. The targeting of miRNA to Conclusion select mRNA sites relies primarily on seed region complementar- Within the past decade, many studies have explored the role ity, with further binding from the 3′end of the miRNA only acting of miRNA as a mechanism to reversibly and rapidly regulate the to stabilize and supplement the interaction. Critically, it has cellular landscape and enable survival during periods of extreme been previously reported that the thermodynamic threshold stress. The overall impact of these studies has resulted in a (mean free energy; mfe) used to predict whether a miRNA:mRNA common theme of stress-responsive miRNA expression across target will occur is ∼18 kcal/mol. This threshold, among other phylogeny. Building upon this initial research, and with the structural requirements of the miRNA:mRNA interaction, has growing availability of genomic information and bioinformatic been used in almost all target prediction programs that have power, researchers have now begun to predict the overall cellu- been developed for the identification of human miRNA targets lar impact of stress-responsive miRNA expression in the context in mind (including miRanda, TargetScan, and Diana microT). of global target regulation. As a result, it is clear that miRNA These miRNA target identification programs typically overlook may be involved in the regulation of many cellular processes the possibility of non-human species existing at T values great- that enable efficient utilization of ATP turnover. As we learn er or lower than 37°C. Indeed, given the strong thermodynamic more about miRNA function in different organisms and in requirement for a successful miRNA:mRNA interaction, it is likely response to various environmental conditions, the possibility of that a significant change in T (such as experienced by frozen temperature having a role to play in influencing target selection frogs and turtles, hibernating mammals, and many other over- may also open comparative miRNA research to an immense wintering animals) will have a strong influence on the ability of number of regulatory possibilities that will need to be explored. miRNA target selection. In this way, a decrease in T would likely favorably stabilize miRNA–target interactions that were Acknowledgements once unfavorable, allowing these interactions to become bio- Thanks go to J. Storey (Institute of Biochemistry, Carleton logically relevant in a temperature-dependent manner. University, Canada) for editorial review of the manuscript. Indeed, one study using the FindTar3 miRNA target prediction software, which allows user control over temperature, showed a 16-fold increase in the number of potential mRNA targets when Funding comparing those predicted at freezing temperatures (3°C) and This research was funded by a Discovery grant from the those identified at 37°C(Biggar and Storey, 2015a, b). For Natural Sciences and Engineering Research Council of Canada example, in response to freezing (3°C) in the hatchling painted (NSERC; grant no. 6793) to K.B.S. turtle (Chrysemys picta marginata), miR-21 expression levels Conflict of interest: none declared. were shown to increase by over 2-fold in heart tissue (Biggar and Storey, 2015a, b). When predicting the mRNA targets of miR-21, this study compared the targets predicted at both 37°C References and 3°C, finding an increase from 47 to 756 targets at the lower Adema, C.M., Hillier, L.W., Jones, C.S., et al. (2017). Whole genome analysis of a schistosomiasis-transmitting freshwater snail. Nat. Commun. 8, 15451. temperature. Interestingly, these low-temperature targets were Bansal, S., Luu, B.E., and Storey, K.B. (2016). MicroRNA regulation in heart enriched in metabolic processes (Biggar and Storey, 2015a, b, and skeletal muscle over the freeze-thaw cycle in the freeze tolerant wood 2017; Luu et al., 2016). Within the context of a global decrease frog. J. Comp. Physiol. B 186, 229–241. in gene expression, it is likely that such an increase of miRNA Bartel, D. (2004). MicroRNAs: genomics, biogenesis, mechanism, and func- targets would dramatically alter the miRNA targeting program. tion. Cell 116, 281–297. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 100 j Biggar and Storey Baryshnikova, A. (2016). Systematic functional annotation and visualization Cowan, K.J., and Storey, K.B. (2003). Mitogen-activated protein kinases: new of biological networks. Cell Syst. 2, 412–421. signaling pathways functioning in cellular responses to environmental Biggar, K.K., Dubuc, A., and Storey, K.B. (2009). MicroRNA regulation below stress. J. Exp. Biol. 206, 1107–1115. zero: differential expression of miRNA-21 and miRNA-16 during freezing in Dark, J. (2005). Annual lipid cycles in hibernators: integration of physiology wood frogs. Cryobiology 59, 317–321. and behavior. Annu. Rev. Nutr. 25, 469–497. Biggar, K.K., Kornfeld, S.F., and Storey, K.B. (2011). Amplification and Epperson, L.E., and Martin, S.L. (2002). Quantitative assessment of ground sequencing of mature microRNAs in uncharacterized animal models using squirrel mRNA levels in multiple stages of hibernation. Physiol. Genomics stem-loop reverse transcription-polymerase chain reaction. Anal. Biochem. 10, 93–102. 416, 231–233. Fraser, K.P., Houlihan, D.F., Lutz, P.L., et al. (2001). Complete suppression of Biggar, K.K., Kornfeld, S.F., Maistrovski, Y., et al. (2012). MicroRNA regulation protein synthesis during anoxia with no post-anoxia protein synthesis in extreme environments: differential expression of microRNAs in the inter- debt in the red-eared slider turtle, Trachemys scripta elegans. J. Exp. Biol. tidal snail Littorina littorea during extended periods of freezing and 204, 4353–4360. anoxia. Genomics Proteomics Bioinformatics 10, 302–309. Frerichs, K.U., Smith, C.B., Brenner, M., et al. (1998). Suppression of protein Biggar, K.K., and Storey, K.B. (2012). Evidence for cell cycle suppression and synthesis in brain during hibernation involves inhibition of protein initi- microRNA regulation of cyclin D1 during anoxia exposure in turtles. Cell ation and elongation. Proc. Natl Acad. Sci. USA 95, 14511–14516. Cycle 11, 1705–1713. Florant, G.L. (1998). Lipid metabolism in hibernators: the importance of Biggar, K.K., and Storey, K.B. (2011). The emerging roles of microRNAs in the essential fatty acids. Integr. Comp. Biol. 38, 331–340. molecular responses of metabolic rate depression. J. Mol. Cell Biol. 3, Frigault, J.J., Lang-Ouellette, D., and Morin, P.J. (2016). Up-regulation of non- 167–175. coding RNA Tug1 in hibernating thirteen-lined ground squirrels, Ictidomys Biggar, K.K., and Storey, K.B. (2014a). Identification and expression of tridecemlineatus. Genomics Proteomics Bioinformatics 14, 113–118. microRNA in the brain of hibernating bats, Myotis lucifugus. Gene 544, Frigault, J.J., Morin, M.D., and Morin, P.J. (2017). Differential expression and 67–74. emerging functions of non-coding RNAs in cold adaptation. J. Comp. Biggar, K.K., and Storey, K.B. (2014b). Insight into temperature-dependent Physiol. B 187, 19–28. microRNA function in mammalian hibernators: perspectives on cold- Friedlander, M.R., Mackowiak, S.D., Li, N., et al. (2012). MiRDeep2 accurately influenced microRNA/target interaction. Temperature 1, 84–86. identifies known and hundreds of novel microRNA genes in seven animal Biggar, K.K., and Storey, K.B. (2015a). Insight into post-transcriptional gene clades. Nucleic Acids Res. 40, 37–52. regulation: stress-responsive microRNAs and their role in the environmen- Friedman, R.C., Farh, K.K.H., Burge, C.B., et al. (2009). Most mammalian tal stress survival of tolerant animals. J. Exp. Biol. 218, 1281–1289. mRNAs are conserved targets of microRNAs. Genome Res. 19, 92–105. Biggar, K.K., and Storey, K.B. (2015b). Low-temperature microRNA expres- Gabrial, U.I., Mirela, S., and Ionel, J. (2011). Quantification of mucoproteins sion in the painted turtle, Chrysemys picta during freezing stress. FEBS (glycoproteins) from snail mucus, Helix aspersa and Helix pomatia. J. Agro. Lett. 589, 3665–3670. Proc. Tech. 17, 410–413. Biggar, K.K., and Storey, K.B. (2017). Exploration of low temperature Guppy, M., and Withers, P. (1999). Metabolic depression in animals: physio- microRNA function in an anoxia tolerant vertebrate ectotherm, the red logical perspectives and biochemical generalizations. Biol. Rev. Camb. eared slider turtle (Trachemys scripta elegans). J. Therm. Biol. 68, Philos. Soc. 74, 1–40. 139–146. Hadj-Moussa, H., Moggridge, J.A., Luu, B.E., et al. (2016). The hibernating Biggar, K.K., Wu, C.W., and Storey, K.B. (2014). High-throughput amplifica- South American marsupial, Dromiciops gliroides, displays torpor-sensitive tion of mature microRNAs in uncharacterized animal models using polya- microRNA expression patterns. Sci. Rep. 6, 24627. denylated RNA and stem-loop reverse transcription polymerase chain Hittel, D., and Storey, K.B. (2002). The translation state of differentially reaction. Anal. Biochem. 462, 32–34. expressed mRNAs in the hibernating 13-lined ground squirrel Britton, C., Winter, A.D., Gillan, V., et al. (2014). MicroRNAs of parasitic hel- (Spermophilus tridecemlineatus). Arch. Biochem. Biophys. 401, 244–254. minths – identification, characterization and potential as drug targets. Int. Hochachka, P.W., Buck, L.T., Doll, C.J., et al. (1996). Unifying theory of hyp- J. Parasitol. 4, 85–94. oxia tolerance: molecular/metabolic defense and rescue mechanisms for Brooks, S.P.J., and Storey, K.B. (1993). De novo protein synthesis and protein surviving oxygen lack. Proc. Natl Acad. Sci. USA 93, 9493–9498. phosphorylation during anoxia and recovery in the red-eared turtle. Am. J. John, B., Enright, A.J., Aravin, A., et al. (2005). MiRanda application: Human Physiol. 265,R1380–R1386. microRNA targets. PLoS Biol. 3,e264. Brooks, S.P.J., and Storey, K.B. (1995). Protein phosphorylation patterns dur- Kornfeld, S.F., Biggar, K.K., and Storey, K.B. (2012). Differential expression of ing aestivation in the land snail Otala lactea. Mol. Cell. Biochem. 143, mature microRNAs involved in muscle maintenance of hibernating little 7–13. brown bats, Myotis lucifugus: a model of muscle atrophy resistance. Carmel, I., Shomron, N., and Heifetz, Y. (2012). Does base-pairing strength Genomics Proteomics Bioinformatics 10, 295–301. play a role in microRNA repression? RNA 18, 1947–1956. Krol, J., Loedige, I., and Filipowicz, W. (2010). The widespread regulation of Chen, M., Zhang, X., Liu, J., et al. (2013). High-throughput sequencing reveals microRNA biogenesis, function and decay. Nat. Rev. Genet. 11, 597–610. differential expression of miRNAs in intestine from sea cucumber during Land, S.C., Buck, L.T., and Hochachka, P.W. (1993). Response of protein syn- aestivation. PLoS One 8,e76120. thesis to anoxia and recovery in anoxia-tolerant hepatocytes. Am. J. Chen, M., and Storey, K.B. (2014). Large-scale identification and comparative Physiol. 265,R41–R48. analysis of miRNA expression profile in the respiratory tree of the sea Lang-Ouellette, D., and Morin, P.J. (2014). Differential expression of miRNAs cucumber Apostichopus japonicus. Mar. Genomics 13, 39–44. with metabolic implications in hibernating thirteen-lined ground squirrels, Cipolla, G.A. (2014). A non-canonical landscape of the microRNA system. Ictidomys tridecemlineatus. Mol. Cell. Biochem. 394, 291–298. Front. Genet. 5, 337. Larade, K., and Storey, K.B. (2002). Reversible suppression of protein synthe- Courteau, L.A., Storey, K.B., and Morin, P.J. (2012). Differential expression of sis in concert with polysome disaggregation during anoxia exposure in microRNA species in a freeze tolerant insect, Eurosta solidaginis. Littorina littorea. Mol. Cell. Biochem. 232, 121–127. Cryobiology 65, 210–214. Lee, Y.J., Johnson, K.R., and Hallenbeck, J.M. (2012). Global protein con- Cowan, K.J., MacDonald, J.A., Storey, J.M., et al. (2000). Metabolic reorgan- jugation by ubiquitin-like-modifiers during ischemic stress is regulated ization and signal transduction during estivation in the spadefoot toad. by microRNAs and confers robust tolerance to ischemia. PLoS One 7, Exp. Biol. Online 5, 1–25. e47787. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 101 Liu, Y., Hu, W., Wang, H., et al. (2010). Genomic analysis of miRNAs in an Shaffer, H.B., Minx, P., Warren, D.E., et al. (2013). The western painted turtle extreme mammalian hibernator, the Arctic ground squirrel. Physiol. genome, a model for the evolution of extreme physiological adaptations in Genomics 42A, 39–51. a slowly evolving lineage. Genome Biol. 14,R28. Luu, B.E., Biggar, K.K., Wu, C.W., et al. (2016). Torpor-responsive expression Storey, K.B. (2015). Regulation of hypometabolism: insights into epigenetics of novel microRNA regulating metabolism and other cellular pathways in controls. J. Exp. Biol. 218, 150–159. the thirteen-lined ground squirrel, Ictidomys tridecemlineatus. FEBS Lett. Storey, K.B., and Storey, J.M. (2004). Metabolic depression in animals: tran- 590, 3574–3582. scriptional and translational controls. Biol. Rev. 79, 207–233. Luu, B.E., and Storey, K.B. (2015). Dehydration triggers differential microRNA Tessier, S.N., Audas, T.E., and Wu, C.W. (2014). The involvement of mRNA expression in Xenopus laevis brain. Gene 573, 64–69. processing factors TIA-1, TIAR, and PABP-1 during mammalian hibernation. Lyons, P.J., Crapoulet, N., Storey, K.B., et al. (2015a). Identification and pro- Cell Stress Chaperones 19, 813–825. filing of miRNAs in the freeze-avoiding gall moth Epiblema scudderiana via Volk, T., Wang, S., Rotstein, B., et al. (2014). Matricellular proteins in devel- next-generation sequencing. Mol. Cell. Biochem. 410, 155–163. opment: perspectives from the Drosophila heart. Matrix. Biol. 37, Lyons, P.J., Govaere, L., Crapoulet, N., et al. (2016). Characterization of cold- 162–166. associated microRNAs in the freeze-tolerant gall fly Eurosta solidaginis Wu, C., So, J., Davis-Dusenbery, B.N., et al. (2011). Hypoxia potentiates using high-throughput sequencing. Comp. Biochem. Physiol. Part D microRNA-mediated gene silencing through posttranslational modification Genomics Proteomics 20, 95–100. of argonaute 2. Mol. Cell. Biol. 31, 4760–4774. Lyons, P.J., Poitras, J.J., Courteau, L.A., et al. (2013). Identification of differen- Wu, C.W., Biggar, K.K., and Storey, K.B. (2013). Biochemical adaptations of tially regulated microRNAs in cold-hardy insects. Cryo. Lett. 34, 83–89. mammalian hibernation: exploring squirrels as a perspective model for Lyons, P.J., Storey, K.B., and Morin, P.J. (2015b). Expression of miRNAs in naturally induced reversible insulin resistance. Braz. J. Med. Biol. Res. 46, response to freezing and anoxia stresses in the freeze tolerant fly Eurosta 1–13. solidaginis. Cryobiology 71, 97–102. Wu, C.W., Biggar, K.K., and Storey, K.B. (2014a). Expression profiling and Maistrovski, Y., Biggar, K.K., and Storey, K.B. (2012). HIF-1α regulation in structural characterization of microRNAs in adipose tissues of hibernating mammalian hibernators: role of non-coding RNA in HIF-1α control during ground squirrels. Genomics Proteomics Bioinformatics 12, 284–291. torpor in ground squirrels and bats. J. Comp. Physiol. B 182, 849–859. Wu, C.W., Biggar, K.K., and Storey, K.B. (2014b). Dehydration mediated Marxen, J.C., and Becker, W. (2011). Calcium binding constituents of the microRNA response in the African clawed frog, Xenopus laevis. Gene 529, organic shell matrix from the freshwater snail Biomphalaria glabrata. 3376–3383. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 127, 235–242. Wu, C.W., Biggar, K.K., Luu, B.E., et al. (2016). Analysis of microRNA expres- Morin, P.J., Dubuc, A., and Storey, K.B. (2007). Differential expression of sion during the torpor-arousal cycle of a mammalian hibernator, the 13- microRNA species in organs of hibernating ground squirrels: a role in transla- lined ground squirrel. Physiol. Genomics 48, 388–396. tional suppression during torpor. BBA Gene Regul. Mech. 1779, 628–633. Wu, C.W., and Storey, K.B. (2013). Regulation of the mTOR signaling network Peace, R.J., Biggar, K.K., Storey, K.B., et al. (2015). A framework for improving in hibernating thirteen-lined ground squirrels. J. Exp. Biol. 215, microRNA prediction in non-human genomes. Nucleic Acids Res. 43,e138. 1720–1727. Podrabsky, J.E., and Hand, S.C. (2000). Depression of protein synthesis dur- Yamasaki, S., and Anderson, P. (2008). Reprogramming mRNA translation ing diapause in embryos of the annual killifish, Austrofundulus limnaeus. during stress. Curr. Opin. Cell Biol. 20, 222–226. Physiol. Biochem. Zool. 73, 799–808. Yi, R., Qin, Y., Macara, I.G., et al. (2003). Exportin-5 mediates the nuclear Pothof, J., Verkaik, N.S., Hoeijmakers, J.H., et al. (2009). MicroRNA responses export of pre-microRNAs and short hairpin RNAs. Genes Dev. 17, and stress granule formation modulate the DNA damage response. Cell 3011–3016. Cycle 8, 3462–3468. Zhang, J., Biggar, K.K., and Storey, K.B. (2013). Regulation of p53 by revers- Rouble, A.N., Biggar, K.K., and Storey, K.B. (2014). A high-throughput proto- ible post-transcriptional and post-translational mechanisms in liver and col for message RNA quantification using RNA dot-blots. Anal. Biochem. skeletal muscle of an anoxia tolerant turtle, Trachemys scripta elegans. 452, 31–33. Gene 513, 147–155. Schaap, P., Barrantes, I., Minx, P., et al. (2016). The Physarum polycephalum Zeng, Y., Yi, R., and Cullen, B.R. (2005). Recognition and cleavage of primary genome reveals extensive use of prokaryotic two-component and microRNA precursors by the nuclear processing enzyme Drosha. EMBO J. metazoan-type tyrosine kinase signaling. Genome Biol. Evol. 8, 109–125. 24, 138–148. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal of Molecular Cell Biology Oxford University Press

Functional impact of microRNA regulation in models of extreme stress adaptation

Biggar, Kyle K; Storey, Kenneth B; Wang, Zefeng
Journal of Molecular Cell Biology , Volume Advance Article (2) – Jan 9, 2018
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Abstract

doi:10.1093/jmcb/mjx053 Journal of Molecular Cell Biology (2018), 10(2), 93–101 j 93 Published online January 9, 2018 Review Functional impact of microRNA regulation in models of extreme stress adaptation Kyle K. Biggar and Kenneth B. Storey Institute of Biochemistry & Department of Biology, Carleton University, Ottawa, ON K1S 5B6, Canada * Correspondence to: Kenneth B. Storey, E-mail: kenneth_storey@carleton.ca Edited by Zefeng Wang When confronted with severe environmental stress, some animals are able to undergo a substantial reorganization of their cellular environment that enables long-term survival. One molecular mechanism of adaptation that has received considerable attention in recent years has been the action of reversible transcriptome regulation by microRNA. The implementation of new computational and high-throughput experimental approaches has started to uncover the vital contributions of microRNA towards stress adaptation. Indeed, recent studies have suggested that microRNA may have a major regulatory influence over a number of cellular processes that are essential to prolonged environmental stress survival. To date, a number of studies have highlighted the role of microRNA in the regulation of a metabolically depressed state, documenting stress-responsive microRNA expression during mammalian hibernation, frog and insect freeze tolerance, and turtle and marine snail anoxia tolerance. These studies collectively indicate a conserved principle of microRNA stress response across phylogeny. As we are on the verge of dissecting the role of microRNA in environmental stress adaptation, this review summarizes recent research advances and the hallmark expression patterns that facilitate stress survival. Keywords: hypometabolism, metabolism, temperature, metabolic rate depression, stress response Introduction readily reversible, and capable of eliciting selective control over A select few animals have developed an incredible ability to both essential and non-essential cellular processes. This review survive and overcome severe environmental challenges that will discuss new advances in the roles of non-coding RNA in pro- include dehydration, oxygen deprivation (i.e. anoxia), as well as moting animal survival in extreme environments, as well as the temperature changes and even freezing of body fluids (Storey transition into (and survival within) the hypometabolic state. The and Storey, 2004). These fascinating feats and the study of how function of small non-coding RNA, namely microRNA (miRNA), will these resilient animals are able to survive such extreme stresses be a primary focus given the extensive amount of miRNA research has captivated researchers for many years. In particular, the past that has taken place in this field over the past 10 years (Morin few decades research in comparative stress biology has focused et al., 2007; Biggar and Storey, 2015a, b; Frigault et al., 2017). on elucidating the molecular survival mechanisms at play, those Mechanisms that help to coordinate stress survival must be easily working to reorganize the cellular landscape for life in a new inducible and reversible so that they can be initiated once a stress is environmental extreme (Storey and Storey, 2004; Storey, 2015). detected. The contribution of readily reversible protein modifica- These mechanisms work to reprioritize cellular processes, such tions, such as Ser/Thr/Tyr phosphorylation, has been extensively that these animals can conserve vital energy stores, with some studied as a dynamic mechanism to quicklymodifyprotein function entering a state of dormancy that is characterized by the depres- and achieve hypometabolism. These studies established that the sion of metabolic rate to 10%–30% of basal levels (or even basic principles involved in regulating stress-responsive signal trans- more), a state commonly referred to as hypometabolism (Guppy duction events included the coordinated reversible phosphorylation and Withers, 1999). Given the relatively rapid onset of many of many proteins, setting this modification as one of the hallmarks of environmental stresses, molecular mechanisms that help to gov- the central mechanisms of metabolic reorganization for many years ern the transition into a hypometabolic state must be rapid, (Cowan and Storey, 2003). Indeed, reversible post-translational modifications have been shown to affect the function of numerous cellular proteins and to serve as a common response element in Received August 13, 2017. Revised November 20, 2017. Accepted November 30, the survival of many environmental challenges and metabolic stresses (Brooks and Storey, 1995; Cowan et al., 2000; Biggar © The Author (2018). Published by Oxford University Press on behalf of Journal of and Storey, 2012; Wu and Storey, 2013). Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 94 j Biggar and Storey Mechanisms of reversible control over protein translation mRNA content, but rather achieved through post-translational Beginning in the early 1990s, it was established that an over- modifications made to translational machinery. all suppression of the energetically costly cellular process of Despite a possible global shutdown of protein translation protein translation was an essential part of hypometabolism pathways, there still remains the question of how the proteins (Brooks and Storey, 1993; Land et al., 1993; Frerichs et al., that are essential for survival continue to be created in the 1998; Fraser et al., 2001; Larade and Storey, 2002; Hittel and hypometabolic state. There must be mechanisms at play that Storey, 2002). Indeed, a depression of translational rate neces- allow for essential mRNA to be translated, while non-essential sarily contributes to energy rationing, thereby helping to reserve mRNA are blocked from translation. This requirement highlights ATP turnover for essential survival-related cellular processes. the regulatory influence of miRNA in the translational control of For example, the rate of protein translation in oxygen-deprived specific mRNA. Perhaps such regulatory mechanisms would turtles was shown to decrease below measurable levels follow- allow gene-specific control over protein translation during peri- ing 3 h of anoxic exposure (at 23°C) as compared with normoxic ods of cell stress. Indeed, this area of research has gained con- values at the same temperature (Fraser et al., 2001). Similarly, siderable interest in recent years within the field of comparative in the brain of torpid thirteen-lined ground squirrels, the rate of biochemistry. protein translation was documented to be only 0.04% of euther- mic rates (Frerichs et al., 1998) and, when corrected for differ- Reversible control of protein translation by microRNA ences in body temperature between euthermic and torpid state, MiRNA are short, non-coding RNA capable of rapidly and rates in torpor were still approximately only one-third of the reversibly regulating the expression of targeted proteins within euthermic values. Together, these results demonstrated that a cell and have been found in plants, animals, bacteria, as well temperature influences alone do not explain reductions in trans- as some viruses. It is well documented that these 18–24 nt tran- lational rates and that the depression of protein translation scripts are able to bind, with full or partial complementarity, to likely involves other molecular mechanisms. Indeed, the sup- specific mRNA targets, resulting in either the inhibition of trans- pression of protein translation in the hypometabolic state is a lation or degradation of the target mRNA (Bartel, 2004). As a multifaceted event, one that is likely achieved through a com- reflection of their regulatory potential, miRNA have been pre- bination of different mechanisms that include reductions in dicted to be involved in all biological process in some aspect gene transcription, inactivation of the translational machinery, throughout the animal kingdom (Friedman et al., 2009). and/or through mRNA-specific interference by miRNA. The biogenesis of a mature miRNA begins with the initial tran- As a result of the high energetic costs of protein synthesis scription of a primary miRNA (pri-miRNA). The pri-miRNA tran- and mRNA transcription (accounting for greater than 25%of script is much larger than the final mature miRNA product, basal ATP turnover in many instances), it would not be surpris- ranging from hundreds to thousands of nucleotides in length ing to see that a strong stress-responsive suppression of mRNA (Krol et al., 2010). Following transcription, the pri-miRNA is then transcription would accompany the observed stress-responsive cleaved to form a hairpin RNA structure called a precursor decrease in translational rate (Hochachka et al., 1996; Podrabsky miRNA (pre-miRNA) and is exported from the nucleus (Yi et al., and Hand, 2000). In this regard, previously studies have shown 2003; Zeng et al., 2005; Krol et al., 2010). Once in the cyto- that reversible control over translational machinery may be a plasm, the pre-miRNA is further processed into a mature miRNA powerful driving force behind global translational silencing during duplex by Dicer, an riboendonuclease protein. From the mature hypometabolism. For example, research on protein translation in miRNA duplex, it is suggested that only one strand will function hibernating thirteen-lined ground squirrels has documented a as a mature miRNA, leaving the passenger strand to be dynamic stress-responsive increase in the relative phosphorylation degraded (Krol et al., 2010). Although this is generally thought of the translational initiation factor eIF2α (Hittel and Storey, to be the case, the idea of passenger miRNA degradation has 2002). The phosphorylation of eIF2α is a well-established marker been challenged in light of recent research suggesting that the of both translational arrest and the disruption of active polyribo- abundance, possible function, and physiological relevance of somes (Yamasaki and Anderson, 2008). Indeed, the same study of passenger miRNA has been underestimated (Cipolla, 2014). hibernating ground squirrels also demonstrated a hibernation- Following mature miRNA processing, the mature miRNA is then responsive disaggregation of polyribosomes (Hittel and Storey, loaded into a protein complex referred to as the RNA-induced 2002). Both of these results indicate that a translationally silent silencing complex (RISC). Interestingly, the miRNA–RISC and state is imposed by a stress-responsive inactivation of the transla- bound mRNA complex have also been found to aggregate within tional machinery. Furthermore, neither the total cellular polyade- stress granules and p-bodies in a stress-responsive manner, nylated RNA content nor the mRNA transcript levels of many such as in response to DNA damage or hypoxia stresses, and genes were found to decrease in hibernation or anoxia models of are eventually either degraded or directed into translation hypometabolism (Epperson and Martin, 2002; Storey and Storey, once the stress is lifted (Pothof et al., 2009; Wu et al., 2011). 2004; Rouble et al., 2014). Therefore, it is reasonable to conclude Indeed, the stress-responsive formation of reversible stress that the suppression of global protein translation rates during granules have been shown to occur in response to mammalian hypometabolism may not be a direct result of altered cellular hibernation (Tessier et al., 2014). Take together, this reversible Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 95 mRNA storage system represents a promising mechanism for as part of winter survival by freeze-tolerant (Littorina littorea, reversible translational arrest in models of hypometabolism. Eurosta solidaginis) and freeze-avoidant (Epiblema scudderiana) species (Biggar et al., 2012; Courteau et al., 2012; Lyons et al., MicroRNA discovery in non-model organisms 2013, 2015a, b, 2016). In general, miRNA and their target recognition sites have Given the inherent difficulty of studying poorly conserved been subject to exquisite conservation across phylogeny. The miRNA and an interest in discovering possible specifies-specific conservation of miRNA by >90% among vertebrates is reflective miRNA, computation-based methods have recently been devel- of the important regulatory function they impose in normal cell oped specifically to identify (i) highly conserved, (ii) poorly con- biology. This high level of sequence homology between species served, and (iii) species-specific miRNA from organisms with has made it possible to study miRNA expression patterns in newly sequenced genomes. The widely used miRDeep2 platform many different species with relative ease. Indeed, multiple stud- accomplishes this by using both an available genome sequence ies have been able to explore the dynamic expression of miRNA and small RNA sequencing information to map mature miRNA in response to a variety of environmental stresses in a diverse sequences back to a reference genomic location and to then array of animals for which little or no genomic sequence infor- evaluate the associated pre-miRNA structure and identify high- mation is available. To date, stress-responsive miRNA expres- confidence miRNA (Friedlander et al., 2012). Given the import- sion has been studied in models of hypometabolism that ance miRNA sequence on its function, it is essential to validate include ground squirrels, bats, frogs, turtles, and many inverte- all computationally predicted candidate miRNAs experimentally, brate species (Table 1 with references). Indeed the dynamic including PCR-based methods, such that their expression and change in stress-responsive miRNA expression may be the result sequence of their mature forms can be verified. Recently, small of multiple regulatory steps, including transcriptional modula- RNA sequencing and miRDeep2 have been used to identify tion, microprocessor or miRNA biogenesis co-factor regulation, novel miRNA regulating gene expression during hibernation in changes in RNA/protein complex localization, and modifications thirteen-lined ground squirrels, Ictidomys tridecemlineatus (Luu of miRNA ends. Although the exact mechanisms controlling et al., 2016). The study identified and experimentally validated stress-induced miRNA expression are not yet known, some of 17 novel miRNA and characterized their relative expression in these steps may play pivotal roles in cellular homeostasis in the liver, skeletal muscle, and heart tissues over the torpor–arousal context of the stress response. cycle. Interestingly, these squirrel-specific miRNA were predicted Much of the initial hypometabolism-focused miRNA research to target mRNA enriched in biological processes that are known was carried out on vertebrate and invertebrate species with lim- to be dynamically regulated during hibernation, including lipid ited genomic sequence information available (Morin et al., metabolism, ion-transport ATPases, and various cellular signal- 2007; Biggar et al., 2009, 2012; Courteau et al., 2012; Lyons ing cascades (Luu et al., 2016). This study provides an example et al., 2013). Given this restriction, these studies were limited to of how computational identification of new miRNA and analysis the analysis of highly conserved miRNA using methods that had of their cellular function is of growing interest in comparative been developed to aid in the amplification, sequencing, and val- research on environmental stress adaptation. Indeed, such idation of these conserved miRNA (Biggar et al., 2011, 2014). knowledge may be crucial to developing a deeper understand- However, while vertebrate miRNA are highly conserved, low con- ing of the roles that miRNA may play in the hypometabolic servation between vertebrate and invertebrate miRNA slowed stress response. The major restriction of this particular platform the initial progress of miRNA research in invertebrate species. is the requirement for small RNA sequencing information. In the Despite this challenge, several studies have explored the regula- absence of this information, another platform, called SMIRP, tion of various relatively conserved invertebrate miRNA, particularly has been developed recently to identify species-specific miRNA Table 1 Studies exploring the role of stress-responsive miRNA regulation in animal models of hypometabolism. Stress Species References Hibernation Ictidomys tridecemlineatus Morin et al. (2007), Maistrovski et al. (2012), Lang-Ouellette and Morin (2014), Wu et al. (2014a), Luu et al. (2016), Wu et al. (2016), Frigault et al. (2016) Spermophilus parryii Liu et al. (2010) Myotis lucifigus Biggar and Storey (2014a, b), Kornfeld et al. (2012), Maistrovski et al. (2012) Dromiciops gliroides Hadj-Moussa et al. (2016) Freezing Rana sylvatica Biggar et al. (2009), Bansal et al. (2016) Littorina littorea Biggar et al. (2012) Eurosta solidaginis Courteau et al. (2012), Lyons et al. (2013), Lyons et al. (2015a, b), Lyons et al. (2016) Chrysemys picta Shaffer et al. (2013), Biggar and Storey (2015a, b) Freeze avoidance Epiblema scudderiana Lyons et al. (2015a, b) Oxygen restriction Littorina littorea Biggar et al. (2012) Trachemys scripta elegans Biggar and Storey (2011), Zhang et al. (2013) Ictidomys tridecemlineatus Lee et al. (2012) Aestivation Xenopus laevis Wu et al. (2013), Luu and Storey (2015) Apostichopus japonicus Chen et al. (2013), Chen and Storey (2014) Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 96 j Biggar and Storey de novo directly from an available genome (Peace et al., 2015). ground squirrels, documenting the torpor-responsive expression The SMIRP platform identifies both known and novel species- of 117 miRNA across four stages of the torpor–arousal cycle specific miRNA by scanning the genome for pre-miRNA-like (Wu et al., 2016). A follow-up study identified 17 additional novel hairpins and evaluating select features of these hairpins (i.e. torpor-responsive miRNA, currently thought to be unique to the % nucleotide content, RNA folding measures, and other topo- thirteen-lined ground squirrel (Luu et al., 2016). Another study uti- logical descriptors) against miRNA that have been previously lized advances in parallel RNA sequencing to study the expression identified from related species (Peace et al., 2015; Schaap of >200 ground squirrel miRNA, including 18 novel miRNA, in the et al., 2016; Adema et al., 2017). liver of the hibernating Arctic ground squirrel, Spermophilus par- Recently, the SMIRP platform was used to identify species- ryii (Liu et al., 2010). To date, the regulatory influence of miRNA specific miRNA from the schistosomiasis-transmitting freshwater expression has been most extensively explored in the context of Ramshorn snail (Biomphalara glabrata)(Adema et al., 2017). mammalian hibernation (Table 1). These studies, and those of a Identification of these miRNA provided information on miRNA variety of other animals, have demonstrated the importance of evolution, conservation, and suggested influence on transla- studying these small regulatory molecules as a conserved dynamic tional regulation that may lead to possible mechanisms for response to environmental stress and adaptation across phyl- population control in this snail species. Researchers from this ogeny. Given the growing implication of miRNA in the regulation of study performed an in silico prediction of miRNA using SMIRP environmental stress tolerance, studies have now begun to dig within the B. glabrata genome scaffolds. Uniquely, SMIRP lever- deeper into miRNA function, asking specific research questions, aged a novel approach to miRNA classifier construction in which such as what miRNA are stress responsive? what mRNA are stress- models are built dynamically and are targeted toward B. glabra- responsive miRNA targeting? what are the global functional impli- ta. This differs from other methods (e.g. precursor miRNA pre- cations of miRNA? diction methods) that attempt to produce generalized models that are applicable to a large number of species. From this MicroRNA-influenced control over cellular pathways and approach, 202 pre-miRNA (95 known and 107 novels) and asso- processes ciated mature miRNA from B. glabrata were identified (Adema It is well established that a single miRNA sequence can exert et al., 2017). No homologous sequences to the identified novel regulatory effects on numerous different targets. Similarly, a single B. glabrata precursor miRNA were found in either the sea slug mRNA is likely the regulatory target of multiple miRNA, with differ- (Aplysia californica) or the sea snail (Lottia gigantea) annotated ent miRNA binding at varied locations within the 3′UTR. This com- miRNA, or present within their available genomes. Interestingly, plex regulatory system creates a model of enormous regulatory this study also identified the possible biological context of novel potential that cannot be ignored when studying miRNA function. B. glabrata miRNA, predicting mRNA targets from the 3′UTR of However, given the limited genomic resources for many experi- available B. glabrata transcripts (John et al., 2005). A significant mental models of hypometabolism, initial explorations into miRNA proportion of the identified target genes of these novel miRNA regulatory potential have been appropriately limited in scope. included multi-miRNA gene regulation of proteins involved in cel- These studies primarily characterized miRNA and target expression lular processes such as secretory mucal proteins (mucin-21-like) patterns in asinglemiRNA,singlemRNAtargetformat (Morin (Gabrial et al., 2011), matricellular proteins (thrombospondin-3b- et al., 2007; Biggar et al, 2009, 2012). One good example from like) (Marxen and Becker, 2011) and shell formation proteins (den- early miRNA research on models of hypometabolism is the stress- tin sialophosphoprotein-like) (Volk et al., 2014)(Figure 1). These responsive characterization of miR-21. This miRNA has been stud- newly identified miRNA greatly enrich the repertoire of known mol- ied in multiple animals in the context of regulating anti-apoptotic lusk miRNA, providing insights into mollusk miRNA function, as well genes in response to environmental stress (Morin et al., 2007; as their evolution and biogenesis. Such species-specific miRNA can Biggar et al., 2009; Biggar and Storey, 2011, 2012; Wu et al., also provide possible biocontrol targets for B. glabrata population 2014b). Following the development of bioinformatics methods that control, or may even play a role in the control of aestivation in this allowed for the widespread study of multiple miRNA from a single species (Britton et al., 2014). set of RNA samples (Biggar et al., 2014), recent studies began to move away from single miRNA:target-based candidate characteriza- MicroRNA and adaptation to extreme environments tion. The scope of current studies is growing larger as we begin to A growing number of studies have begun to highlight the con- learn more about miRNA target selection and begin to apply new served response of miRNA in the regulation of the hypometabolic computational-based methods to explore the regulatory impact of a state. Beginning in 2007, the first study of this type explored the greater set of stress-responsive miRNA on the complete system of expression of several well-conserved miRNA in response to sea- cellular processes (Luu et al., 2016; Wu et al., 2016). sonal hibernation in the thirteen-lined ground squirrel (Morin In 2016, an expansive study looking at the torpor-responsive et al., 2007). Although only exploring the expression of a handful expression of 117 conserved miRNA in hibernating thirteen-lined of miRNA, this was a keystone paper in introducing the possible ground squirrels over four stages of the torpor–arousal cycle influence of miRNA as a mechanism to reversibly control mRNA (euthermia, early torpor, late torpor, and interbout arousal) (Wu translation in models of hypometabolism. More recently, several et al., 2016). Moving away from candidate miRNA expression other studies have explored miRNA expression in hibernation of analysis, this study found significant differential expression of a Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 97 Figure 1 Prediction of species-specific miRNA function from the freshwater Ramshorn snail, Biomphalaria glabrata. Novel miRNA were iden- tified from the B. glabrata genome (BgalB1; vectorbase.org) using SMIRP. Targets of novel snail miRNA were then identified using miRDeep2 from the available transcript assembly (BgalB1.5; vectorbase.org). Target genes of novel miRNA included multi-miRNA gene regu- lation of proteins involved in cellular processes such as secretory mucal proteins (mucin-21-like), matricellular proteins (thrombospondin- 3b-like), and shell formation (dentin sialophosphoprotein-like). number of miRNA in both a tissue and torpor stage-specific manner, clearly demonstrating that miRNA likely play an active role in mammalian hibernation and dynamic metabolic regulation. Although miRNA expression profiles were largely tissue-specific for the three organs studied (liver, heart, skeletal muscle), gene ontology (GO) annotation analysis revealed that the putative tar- gets of the upregulated miRNA were commonly enriched in cellular processes involved in the suppression of pro-growth. For example, in liver tissue, upregulated miRNA targeted genes enriched in cel- lular processes such as growth factor receptor signaling pathways, regulation of nuclear division, and glycolysis during the early tor- por stage (Wu et al., 2016). To further expand upon this analysis, we worked to elucidate the organization of the miRNA-targeted cellular system using the targets of the significantly upregulated miRNA from the ET stage of liver tissue. By mapping the collective targets of torpor- responsive miRNA with their known protein interactions, we were able to obtain an intuitive representation of miRNA-regulated Figure 2 Predicted functional interaction map of stress-responsive processes within a functional network. To accomplish this, we miRNA targets from the hibernating thirteen-lined ground squirrel. A used the spatial analysis of functional enrichment (SAFE) tool protein interaction map of all predicted targets of miRNA found to within the Cytoscape software (v3.4.0) (Figure 2). For each be significantly overexpressed during the early torpor (ET) stage of miRNA target within the network, SAFE defined the local neigh- ground squirrel hibernation. Network was constructed using known borhood of targets by identifying those that look to be clustered human protein interactions from the STRING database (https:// within the network. GO analysis was then carried to determine string-db.org/), contained 532 miRNA targets (i.e. nodes) and 1433 enriched miRNA-regulated cellular processes (Baryshnikova, known interactions (i.e. edges), and was originally constructed 2016). Similar to the original study, we identified an enrichment using Cytoscape (v3.4.0). Spatial analysis of functional enrichment (SAFE)-based construction of a functional map of the network by in targeting cellular signaling pathways, including growth factor combining all region-specific human GO terms into 12 functional signaling and carboxylic acid catabolism. We also identified a domains based on the similarity of their enrichment landscapes significant enrichment in the regulation of lipid biosynthesis, −4 (P values < 2 × 10 , Fisher’s exact test). Different colors represent hormone response signaling, and RNA processing (Figure 2). different functional domains. Each domain is labeled with a tag list, Lipid metabolism is known to be under tight regulatory control composed of the five words that occur most frequently within the during hibernation as triglycerides are the primary energy source names of the associated GO terms. for the hibernating mammal and are known to influence the Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 98 j Biggar and Storey length of torpor bouts and metabolic rate (Florant, 1998; Dark, and TIAR function during stress suggested that enhanced protein 2005; Wu et al., 2013). Since the entry into and arousal from aggregation was not present during torpor. Our identification of torpor is a short process that requires rapid changes in many miRNA-targeted regulation of RNA processing agrees with this study metabolic processes, this particular result is not surprising. as it identifies posttranscriptional regulatory mechanisms at play in Furthermore, with regards to the predicted enrichment of RNA reducing translational rates and/or mRNA processing. As demon- processing in torpid thirteen-lined ground squirrels, a previous strated by our deeper network-based functional analysis into torpor- study has characterized the roles of three RNA binding proteins: responsive miRNA function (Figure 2), a global analysis of miRNA T-cell intracellular antigen 1 (TIA-1), TIA-1 related (TIAR), and targets can help to provide deeper, meaningful, insight into the poly(A)-binding protein (PABP-1)(Tessier et al., 2014). TIA-1 role of miRNA in helping to coordinate the torpor–arousal cycle. was identified as a major component of sub-nuclear structures with up to a 7-fold increase in relative protein levels found in Temperature influence over microRNA–target interaction the nucleus during hibernation. Additionally, analysis of the Given the state of miRNA research within the field of com- formation of reversible aggregates that are associated with TIA-1 parative biochemistry and the functional insight that it has Figure 3 Temperature-associated regulation of miRNA function. (A) The average G/C content of complete miRNA sequences as well as their seed regions plotted against the physiological temperature of each organism. (B) For each of the organisms, miRNAs were divided into two subsets that contain (i) miRNAs that are specific to a taxonomic group and (ii) the rest miRNAs that are shared by other taxonomic groups. Each pair of bars shows the difference in G/C content between these two subsets for miRNAs (dark gray) and for seeds (light gray). Error bars show standard error of the difference between two means; (*) P < 0.05 and (**) P < 0.0005.(C) Depiction of proposed temperature influence in RNA binding thermodynamics. It is hypothesized that low temperature could act to stabilize miRNA:target interactions that were once unfavorable or unstable at higher temperatures. Data for A and B were derived from Carmel et al. (2012). Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 99 provided to date, it is becoming increasingly clear that these Overall, the increasing realization that T will likely have a small regulatory RNA are an important component of environ- dramatic impact on miRNA function, presents the possibility that mental stress survival and the hypometabolic response. Given distinct temperature-induced miRNA targeting programs may be this interest, there has also been an increasing attempt to at play and help to facilitate cellular function at various tem- explore the targets that these stress-responsive miRNA regulate peratures. In this way, there may exist distinct cold-influenced (Figure 3). In recent years, there has also been a growing inter- miRNA targeting programs that facilitate unique hypometabolic est in the possibility of temperature influencing the regulatory survival of extreme stresses. Given that the strict seed pairing function of miRNA (Figure 3). This has been previously dis- requirement of miRNA would still be required, it is possible that cussed both in terms of miRNA base-content and its relationship the relatively lower G/C content within the seed region of organ- to T (Figure 3A and B), as well as in the context of low-temperature isms living at lower temperatures may also have a role in aiding influence over miRNA:mRNA binding thermodynamics (Figure 3C) this temperature-sensitive process (Figure 3). Thus, it is interesting (Carmel et al., 2012; Biggar and Storey, 2014a, b, 2015a, b, 2017). to propose that miRNA could play a specific temperature-sensitive Given the relationship between miRNA seed binding potential role in helping various species tocopewithtemperature-related (i.e. G/C content) and temperature, the question as to whether stress. there are other mechanisms by which temperature can influence miRNA function should be asked. The targeting of miRNA to Conclusion select mRNA sites relies primarily on seed region complementar- Within the past decade, many studies have explored the role ity, with further binding from the 3′end of the miRNA only acting of miRNA as a mechanism to reversibly and rapidly regulate the to stabilize and supplement the interaction. Critically, it has cellular landscape and enable survival during periods of extreme been previously reported that the thermodynamic threshold stress. The overall impact of these studies has resulted in a (mean free energy; mfe) used to predict whether a miRNA:mRNA common theme of stress-responsive miRNA expression across target will occur is ∼18 kcal/mol. This threshold, among other phylogeny. Building upon this initial research, and with the structural requirements of the miRNA:mRNA interaction, has growing availability of genomic information and bioinformatic been used in almost all target prediction programs that have power, researchers have now begun to predict the overall cellu- been developed for the identification of human miRNA targets lar impact of stress-responsive miRNA expression in the context in mind (including miRanda, TargetScan, and Diana microT). of global target regulation. As a result, it is clear that miRNA These miRNA target identification programs typically overlook may be involved in the regulation of many cellular processes the possibility of non-human species existing at T values great- that enable efficient utilization of ATP turnover. As we learn er or lower than 37°C. Indeed, given the strong thermodynamic more about miRNA function in different organisms and in requirement for a successful miRNA:mRNA interaction, it is likely response to various environmental conditions, the possibility of that a significant change in T (such as experienced by frozen temperature having a role to play in influencing target selection frogs and turtles, hibernating mammals, and many other over- may also open comparative miRNA research to an immense wintering animals) will have a strong influence on the ability of number of regulatory possibilities that will need to be explored. miRNA target selection. In this way, a decrease in T would likely favorably stabilize miRNA–target interactions that were Acknowledgements once unfavorable, allowing these interactions to become bio- Thanks go to J. Storey (Institute of Biochemistry, Carleton logically relevant in a temperature-dependent manner. University, Canada) for editorial review of the manuscript. Indeed, one study using the FindTar3 miRNA target prediction software, which allows user control over temperature, showed a 16-fold increase in the number of potential mRNA targets when Funding comparing those predicted at freezing temperatures (3°C) and This research was funded by a Discovery grant from the those identified at 37°C(Biggar and Storey, 2015a, b). For Natural Sciences and Engineering Research Council of Canada example, in response to freezing (3°C) in the hatchling painted (NSERC; grant no. 6793) to K.B.S. turtle (Chrysemys picta marginata), miR-21 expression levels Conflict of interest: none declared. were shown to increase by over 2-fold in heart tissue (Biggar and Storey, 2015a, b). When predicting the mRNA targets of miR-21, this study compared the targets predicted at both 37°C References and 3°C, finding an increase from 47 to 756 targets at the lower Adema, C.M., Hillier, L.W., Jones, C.S., et al. (2017). Whole genome analysis of a schistosomiasis-transmitting freshwater snail. Nat. Commun. 8, 15451. temperature. Interestingly, these low-temperature targets were Bansal, S., Luu, B.E., and Storey, K.B. (2016). MicroRNA regulation in heart enriched in metabolic processes (Biggar and Storey, 2015a, b, and skeletal muscle over the freeze-thaw cycle in the freeze tolerant wood 2017; Luu et al., 2016). Within the context of a global decrease frog. J. Comp. Physiol. B 186, 229–241. in gene expression, it is likely that such an increase of miRNA Bartel, D. (2004). MicroRNAs: genomics, biogenesis, mechanism, and func- targets would dramatically alter the miRNA targeting program. tion. Cell 116, 281–297. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 100 j Biggar and Storey Baryshnikova, A. (2016). Systematic functional annotation and visualization Cowan, K.J., and Storey, K.B. (2003). Mitogen-activated protein kinases: new of biological networks. Cell Syst. 2, 412–421. signaling pathways functioning in cellular responses to environmental Biggar, K.K., Dubuc, A., and Storey, K.B. (2009). MicroRNA regulation below stress. J. Exp. Biol. 206, 1107–1115. zero: differential expression of miRNA-21 and miRNA-16 during freezing in Dark, J. (2005). Annual lipid cycles in hibernators: integration of physiology wood frogs. Cryobiology 59, 317–321. and behavior. Annu. Rev. Nutr. 25, 469–497. Biggar, K.K., Kornfeld, S.F., and Storey, K.B. (2011). Amplification and Epperson, L.E., and Martin, S.L. (2002). Quantitative assessment of ground sequencing of mature microRNAs in uncharacterized animal models using squirrel mRNA levels in multiple stages of hibernation. Physiol. Genomics stem-loop reverse transcription-polymerase chain reaction. Anal. Biochem. 10, 93–102. 416, 231–233. Fraser, K.P., Houlihan, D.F., Lutz, P.L., et al. (2001). Complete suppression of Biggar, K.K., Kornfeld, S.F., Maistrovski, Y., et al. (2012). MicroRNA regulation protein synthesis during anoxia with no post-anoxia protein synthesis in extreme environments: differential expression of microRNAs in the inter- debt in the red-eared slider turtle, Trachemys scripta elegans. J. Exp. Biol. tidal snail Littorina littorea during extended periods of freezing and 204, 4353–4360. anoxia. Genomics Proteomics Bioinformatics 10, 302–309. Frerichs, K.U., Smith, C.B., Brenner, M., et al. (1998). Suppression of protein Biggar, K.K., and Storey, K.B. (2012). Evidence for cell cycle suppression and synthesis in brain during hibernation involves inhibition of protein initi- microRNA regulation of cyclin D1 during anoxia exposure in turtles. Cell ation and elongation. Proc. Natl Acad. Sci. USA 95, 14511–14516. Cycle 11, 1705–1713. Florant, G.L. (1998). Lipid metabolism in hibernators: the importance of Biggar, K.K., and Storey, K.B. (2011). The emerging roles of microRNAs in the essential fatty acids. Integr. Comp. Biol. 38, 331–340. molecular responses of metabolic rate depression. J. Mol. Cell Biol. 3, Frigault, J.J., Lang-Ouellette, D., and Morin, P.J. (2016). Up-regulation of non- 167–175. coding RNA Tug1 in hibernating thirteen-lined ground squirrels, Ictidomys Biggar, K.K., and Storey, K.B. (2014a). Identification and expression of tridecemlineatus. Genomics Proteomics Bioinformatics 14, 113–118. microRNA in the brain of hibernating bats, Myotis lucifugus. Gene 544, Frigault, J.J., Morin, M.D., and Morin, P.J. (2017). Differential expression and 67–74. emerging functions of non-coding RNAs in cold adaptation. J. Comp. Biggar, K.K., and Storey, K.B. (2014b). Insight into temperature-dependent Physiol. B 187, 19–28. microRNA function in mammalian hibernators: perspectives on cold- Friedlander, M.R., Mackowiak, S.D., Li, N., et al. (2012). MiRDeep2 accurately influenced microRNA/target interaction. Temperature 1, 84–86. identifies known and hundreds of novel microRNA genes in seven animal Biggar, K.K., and Storey, K.B. (2015a). Insight into post-transcriptional gene clades. Nucleic Acids Res. 40, 37–52. regulation: stress-responsive microRNAs and their role in the environmen- Friedman, R.C., Farh, K.K.H., Burge, C.B., et al. (2009). Most mammalian tal stress survival of tolerant animals. J. Exp. Biol. 218, 1281–1289. mRNAs are conserved targets of microRNAs. Genome Res. 19, 92–105. Biggar, K.K., and Storey, K.B. (2015b). Low-temperature microRNA expres- Gabrial, U.I., Mirela, S., and Ionel, J. (2011). Quantification of mucoproteins sion in the painted turtle, Chrysemys picta during freezing stress. FEBS (glycoproteins) from snail mucus, Helix aspersa and Helix pomatia. J. Agro. Lett. 589, 3665–3670. Proc. Tech. 17, 410–413. Biggar, K.K., and Storey, K.B. (2017). Exploration of low temperature Guppy, M., and Withers, P. (1999). Metabolic depression in animals: physio- microRNA function in an anoxia tolerant vertebrate ectotherm, the red logical perspectives and biochemical generalizations. Biol. Rev. Camb. eared slider turtle (Trachemys scripta elegans). J. Therm. Biol. 68, Philos. Soc. 74, 1–40. 139–146. Hadj-Moussa, H., Moggridge, J.A., Luu, B.E., et al. (2016). The hibernating Biggar, K.K., Wu, C.W., and Storey, K.B. (2014). High-throughput amplifica- South American marsupial, Dromiciops gliroides, displays torpor-sensitive tion of mature microRNAs in uncharacterized animal models using polya- microRNA expression patterns. Sci. Rep. 6, 24627. denylated RNA and stem-loop reverse transcription polymerase chain Hittel, D., and Storey, K.B. (2002). The translation state of differentially reaction. Anal. Biochem. 462, 32–34. expressed mRNAs in the hibernating 13-lined ground squirrel Britton, C., Winter, A.D., Gillan, V., et al. (2014). MicroRNAs of parasitic hel- (Spermophilus tridecemlineatus). Arch. Biochem. Biophys. 401, 244–254. minths – identification, characterization and potential as drug targets. Int. Hochachka, P.W., Buck, L.T., Doll, C.J., et al. (1996). Unifying theory of hyp- J. Parasitol. 4, 85–94. oxia tolerance: molecular/metabolic defense and rescue mechanisms for Brooks, S.P.J., and Storey, K.B. (1993). De novo protein synthesis and protein surviving oxygen lack. Proc. Natl Acad. Sci. USA 93, 9493–9498. phosphorylation during anoxia and recovery in the red-eared turtle. Am. J. John, B., Enright, A.J., Aravin, A., et al. (2005). MiRanda application: Human Physiol. 265,R1380–R1386. microRNA targets. PLoS Biol. 3,e264. Brooks, S.P.J., and Storey, K.B. (1995). Protein phosphorylation patterns dur- Kornfeld, S.F., Biggar, K.K., and Storey, K.B. (2012). Differential expression of ing aestivation in the land snail Otala lactea. Mol. Cell. Biochem. 143, mature microRNAs involved in muscle maintenance of hibernating little 7–13. brown bats, Myotis lucifugus: a model of muscle atrophy resistance. Carmel, I., Shomron, N., and Heifetz, Y. (2012). Does base-pairing strength Genomics Proteomics Bioinformatics 10, 295–301. play a role in microRNA repression? RNA 18, 1947–1956. Krol, J., Loedige, I., and Filipowicz, W. (2010). The widespread regulation of Chen, M., Zhang, X., Liu, J., et al. (2013). High-throughput sequencing reveals microRNA biogenesis, function and decay. Nat. Rev. Genet. 11, 597–610. differential expression of miRNAs in intestine from sea cucumber during Land, S.C., Buck, L.T., and Hochachka, P.W. (1993). Response of protein syn- aestivation. PLoS One 8,e76120. thesis to anoxia and recovery in anoxia-tolerant hepatocytes. Am. J. Chen, M., and Storey, K.B. (2014). Large-scale identification and comparative Physiol. 265,R41–R48. analysis of miRNA expression profile in the respiratory tree of the sea Lang-Ouellette, D., and Morin, P.J. (2014). Differential expression of miRNAs cucumber Apostichopus japonicus. Mar. Genomics 13, 39–44. with metabolic implications in hibernating thirteen-lined ground squirrels, Cipolla, G.A. (2014). A non-canonical landscape of the microRNA system. Ictidomys tridecemlineatus. Mol. Cell. Biochem. 394, 291–298. Front. Genet. 5, 337. Larade, K., and Storey, K.B. (2002). Reversible suppression of protein synthe- Courteau, L.A., Storey, K.B., and Morin, P.J. (2012). Differential expression of sis in concert with polysome disaggregation during anoxia exposure in microRNA species in a freeze tolerant insect, Eurosta solidaginis. Littorina littorea. Mol. Cell. Biochem. 232, 121–127. Cryobiology 65, 210–214. Lee, Y.J., Johnson, K.R., and Hallenbeck, J.M. (2012). Global protein con- Cowan, K.J., MacDonald, J.A., Storey, J.M., et al. (2000). Metabolic reorgan- jugation by ubiquitin-like-modifiers during ischemic stress is regulated ization and signal transduction during estivation in the spadefoot toad. by microRNAs and confers robust tolerance to ischemia. PLoS One 7, Exp. Biol. Online 5, 1–25. e47787. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018 MicroRNA regulation and stress tolerance j 101 Liu, Y., Hu, W., Wang, H., et al. (2010). Genomic analysis of miRNAs in an Shaffer, H.B., Minx, P., Warren, D.E., et al. (2013). The western painted turtle extreme mammalian hibernator, the Arctic ground squirrel. Physiol. genome, a model for the evolution of extreme physiological adaptations in Genomics 42A, 39–51. a slowly evolving lineage. Genome Biol. 14,R28. Luu, B.E., Biggar, K.K., Wu, C.W., et al. (2016). Torpor-responsive expression Storey, K.B. (2015). Regulation of hypometabolism: insights into epigenetics of novel microRNA regulating metabolism and other cellular pathways in controls. J. Exp. Biol. 218, 150–159. the thirteen-lined ground squirrel, Ictidomys tridecemlineatus. FEBS Lett. Storey, K.B., and Storey, J.M. (2004). Metabolic depression in animals: tran- 590, 3574–3582. scriptional and translational controls. Biol. Rev. 79, 207–233. Luu, B.E., and Storey, K.B. (2015). Dehydration triggers differential microRNA Tessier, S.N., Audas, T.E., and Wu, C.W. (2014). The involvement of mRNA expression in Xenopus laevis brain. Gene 573, 64–69. processing factors TIA-1, TIAR, and PABP-1 during mammalian hibernation. Lyons, P.J., Crapoulet, N., Storey, K.B., et al. (2015a). Identification and pro- Cell Stress Chaperones 19, 813–825. filing of miRNAs in the freeze-avoiding gall moth Epiblema scudderiana via Volk, T., Wang, S., Rotstein, B., et al. (2014). Matricellular proteins in devel- next-generation sequencing. Mol. Cell. Biochem. 410, 155–163. opment: perspectives from the Drosophila heart. Matrix. Biol. 37, Lyons, P.J., Govaere, L., Crapoulet, N., et al. (2016). Characterization of cold- 162–166. associated microRNAs in the freeze-tolerant gall fly Eurosta solidaginis Wu, C., So, J., Davis-Dusenbery, B.N., et al. (2011). Hypoxia potentiates using high-throughput sequencing. Comp. Biochem. Physiol. Part D microRNA-mediated gene silencing through posttranslational modification Genomics Proteomics 20, 95–100. of argonaute 2. Mol. Cell. Biol. 31, 4760–4774. Lyons, P.J., Poitras, J.J., Courteau, L.A., et al. (2013). Identification of differen- Wu, C.W., Biggar, K.K., and Storey, K.B. (2013). Biochemical adaptations of tially regulated microRNAs in cold-hardy insects. Cryo. Lett. 34, 83–89. mammalian hibernation: exploring squirrels as a perspective model for Lyons, P.J., Storey, K.B., and Morin, P.J. (2015b). Expression of miRNAs in naturally induced reversible insulin resistance. Braz. J. Med. Biol. Res. 46, response to freezing and anoxia stresses in the freeze tolerant fly Eurosta 1–13. solidaginis. Cryobiology 71, 97–102. Wu, C.W., Biggar, K.K., and Storey, K.B. (2014a). Expression profiling and Maistrovski, Y., Biggar, K.K., and Storey, K.B. (2012). HIF-1α regulation in structural characterization of microRNAs in adipose tissues of hibernating mammalian hibernators: role of non-coding RNA in HIF-1α control during ground squirrels. Genomics Proteomics Bioinformatics 12, 284–291. torpor in ground squirrels and bats. J. Comp. Physiol. B 182, 849–859. Wu, C.W., Biggar, K.K., and Storey, K.B. (2014b). Dehydration mediated Marxen, J.C., and Becker, W. (2011). Calcium binding constituents of the microRNA response in the African clawed frog, Xenopus laevis. Gene 529, organic shell matrix from the freshwater snail Biomphalaria glabrata. 3376–3383. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 127, 235–242. Wu, C.W., Biggar, K.K., Luu, B.E., et al. (2016). Analysis of microRNA expres- Morin, P.J., Dubuc, A., and Storey, K.B. (2007). Differential expression of sion during the torpor-arousal cycle of a mammalian hibernator, the 13- microRNA species in organs of hibernating ground squirrels: a role in transla- lined ground squirrel. Physiol. Genomics 48, 388–396. tional suppression during torpor. BBA Gene Regul. Mech. 1779, 628–633. Wu, C.W., and Storey, K.B. (2013). Regulation of the mTOR signaling network Peace, R.J., Biggar, K.K., Storey, K.B., et al. (2015). A framework for improving in hibernating thirteen-lined ground squirrels. J. Exp. Biol. 215, microRNA prediction in non-human genomes. Nucleic Acids Res. 43,e138. 1720–1727. Podrabsky, J.E., and Hand, S.C. (2000). Depression of protein synthesis dur- Yamasaki, S., and Anderson, P. (2008). Reprogramming mRNA translation ing diapause in embryos of the annual killifish, Austrofundulus limnaeus. during stress. Curr. Opin. Cell Biol. 20, 222–226. Physiol. Biochem. Zool. 73, 799–808. Yi, R., Qin, Y., Macara, I.G., et al. (2003). Exportin-5 mediates the nuclear Pothof, J., Verkaik, N.S., Hoeijmakers, J.H., et al. (2009). MicroRNA responses export of pre-microRNAs and short hairpin RNAs. Genes Dev. 17, and stress granule formation modulate the DNA damage response. Cell 3011–3016. Cycle 8, 3462–3468. Zhang, J., Biggar, K.K., and Storey, K.B. (2013). Regulation of p53 by revers- Rouble, A.N., Biggar, K.K., and Storey, K.B. (2014). A high-throughput proto- ible post-transcriptional and post-translational mechanisms in liver and col for message RNA quantification using RNA dot-blots. Anal. Biochem. skeletal muscle of an anoxia tolerant turtle, Trachemys scripta elegans. 452, 31–33. Gene 513, 147–155. Schaap, P., Barrantes, I., Minx, P., et al. (2016). The Physarum polycephalum Zeng, Y., Yi, R., and Cullen, B.R. (2005). Recognition and cleavage of primary genome reveals extensive use of prokaryotic two-component and microRNA precursors by the nuclear processing enzyme Drosha. EMBO J. metazoan-type tyrosine kinase signaling. Genome Biol. Evol. 8, 109–125. 24, 138–148. Downloaded from https://academic.oup.com/jmcb/article-abstract/10/2/93/4682967 by Ed 'DeepDyve' Gillespie user on 20 June 2018

Journal

Journal of Molecular Cell Biology – Oxford University Press

Published: Jan 9, 2018

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