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doi: 10.1104/pp.19.00389pmid: 33814620
ABC Transporter Gene Enhances Chickpea Yield The development of crop cultivars with increased seed number or seed size and weight (SW) is critical for ensuring global food and nutritional security. Because of the importance of SW, substantial efforts have been directed toward identifying genetic factors regulating this yield-contributing trait. These efforts have uncovered various signaling pathways that function in synchrony to regulate SW. These signaling pathways predominantly function through modulation of cell proliferation and cell expansion during seed development. Chickpea (Cicer arietinum), the genome of which has recently been sequenced, exhibits distinct phenotypic variation for diverse agromorphological traits, including pod and seed yield component traits. Establishing a comprehensive understanding of the complex genetic architecture of SW traits is essential for facilitating future efforts to genetically enhance chickpea. Using a combinatorial genomics-assisted breeding and functional genomics strategy, Basu et al. (pp. 253–275) have uncovered an ABCC3-type ABC transporter gene that regulates SW in two chickpea cultivars. The ABC transporter gene identified regulates SW by modulating glutathione conjugate transport into vacuoles. The superior allele/haplotype of this gene in near-isogenic lines enhances the seed weight, yield, productivity, and multiple desirable plant architecture and seed quality traits without compromising agronomic performance. These findings will facilitate the development of nutritionally enriched high-yielding cultivars in chickpea. Readers interested in the control of SW may also wish to examine the contribution by Shi et al. (pp. 381–391), wherein the authors characterize a dominant gain-of-function rice (Oryza sativa) mutant that develops significantly enlarged grains. Screening For Blockers of Plant Calcium Channels The importance of Ca2+ in animal physiology led to the development of an extensive pharmacological toolbox to manipulate specific groups of Ca2+ channels. Simple drug treatments allow for the manipulation of specific Ca2+ channels, thereby providing insights of which Ca2+ channels are involved in any process of interest. Unfortunately, the Ca2+ signaling machinery in plants has diverged significantly from the one in the animal kingdom, with many of the Ca2+ signaling components of animals being absent in plants and vice versa. For instance, channels associated with muscle and nerve Ca2+ signal transduction in animals, such as l-type voltage-dependent Ca2+ channels, inositol 1,4,5-triphosphate receptors, and ryanodine receptors, are either missing or significantly different in plants. Hence, many Ca2+ channel inhibitors established in animal systems are arguably of limited use in plants. Thus, there is an important need to develop Ca2+ channel inhibitors directly in plant systems to stimulate research in plant Ca2+ signaling. De Vriese et al. (pp. 480–496) have developed a chemical screen to identify small molecules that modify the shape and amplitude of auxin-induced cytosolic Ca2+ dynamics and use them to further explore the role of Ca2+ signaling in cellular auxin responses. They report that auxin-induced Ca2+ signaling is highly sensitive to disruption of membrane proton gradients and the mammalian Ca2+ channel inhibitor bepridil. Whereas protonophores nonselectively inhibited auxin-induced and osmotic stress-induced Ca2+ signals, bepridil specifically inhibited auxin-induced Ca2+. Bepridil also severely alters vacuolar morphology and antagonizes auxin-induced vacuolar remodeling. Further exploration of this plant-tailored collection of inhibitors will lead to a better understanding of auxin-induced Ca2+ entry and its relevance for auxin responses. Evolution of Frost Tolerance in the Pooideae Frost is one of the most severe abiotic stresses a plant can experience, and exposure to frost is a limiting factor for many species in temperate and arctic regions. Although the grass subfamily Pooideae dominates temperate and arctic grass floras, the ancestors of this group were most likely adapted to tropical or subtropical climates. Cold acclimation, wherein plants increase their frost tolerance in response to gradually falling temperatures and shorter days in the autumn, had to have been a critical adaptation in the spread of Pooideae to temperate and arctic regions. Understanding how complex traits such as cold acclimation evolved remains a major challenge in evolutionary biology. Schubert et al. (pp. 404–419) report on their investigations concerning the evolution of cold acclimation in the Pooideae. The authors’ examination of differential gene expression after cold treatment in the transcriptomes of five phylogenetically diverse species revealed widespread species-specific responses of genes with conserved sequences. The evolution of both protein-coding and regulatory DNA sequences as well as the de novo origin of novel genes and functions contributed toward the evolution of the cold acclimation response in the Pooideae. These results suggest that selection pressure resulting from global cooling during the Cenozoic must have acted on already diverged lineages. Nevertheless, conservation of cold-induced gene expression of certain genes indicates that the Pooideae ancestor may have possessed some molecular machinery to mitigate cold stress. In toto, however, the data suggest a mostly independent evolution of cold acclimation responses in the Pooideae. Effects of Isoprene on Plant Growth Abiotic stress conditions such as drought and salt stress increase isoprene synthesis and emission in many plant species. The carbon and energy costs of isoprene production have been found to vary depending on the plant species and environmental conditions. For example, trees that normally produce isoprene, such as eastern cottonwood (Populus deltoides), consume about 2% of photosynthetic C for isoprene production, while kudzu (Pueraria montana) can consume up to 64%. Given the potential costs associated with isoprene emission, it is not surprising that the property of isoprene emission has been lost in some species: Arabidopsis (Arabidopsis thaliana) and tobacco (Nicotiana tabacum), for example, do not produce isoprene. A key enzyme in isoprene production is isoprene synthase (ISPS), which converts dimethylallyl diphosphate into isoprene. Understanding how isoprene affects plant growth and physiology will allow researchers to determine whether ISPS will be a beneficial trait to be reintroduced to plants, especially for the purpose of crop improvement. With this strategy in mind, Zuo et al. (pp. 124–152) have transformed Arabidopsis with an ISPS gene from Eucalyptus globulus. The regulatory mechanisms of photosynthesis and isoprene emission in these transformed plants were found to be similar to those of native emitters, indicating that the regulatory components of isoprene emission are not specific to isoprene-emitting species. The leaf chlorophyll and carotenoid contents of the Arabidopsis transformants were enhanced by isoprene, which also had a marked positive effect on hypocotyl, cotyledon, leaf, and inflorescence growth. By contrast, leaf and stem growth was reduced in tobacco engineered to emit isoprene. The expressions of genes belonging to signaling networks or associated with specific growth regulators (e.g. gibberellic acid and jasmonic acid) were altered by isoprene emission, as were genes involved in stress tolerance. The authors propose that isoprene likely executes its effects on growth and stress tolerance through direct regulation of gene expression and that the enhancement of jasmonic acid-mediated defense signaling by isoprene may trigger a growth-defense tradeoff leading to variations in the growth response. Systems Biology of Deetiolation Upon exposure to light, many leaf and stem cells acquire photosynthetic competence by converting pale etioplasts into green chloroplasts. Deetiolation involves the concerted and synchronized activity of a highly complex biogenesis program. Thylakoid membranes must develop from disassembling prolamellar bodies and prothylakoids and from newly synthesized lipids. Moreover, large protein complexes containing dozens of protein subunits and hundreds of pigments and cofactors must be inserted into the budding membrane in strictly defined stoichiometric ratios. The protein complexes involved consist of polypeptides originating from two evolutionarily distinct compartments, the nucleus and the plastid, which must be expressed, processed, targeted, and inserted into the membrane in a highly coordinated manner. These processes are dependent upon and controlled by a wide range of assembly chaperones and other biogenesis factors, which are not or only poorly understood. In spite of the complexity of thylakoid biogenesis, the etioplast-to-chloroplast transition can occur astoundingly rapidly. Armarego-Marriott et al. (pp. 654–681) have developed a system to study both the deetiolation process and the process of photosynthetic maturation in leaves of tobacco at high temporal resolution. Targeted and nontargeted approaches were undertaken to define the dynamic changes in the transcriptomes of the nucleus and the plastid. In addition, the accumulation kinetics of pigments, lipids, soluble metabolites, and photosynthetic proteins and their activities were determined and correlated with the physical changes in membrane ultrastructure. This work provides a comprehensive systems-level description of thylakoid development and the etioplast-to-chloroplast differentiation process and also reveals candidate genes involved in chloroplast biogenesis and the acquisition of photosynthetic competence. A Tonoplast Calcineurin B-Like Protein and Stomatal Movement SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) comprise a highly conserved superfamily of proteins in all eukaryotic cells and play important roles in membrane fusion events involved in the delivery of membranes, proteins, and soluble cargos. SNARES form a core complex to bring vesicle and target membrane surfaces together, thereby driving secretion as well as the traffic of vesicles between endosomal compartments. Beyond their canonical role in membrane fusion, a few SNAREs are also known to interact with ion channels and affect their regulation. The plasma membrane SNARE SYP121 of Arabidopsis is the best-known example. More specifically, SYP121 interacts with the K+ channels KC1 and KAT1, altering channel gating to promote K+ uptake. Channel binding is specific for SYP121: it depends on a conserved N-terminal motif defined by the sequence F9xRF within SYP121. Much of vesicle traffic at the Arabidopsis plasma membrane, however, is subject to the protein SEC11, which also selectively binds with SYP121. How the binding of SEC11 with SYP121 is coordinated with SYP121 interactions with K+ channels is poorly understood, as both SEC11 and the channels are thought to compete for the same SNARE binding site. Zhang et al. (pp. 228–239) now identify a second binding motif within the N terminus of SYP121 and demonstrate that this motif affects SEC11 binding independently of the F9xRF motif that is shared with the K+ channels. This second, previously unrecognized motif is centered on residues R20R21 of SYP121 and is essential for SEC11 interaction with SYP121. Mutation of the R20R21 motif blocked vesicle traffic without uncoupling the effects of SYP121 on solute and K+ uptake associated with the F9xRF motif. Author notes www.plantphysiol.org/cgi/doi/10.1104/pp.19.00389 © 2019 American Society of Plant Biologists. All Rights Reserved. This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model)
doi: 10.1104/pp.19.00260pmid: 31053670
For a number of crops, agricultural yields have been boosted by the phenomenon of heterosis or hybrid vigor, where a hybrid outperforms either parent. Understanding the various mechanisms by which heterosis arises may facilitate the breeding of hybrids with increased vigor for further crop varieties or species. Vigor can be determined by measurement of a number of traits from seed yield and plant biomass through to biotic and abiotic stress resistance. Tradeoffs may exist between some of these traits such that breeding for one may negatively affect another desirable phenotype (e.g. defense versus growth; Albrecht and Argueso, 2017). A number of plant hormones act in regulating plant growth and defense with both synergistic and antagonistic effects. Maize (Zea mays) and rice (Oryza sativa) are two key crops for which heterosis has increased yields. In rice, altered regulation of gibberellic acid and abscisic acid biosynthesis genes can lead to hybrid vigor (Chen et al., 2018), whereas factors that contribute to heterosis in maize include the masking of recessive deleterious mutations (Yang et al., 2017) and changes in the epigenetic landscape associated with subsequent gene expression (Seifert et al., 2018). As for many biological processes, Arabidopsis (Arabidopsis thaliana) has been used as a model in the study of hybrid vigor, with a number of pathways and genes that contribute to heterosis identified over the past decade. For Ler × C24 hybrids of Arabidopsis, auxin signaling and transport pathways, as well as epigenetic regulation of genes, have been identified as mechanisms of heterosis (Groszmann et al., 2011; Shen et al., 2012). These studies confirm the potential for hormone signaling and epigenetic regulation of gene expression to contribute to hybrid vigor. For C24 × Col-0 hybrids, heterosis in rosette growth is linked to upregulation of photosynthesis genes and downregulation of stress response genes that are coregulated by the circadian clock (Miller et al., 2015). Although far from comprehensive, these previous studies demonstrate the complexity of heterosis, which has different causes between hybrids and species and multiple factors contributing to heterosis within a single hybrid. In contrast to the varied mechanisms of heterosis, the opposite phenomenon of hybrid incompatibility or hybrid weakness is frequently based on upregulation of defense response genes (Chae et al., 2014). In this issue of Plant Physiology, Ian Greaves’ group at CSIRO Agriculture and Food in Canberra further explored the basis of heterosis in C24 × Ler hybrids of Arabidopsis (Gonzalez-Bayon et al., 2019). This study builds on their previous body of work showing that hormone pathway and biotic response genes are differently expressed in hybrids of C24 × Ler, C24 × Col-0, and Col-0 × Ler (Groszmann et al., 2015). The group has previously shown that a reduction in the defense hormone salicylic acid leads to increased vigor in C24 plants, whereas decreased salicylic acid and auxin responses are present in C24 × Ler and C24 ± Col-0 hybrids (Groszmann et al., 2015). Based on these results, the authors hypothesized that hybrid vigor can in some instances be the reverse of hybrid incompatibility, i.e. that downregulation of defense response genes in a hybrid may lead to heterosis. It is important to note that C24 is physiologically an outlier compared to other Arabidopsis accessions (Ferguson et al., 2018). With reduced stomatal conductance, possibly linked to the higher salicylic acid levels, C24 is drought-, heat-, and ozone-tolerant, but more sensitive to some other forms of abiotic stress (Bechtold et al., 2010, 2018; Brosché et al., 2010; Xu et al., 2015). C24 has been known for almost a decade to have constitutively high salicylic acid levels with small rosettes but normal levels of seed yield when not stressed (Bechtold et al., 2010). Thus, the use of rosette biomass as an indicator of overall fitness does not apply to C24. In this article, the authors have explored further the role of salicylic acid in C24 and C24 × Ler hybrids. The authors confirmed their previous finding that decreasing salicylic acid levels in C24, through the introduction of a bacterial NahG gene that converts the hormone into inactive catechol, leads to an increase in plant size after 21 d of growth. Whether a further reduction in salicylic acid levels in the C24 × Ler hybrid would also lead to an increase in growth has not been not tested, although the fresh weight of mature C24 NahG rosettes was equivalent to that of the C24 × Ler F1 plants. However, the hybrids were larger than either parent or the C24 NahG plants early in growth, indicating that there is an additional heterosis mechanism other than salicylic acid active during seedling establishment. Through a detailed transcriptome comparison of C24 NahG, Ler NahG, and F1 C24 × Ler plants to the midparental value, Gonzalez-Bayon et al. (2019) found that the majority of differentially expressed genes were downregulated, rather than upregulated, and therefore focused on this subset of 771 repressed genes. Unsurprisingly, genes within the salicylic acid biosynthesis pathway were downregulated in the C24 NahG and hybrid lines, consistent with salicylic acid levels being lower than the C24 value in these lines. In the Ler NahG line, where salicylic acid levels were not significantly reduced beyond the already low levels of the hormone in the Ler wild type, various salicylic acid-responsive defense genes were also downregulated, but to a lesser extent. In the C24 NahG and hybrid lines, lower gene expression was largely due to a reduction in expression of C24-derived genes rather than the Ler genes. That downregulation of defense responses may facilitate increased plant growth as a trade-off between growth and defense generally is accepted in plant biology (Karasov et al., 2017), although this connection is not absolute (see Campos et al., 2016). One important salicylic acid-inducible regulatory gene that has been linked to the balance between growth and defense is TL1 BINDING TRANSCRIPTION FACTOR1 (Pajerowska-Mukhtar et al., 2012), a gene whose transcript levels were significantly reduced in the F1 hybrids and C24 NahG plants. Likewise, genes connected to salicylic-acid–induced senescence were downregulated in these lines, which Gonzalez-Bayon et al. (2019) linked to a brief delay in senescence providing leaves with a longer period of maximum photosynthetic potential. Previously, the Greaves group has reported that increased plant size in C24 × Ler F1 hybrids and C24 NahG plants is partially due to increase in cell size (Groszmann et al., 2015). In this article, they expand on this interesting result, showing that a handful of genes associated with increased cell size (encoding expansins and xyloglucan-modifying proteins) were upregulated in these lines, whereas extensins that limit cell size were downregulated. Whether these genes act downstream of salicylic acid remains to be explored. In summary, downregulation of salicylic acid levels can increase growth in an accession with unusually high salicylic acid levels. In comparison, a hybrid from this accession that exhibits heterosis also showed downregulation of salicylic acid-regulated defense genes. Both genotypes exhibit delayed senescence, allowing leaves to contribute more to growth through a prolonged period of maximum photosynthetic capacity. However, as evidenced by (1) the small size of the Ler parental line despite its low levels of salicylic acid, and (2) previous literature, hormone levels are only one part of the jigsaw puzzle in plant growth and heterosis. The wider impact of the research presented by Gonzalez-Bayon et al. (2019) is that selection for downregulation of the salicylic-acid pathway in inbred accessions may prove to be a viable strategy to increase plant size and hence crop yields. Whether this approach is also useful for hybrids will require further investigation and may only be effective in cases where one parent has uncommonly high salicylic acid levels to begin with. LITERATURE CITED Albrecht T , Argueso CT ( 2017 ) Should I fight or should I grow now? The role of cytokinins in plant growth and immunity and in the growth-defence trade-off . Ann Bot 119 : 725 – 735 Google Scholar PubMed OpenURL Placeholder Text WorldCat Bechtold U , Lawson T, Mejia-Carranza J, Meyer RC, Brown IR, Altmann T, Ton J, Mullineaux PM ( 2010 ) Constitutive salicylic acid defences do not compromise seed yield, drought tolerance and water productivity in the Arabidopsis accession C24 . Plant Cell Environ 33 : 1959 – 1973 Google Scholar Crossref Search ADS PubMed WorldCat Bechtold U , Ferguson JN, Mullineaux PM ( 2018 ) To defend or to grow: Lessons from Arabidopsis C24 . J Exp Bot 69 : 2809 – 2821 Google Scholar Crossref Search ADS PubMed WorldCat Brosché M , Merilo E, Mayer F, Pechter P, Puzõrjova I, Brader G, Kangasjärvi J, Kollist H ( 2010 ) Natural variation in ozone sensitivity among Arabidopsis thaliana accessions and its relation to stomatal conductance . Plant Cell Environ 33 : 914 – 925 Google Scholar Crossref Search ADS PubMed WorldCat Campos ML , Yoshida Y, Major IT, de Oliveira Ferreira D, Weraduwage SM, Froehlich JE, Johnson BF, Kramer DM, Jander G, Sharkey TD, et al. ( 2016 ) Rewiring of jasmonate and phytochrome B signalling uncouples plant growth-defense tradeoffs . Nat Commun 7 : 12570 Google Scholar Crossref Search ADS PubMed WorldCat Chae E , Bomblies K, Kim ST, Karelina D, Zaidem M, Ossowski S, Martín-Pizarro C, Laitinen RA, Rowan BA, Tenenboim H, et al. ( 2014 ) Species-wide genetic incompatibility analysis identifies immune genes as hot spots of deleterious epistasis . Cell 159 : 1341 – 1351 Google Scholar Crossref Search ADS PubMed WorldCat Chen L , Bian J, Shi S, Yu J, Khanzada H, Wassan GM, Zhu C, Luo X, Tong S, Yang X, et al. ( 2018 ) Genetic analysis for the grain number heterosis of a super-hybrid rice WFYT025 combination using RNA-Seq . Rice (NY) 11 : 37 Google Scholar Crossref Search ADS WorldCat Ferguson JN , Humphry M, Lawson T, Brendel O, Bechtold U ( 2018 ) Natural variation of life‐history traits, water use, and drought responses in Arabidopsis . Plant Direct 2 : e00035 Google Scholar Crossref Search ADS PubMed WorldCat Gonzalez-Bayon R , Shen Y, Groszmann M, Zhu A, Wang A, Allu AD, Dennis LS, Peacock WJ, Greaves IK ( 2019 ) Senescence and defense pathways contribute to heterosis . Plant Physiol 180 : 240 – 252 Google Scholar Crossref Search ADS PubMed WorldCat Groszmann M , Greaves IK, Albertyn ZI, Scofield GN, Peacock WJ, Dennis ES ( 2011 ) Changes in 24-nt siRNA levels in Arabidopsis hybrids suggest an epigenetic contribution to hybrid vigor . Proc Natl Acad Sci USA 108 : 2617 – 2622 Google Scholar Crossref Search ADS PubMed WorldCat Groszmann M , Gonzalez-Bayon R, Lyons RL, Greaves IK, Kazan K, Peacock WJ, Dennis ES ( 2015 ) Hormone-regulated defense and stress response networks contribute to heterosis in Arabidopsis F1 hybrids . Proc Natl Acad Sci USA 112 : E6397 – E6406 Google Scholar Crossref Search ADS PubMed WorldCat Karasov TL , Chae E, Herman JJ, Bergelson J ( 2017 ) Mechanisms to mitigate the trade-off between growth and defense . Plant Cell 29 : 666 – 680 Google Scholar Crossref Search ADS PubMed WorldCat Miller M , Song Q, Shi X, Juenger TE, Chen ZJ ( 2015 ) Natural variation in timing of stress-responsive gene expression predicts heterosis in intraspecific hybrids of Arabidopsis . Nat Commun 6 : 7453 Google Scholar Crossref Search ADS PubMed WorldCat Pajerowska-Mukhtar KM , Wang W, Tada Y, Oka N, Tucker CL, Fonseca JP, Dong X ( 2012 ) The HSF-like transcription factor TBF1 is a major molecular switch for plant growth-to-defense transition . Curr Biol 22 : 103 – 112 Google Scholar Crossref Search ADS PubMed WorldCat Seifert F , Thiemann A, Grant-Downton R, Edelmann S, Rybka D, Schrag TA, Frisch M, Dickinson HG, Melchinger AE, Scholten S ( 2018 ) Parental expression variation of small RNAs is negatively correlated with grain yield heterosis in a maize breeding population . Front Plant Sci 9 : 13 Google Scholar Crossref Search ADS PubMed WorldCat Shen H , He H, Li J, Chen W, Wang X, Guo L, Peng Z, He G, Zhong S, Qi Y, et al. ( 2012 ) Genome-wide analysis of DNA methylation and gene expression changes in two Arabidopsis ecotypes and their reciprocal hybrids . Plant Cell 24 : 875 – 892 Google Scholar Crossref Search ADS PubMed WorldCat Xu E , Vaahtera L, Hõrak H, Hincha DK, Heyer AG, Brosché M ( 2015 ) Quantitative trait loci mapping and transcriptome analysis reveal candidate genes regulating the response to ozone in Arabidopsis thaliana . Plant Cell Environ 38 : 1418 – 1433 Google Scholar Crossref Search ADS PubMed WorldCat Yang J , Mezmouk S, Baumgarten A, Buckler ES, Guill KE, McMullen MD, Mumm RH, Ross-Ibarra J ( 2017 ) Incomplete dominance of deleterious alleles contributes substantially to trait variation and heterosis in maize . PLoS Genet 13 : e1007019 Google Scholar Crossref Search ADS PubMed WorldCat Author notes 1 Author for contact: [email protected]. 2 Senior author. www.plantphysiol.org/cgi/doi/10.1104/pp.19.00260 © 2019 American Society of Plant Biologists. All Rights Reserved. This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model)
doi: 10.1104/pp.19.00333pmid: 31053671
The Poaceae is one of the largest plant families in angiosperms, containing more than 12,000 grass species, which are classified into two major clades: 1) Panicoideae, Arundinoideae, Chloridoideae, Micrairoideae, Aristidoideae and Danthonioideae and 2) Bambusoideae, Oryzoideae and Pooideae (Fig. 1; Kellogg, 2015). The Poaceae originally evolved in warm moist habitats, and further successfully adapted to a worldwide range of climatic regimes from tropics to temperate zones including freezing Arctic and Antarctic ecosystems. Two subfamilies, Pooideae and Danthonioideae, predominantly occupy the temperate zones of the Northern and Southern hemisphere, respectively (Kellogg, 2015). The Pooideae is one of the most species-rich Poaceae subfamilies, containing economically important cereal crops such as wheat (Triticum aestivum), barley (Hordeum vulgare), and oats (Avena sativa). Cold tolerance in the Pooideae involves cold acclimation (adaptation to freezing temperatures by pre-exposure to short periods of nonfreezing temperatures in the autumn) and vernalization (flowering after exposure to prolonged cold temperature in early spring). It has been hypothesized that cold adaptation was acquired before or during the early radiation of the Pooideae, which would reasonably result in cold tolerance for the whole subfamily (Zhong et al., 2018). In this issue of Plant Physiology, Schubert et al. (2019) show that gene expression in response to cold stress in diverse Pooideae species is largely species specific, supporting an alternative hypothesis that cold adaptation of the Pooideae occurred after species diversification. Figure 1. Open in new tabDownload slide Simplified phylogeny of the Poaceae. The Pooideae subfamily is shaded in blue, and the Bambusoideae, Oryzoideae and Pooideae (BOP) clade is shaded in yellow. PACMAD, Panicoideae, Arundinoideae, Chloridoideae, Micrairoideae, Aristidoideae and Danthonioideae. Figure 1. Open in new tabDownload slide Simplified phylogeny of the Poaceae. The Pooideae subfamily is shaded in blue, and the Bambusoideae, Oryzoideae and Pooideae (BOP) clade is shaded in yellow. PACMAD, Panicoideae, Arundinoideae, Chloridoideae, Micrairoideae, Aristidoideae and Danthonioideae. To test the evolutionary history of cold tolerance in response to cold acclimation, Schubert et al. (2019) selected diverse species spanning the Pooideae phylogeny, including a representative of the core Pooideae (H. vulgare), a member of the sister clade Brachypodium, and species from three early diverging lineages, Nardus stricta, Stipa lagascae, and Melica nutans (Fig. 1). All of the species show improved freezing tolerance after cold acclimation compared with plants without acclimation, demonstrating a conserved strategy for coping with freezing temperatures across the Pooideae. The authors further performed transcriptomic analysis in response to short-term (8 h) and long-term (4 and 9 weeks) cold treatment (6°C) in all five species. High confidence ortholog groups (HCOG) across the five species were used for cross-species comparisons. Among the differentially expressed HCOG between cold treatments and controls in at least one species, 50% are specific to a single species, and 16 out of 5577 (0.29%) HCOG are shared among all five species (Schubert et al., 2019). Similarly, in a transcriptomic study of cold response in Brachypodium distachyon, Nassella pulchra, and M. nutans, over 75% differentially expressed ortholog groups were specific to a single species (Zhong et al., 2018). These results suggest independent evolution of cold adaptation in different Pooideae lineages, which is consistent with a recent study dating the origin of the Pooideae to the late Cretaceous (66 million years ago), which would imply that the major clades within the Pooideae had already diversified during the global cooling of the Eocene-Oligocene period (34 million years ago; Schubert et al., 2018). Moreover, among the differentially expressed HCOG, positive selection of protein coding sequences has been observed at both the early and late splits of the Pooideae phylogeny, suggesting continuous evolution of cold adaptation (Schubert et al., 2019). Cold-responsive gene family expansion has played critical roles in cold adaptation during Pooideae radiation (Sandve and Fjellheim, 2010; Zhong et al., 2018). Because genes with complex evolutionary history were not included in HCOG, Schubert et al. (2019) specifically evaluated the evolution of five well-known cold-responsive gene families, including those encoding C-repeat binding factors, dehydrins, chloroplast-targeted cold-regulated proteins, ice recrystallization inhibition proteins, and fructosyl transferases. They found that evolution in response to cold acclimation of these gene families occurred at both early and late branches of the Pooideae phylogeny, reflecting diverse evolutionary histories of these genes. Together, the results of this study suggest that ancestral cold-responsive genes may have facilitated migration of the ancestors of the Pooideae into temperate regions, but cold tolerance mostly evolved after species diversification of the Pooideae, via regulation of cold-responsive gene expression, positive selection of coding sequences, and gene duplication and neo-functionalization. LITERATURE CITED Kellogg EA ( 2015 ) Poaceae . In K Kubitzki , ed, Flowering Plants. Monocots . Springer , Berlin , pp 1 – 416 Google Scholar Google Preview OpenURL Placeholder Text WorldCat COPAC Sandve SR , Fjellheim S ( 2010 ) Did gene family expansions during the Eocene-Oligocene boundary climate cooling play a role in Pooideae adaptation to cool climates? Mol Ecol 19 : 2075 – 2088 Google Scholar Crossref Search ADS PubMed WorldCat Schubert M , Marcussen T, Meseguer AS, Fjellheim S ( 2018 ) The grass subfamily Pooideae: Late Cretaceous origin and climate-driven Cenozoic diversification . bioRxiv 462440 , doi:10.1101/462440 Google Scholar OpenURL Placeholder Text WorldCat Schubert M , Groenvold L, Sandve SR, Hvidsten TR, Fjellheim S ( 2019 ) Evolution of cold acclimation and its role in niche transition in the temperate grass subfamily Pooideae . Plant Physiol . 180 : 404 – 419 Google Scholar Crossref Search ADS PubMed WorldCat Zhong J , Robbett M, Poire A, Preston JC ( 2018 ) Successive evolutionary steps drove Pooideae grasses from tropical to temperate regions . New Phytol 217 : 925 – 938 Google Scholar Crossref Search ADS PubMed WorldCat Author notes 1 Author for contact: [email protected]. 2 Senior author. www.plantphysiol.org/cgi/doi/10.1104/pp.19.00333 © 2019 American Society of Plant Biologists. All Rights Reserved. This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model)
doi: 10.1104/pp.19.00374pmid: 31053672
The transcription of many genes is regulated through alternative splicing, with over 60% of genes in Arabidopsis (Arabidopsis thaliana) producing more than one mRNA (Marquez et al., 2012). The most common forms of alternative splicing are intron retention and the use of alternative polyadenylation sites that result in transcripts of different length (Filichkin et al., 2010; Marquez et al., 2012). Alternative splicing can be regulated by environmental conditions that result in a switch between a transcript that is subjected to nonsense-mediated decay and a protein-coding transcript (Hartmann et al., 2016). In some instances, alternative splicing may be controlled through epigenetic modifications (Luco et al., 2011). Foremost among the epigenetic modifications in plants are DNA and histone methylation. While DNA and histone modifications are often associated with heterochromatin and untranscribed DNA, the spread of these marks into active genes can be detrimental, leading to a plethora of developmental abnormalities (Saze et al., 2008; Miura et al., 2009). With a reinforcing feedback loop between CHG methylation by the DNA methyl transferase CHROMOMETHYLASE3 and H3K9 methylation by the histone methyltransferases SU(VAR)3-9 HOMOLOG4 (SUVH4), SUVH5, and SUVH6, active demethylation is required to protect transcribed regions of the genome from accumulation of methylation. H3K9 demethylation is provided by the Jumonji C domain-containing protein INCREASE IN BONSAI METHYLATION1 (IBM1; Saze et al., 2008; Miura et al., 2009; Inagaki et al., 2010), while DNA demethylation is regulated by DNA glycosylases/lyases including REPRESSOR OF SILENCING1 (ROS1) and ROS3 (Agius et al., 2006; Zheng et al., 2008). IBM1 transcripts are alternatively spliced, with the two longer mRNAs that use a distal polyadenylation site providing functional protein (Rigal et al., 2012). Alternative splicing of IBM1 is regulated by a complex containing IBM2 and ENHANCED DOWNY MILDEW2 (EDM2). IBM2 has RNA recognition motifs and a Bromo-Adjacent Homology domain, which is commonly found in chromatin-associated proteins such as DNA methyltransferases. IBM2 is required for production of full-length transcripts from genes with intergenic heterochromatin (Saze et al., 2013). EDM2 contains zinc finger domains and is thought to act as an RNA methyltransferase (Tsuchiya and Eulgem, 2013; Lei et al., 2014). Other targets of the IBM2-EDM2 complex that also contain intronic heterochromatin include the resistance gene RECOGNITION OF PERONOSPORA PARASITICA7 (RPP7; AT1G11270) which encodes an F-box protein, and AT3G05410, which encodes a PSII reaction center PsbP family protein. In this issue of Plant Physiology, Nicolas Bouché’s group at INRA in Versailles further investigated IBM2 function in alternative polyadenylation site selection by identifying an interacting protein (Deremetz et al., 2019). To isolate IBM2 interactors, they performed a forward genetic screen using the ibm2-4 allele and selected for restoration of a wild-type phenotype. To exclude mutations that more broadly affected DNA or histone methylation of IBM1 targets, a secondary screen ascertained whether transcript levels of AT3G05410 were at least partially restored, as transcription of this gene is controlled by IBM2 but not IBM1. Through this approach, a new allele of FPA was identified that has intermediate AT3G05410 transcript levels. FPA was originally identified as a component of the autonomous flowering time pathway (Schomburg et al., 2001), a function that arises from its broader regulation of polyadenylation site selection (Sonmez et al., 2011; Duc et al., 2013). These functions of FPA in flowering time and RNA processing are shared by FCA, another RNA recognition motif-containing protein (Bäurle et al., 2007; Sonmez et al., 2011). Examining genome-wide trends in genic DNA methylation, the authors confirmed that CHG methylation specifically increases in genic regions in the absence of IBM2, a trend that is predominantly reversed in double fpa ibm2-4 mutants. How these changes in DNA methylation translate into transcript levels and selection of alternative polyadenylation sites was explored for a few select targets of IBM2, with the results indicating variable responses between genes. For IBM1, mutation of fpa alone or in an ibm2-4 background resulted in increased transcript levels with the distal polyadenylation site compared with the wild-type Columbia-0, with a concomitant decrease in transcripts terminating at the proximal polyadenylation site. These changes in transcript levels were the reverse of what was observed in ibm2-4 single mutants (Fig. 1). For RPP7, ibm2-4 decreased longer transcript levels; however, transcript levels were almost restored to normal in double fpa ibm2-4 mutants, with a commensurate increase in RPP7-mediated resistance to the Hiks1 strain of downy mildew (Hyaloperonospora parasitica). In contrast, longer transcript levels were only partially restored in fpa mutants for two other targets, AT3G05410 and AT1G11270. Overexpression of FPA repressed the long transcript levels of these targets to varying degrees. Thus, depending on the target locus, the activity of FPA is partially to wholly antagonistic to that of IBM2. Figure 1. Open in new tabDownload slide Effect of FPA and IBM2 mutations on short and long transcript accumulation from IBM1. In the presence of the IBM2/EDM2/AIPP1 complex, FPA may be excluded from binding to RNA transcripts at the proximal polyadenylation site. Levels of short and long mRNAs are indicated by the width of the lines below the gene model. Figure 1. Open in new tabDownload slide Effect of FPA and IBM2 mutations on short and long transcript accumulation from IBM1. In the presence of the IBM2/EDM2/AIPP1 complex, FPA may be excluded from binding to RNA transcripts at the proximal polyadenylation site. Levels of short and long mRNAs are indicated by the width of the lines below the gene model. Examining DNA methylation specifically at IBM1, CHG and CHH methylation over the heterochromatic region of the intron were reduced in fpa and fpa ibm2-4 mutants but were unaffected by ibm2-4 alone. Overexpression of IBM1 long transcripts in a wild-type background did not alter CHG methylation in the IBM1 intron; therefore, the reduction in methylation at IBM1 in fpa mutants is unlikely to be due to IBM1 reducing H3K9 methylation levels and hence limiting action of the DNA/H3K9 methylation feedback loop. Genome-wide, relatively few regions showed changes in DNA methylation in fpa mutants, with only 70 hypomethylated and 86 hypermethylated differentially methylated regions identified by whole-genome bisulfite sequencing. The majority of the hypermethylated regions were within genes, whereas most hypomethylated regions were associated with transposons or were intergenic. These results demonstrating that DNA methylation levels are affected at relatively few loci in fpa are consistent with previous reports that FPA does not have an extensive role in RNA-mediated chromatin silencing (Duc et al., 2013). Methylation was mostly restored to wild-type levels upon complementation of the fpa mutant, indicating that the effects of FPA mutation are transient and methylation can be restored upon reintroduction of FPA. Deremetz et al. (2019) also examined the function of FPA in regulating the transcription of genes that are IBM2 targets where the heterochromatic domain is nonintronic. They identified 18 transposons that were down-regulated in mutants of components of the IBM2-EDM2 complex by using previously published RNA sequencing data (Duan et al., 2017). In two cases, these transposons are nonintronic targets that were down-regulated in expression in ibm2 and fpa ibm2 mutants but not in fpa mutants, indicating that FPA and IBM2 only act antagonistically at genes with intronic heterochromatin. Together, the results by Deremetz et al. (2019) demonstrate that FPA affects DNA methylation of a relatively small number of targets, confirming a limited functional role in polyadenylation site usage. At select loci, IBM2 and FPA act antagonistically to control transcription. These loci include key genes such as IBM1, which encodes a protein involved in H3K9 demethylation, and RPP7, which functions in race-specific disease resistance. Given previous links between FPA and FCA in RNA processing, it will be informative to know whether FCA also functions at IBM2 targets and whether there is direct interaction between the IBM2/EDM2 complex and FPA/FCA. Also, given the limited number of loci affected by FPA, identification of other proteins that interact similarly with IBM2 at further loci where methylation accumulates in ibm2 mutants will be important. LITERATURE CITED Agius F , Kapoor A, Zhu JK ( 2006 ) Role of the Arabidopsis DNA glycosylase/lyase ROS1 in active DNA demethylation . Proc Natl Acad Sci USA 103 : 11796 – 11801 Google Scholar Crossref Search ADS PubMed WorldCat Bäurle I , Smith L, Baulcombe DC, Dean C ( 2007 ) Widespread role for the flowering-time regulators FCA and FPA in RNA-mediated chromatin silencing . Science 318 : 109 – 112 Google Scholar Crossref Search ADS PubMed WorldCat Deremetz A , Le Roux C, Idir Y, Brousse C, Agorio A, Gy I, Parker JE, Bouché N ( 2019 ) Antagonistic actions of FPA and IBM2 regulate transcript processing from genes containing heterochromatin . Plant Physiol 180 : 392 – 403 Google Scholar Crossref Search ADS PubMed WorldCat Duan CG , Wang X, Zhang L, Xiong X, Zhang Z, Tang K, Pan L, Hsu CC, Xu H, Tao WA, et al. ( 2017 ) A protein complex regulates RNA processing of intronic heterochromatin-containing genes in Arabidopsis . Proc Natl Acad Sci USA 114 : E7377 – E7384 Google Scholar Crossref Search ADS PubMed WorldCat Duc C , Sherstnev A, Cole C, Barton GJ, Simpson GG ( 2013 ) Transcription termination and chimeric RNA formation controlled by Arabidopsis thaliana FPA . PLoS Genet 9 : e1003867 Google Scholar Crossref Search ADS PubMed WorldCat Filichkin SA , Priest HD, Givan SA, Shen R, Bryant DW, Fox SE, Wong WK, Mockler TC ( 2010 ) Genome-wide mapping of alternative splicing in Arabidopsis thaliana . Genome Res 20 : 45 – 58 Google Scholar Crossref Search ADS PubMed WorldCat Hartmann L , Drewe-Boß P, Wießner T, Wagner G, Geue S, Lee HC, Obermüller DM, Kahles A, Behr J, Sinz FH, et al. ( 2016 ) Alternative splicing substantially diversifies the transcriptome during early photomorphogenesis and correlates with the energy availability in Arabidopsis . Plant Cell 28 : 2715 – 2734 Google Scholar Crossref Search ADS PubMed WorldCat Inagaki S , Miura-Kamio A, Nakamura Y, Lu F, Cui X, Cao X, Kimura H, Saze H, Kakutani T ( 2010 ) Autocatalytic differentiation of epigenetic modifications within the Arabidopsis genome . EMBO J 29 : 3496 – 3506 Google Scholar Crossref Search ADS PubMed WorldCat Lei M , La H, Lu K, Wang P, Miki D, Ren Z, Duan CG, Wang X, Tang K, Zeng L, et al. ( 2014 ) Arabidopsis EDM2 promotes IBM1 distal polyadenylation and regulates genome DNA methylation patterns . Proc Natl Acad Sci USA 111 : 527 – 532 Google Scholar Crossref Search ADS PubMed WorldCat Luco RF , Allo M, Schor IE, Kornblihtt AR, Misteli T ( 2011 ) Epigenetics in alternative pre-mRNA splicing . Cell 144 : 16 – 26 Google Scholar Crossref Search ADS PubMed WorldCat Marquez Y , Brown JWS, Simpson C, Barta A, Kalyna M ( 2012 ) Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis . Genome Res 22 : 1184 – 1195 Google Scholar Crossref Search ADS PubMed WorldCat Miura A , Nakamura M, Inagaki S, Kobayashi A, Saze H, Kakutani T ( 2009 ) An Arabidopsis jmjC domain protein protects transcribed genes from DNA methylation at CHG sites . EMBO J 28 : 1078 – 1086 Google Scholar Crossref Search ADS PubMed WorldCat Rigal M , Kevei Z, Pélissier T, Mathieu O ( 2012 ) DNA methylation in an intron of the IBM1 histone demethylase gene stabilizes chromatin modification patterns . EMBO J 31 : 2981 – 2993 Google Scholar Crossref Search ADS PubMed WorldCat Saze H , Shiraishi A, Miura A, Kakutani T ( 2008 ) Control of genic DNA methylation by a jmjC domain-containing protein in Arabidopsis thaliana . Science 319 : 462 – 465 Google Scholar Crossref Search ADS PubMed WorldCat Saze H , Kitayama J, Takashima K, Miura S, Harukawa Y, Ito T, Kakutani T ( 2013 ) Mechanism for full-length RNA processing of Arabidopsis genes containing intragenic heterochromatin . Nat Commun 4 : 2301 Google Scholar Crossref Search ADS PubMed WorldCat Schomburg FM , Patton DA, Meinke DW, Amasino RM ( 2001 ) FPA, a gene involved in floral induction in Arabidopsis, encodes a protein containing RNA-recognition motifs . Plant Cell 13 : 1427 – 1436 Google Scholar Crossref Search ADS PubMed WorldCat Sonmez C , Bäurle I, Magusin A, Dreos R, Laubinger S, Weigel D, Dean C ( 2011 ) RNA 3′ processing functions of Arabidopsis FCA and FPA limit intergenic transcription . Proc Natl Acad Sci USA 108 : 8508 – 8513 Google Scholar Crossref Search ADS PubMed WorldCat Tsuchiya T , Eulgem T ( 2013 ) Mutations in EDM2 selectively affect silencing states of transposons and induce plant developmental plasticity . Sci Rep 3 : 1701 Google Scholar Crossref Search ADS PubMed WorldCat Zheng X , Pontes O, Zhu J, Miki D, Zhang F, Li WX, Iida K, Kapoor A, Pikaard CS, Zhu JK ( 2008 ) ROS3 is an RNA-binding protein required for DNA demethylation in Arabidopsis . Nature 455 : 1259 – 1262 Google Scholar Crossref Search ADS PubMed WorldCat Author notes 1 Author for contact: [email protected]. 2 Senior author. www.plantphysiol.org/cgi/doi/10.1104/pp.19.00374 © 2019 American Society of Plant Biologists. All Rights Reserved. © The Author(s) 2019. Published by Oxford University Press on behalf of American Society of Plant Biologists. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
doi: 10.1104/pp.19.00296pmid: 31053673
One goal of synthetic biology is to build artificial pathways for bioengineering of high-value compounds. To this end, pathways are not only split into individual reactions, but the expression of one enzyme is also split into individual parts (e.g. promotors, ribosomal binding sites, coding sequences, and terminators) that are used to optimize flux through the whole pathway. Testing a multitude of combinations of these parts for one step of a pathway would involve numerous cloning steps, which is further increased if whole pathways are built. Traditional cloning methods are too inefficient and laborious for these synthetic biology approaches. To facilitate more efficient high-throughput cloning, the Golden Gate cloning system was developed (Engler et al., 2008; Weber et al., 2011; Fig. 1). This system relies on type IIS restriction enzymes that cut outside of their recognition sequence. These enzymes can generate a high degree of specificity for a ligation reaction from one digestion reaction with 256 theoretically possible four-nucleotide overhangs. This not only allows these enzymes to be used in a high-throughput setting but also facilitates the assembly of multiple parts into one construct with the correct order and orientation, when the same four-nucleotide overhangs are used at the end of one part and the beginning of the next. This allows for easy assembly of multiple combinations, if alternate parts have the same four-nucleotide overhang. Another advantage of the system is that although it was developed by a company (Icon Genetics), it does not contain proprietary tools and reagents, and even more novel parts have been developed and made freely available since publication of the initial tool kit. A Golden Gate-based tool kit for plants, containing vectors and 96 standardized parts, including promotors, terminators, and tags, was subsequently developed (Engler et al., 2014). Figure 1. Open in new tabDownload slide Golden Gate assembly standard. Parts are cloned into Level 0 vectors and are the basic units of genetic grammar; for simplicity, only three parts are shown. Parts are assembled into transcription units in Level 1 vectors, which can be further assembled into larger multiple-transcription unit constructs in Level M and P vectors. The increase of the assembly between Level M and P vectors can be repeated multiple times and is only limited by the overall size of the resulting construct. Figure 1. Open in new tabDownload slide Golden Gate assembly standard. Parts are cloned into Level 0 vectors and are the basic units of genetic grammar; for simplicity, only three parts are shown. Parts are assembled into transcription units in Level 1 vectors, which can be further assembled into larger multiple-transcription unit constructs in Level M and P vectors. The increase of the assembly between Level M and P vectors can be repeated multiple times and is only limited by the overall size of the resulting construct. In this issue of Plant Physiology, Vasudevan et al. (2019) describe the modification of the plant Golden Gate tool kit for use with cyanobacteria. Cyanobacteria are photosynthetically active bacteria and, through an endosymbiosis event, are the most likely progenitor for the plant chloroplast. For this reason, they can help us further understand photosynthesis in plants. Additionally, cyanobacteria have great potential as a photoautotrophic bioengineering platform, and the Golden Gate tool kit will certainly benefit the development of cyanobacteria strains that produce high-value compounds. The authors established vectors for assembly of components for transient expression or integration into the genome. They also developed a multitude of individual parts such as 21 terminators and 12 native (Synechocystis sp. PCC 6803) and 33 heterologous/synthetic promotors, which were tested in the cyanobacteria model organisms Synechocystis sp. PCC 6803 and Synechococcus elongatus UTEX 2973. This large suite of promotors allows the user to choose a desired expression strength for a construct. Moreover, as random integration of constructs into the genome can lead to growth disturbances, linker sequences were developed that target integration of the constructs to neutral sites that have minimal to no consequences for the organism. Finally, the CRISPR/Cas9 system is included as a part that can either be used to generate knockout mutants or introduce specific mutations with guide RNAs, although the latter was not demonstrated in this publication. The development of this CyanoGate will hopefully not only further our knowledge of how photosynthesis in cyanobacteria compares with photosynthesis in plants but will also help us establish cyanobacteria as a photoautotrophic production host for high-value compounds that benefit human health and development without the harmful by-products of chemical synthesis. LITERATURE CITED Engler C , Kandzia R, Marillonnet S ( 2008 ) A one pot, one step, precision cloning method with high throughput capability . PLoS ONE 3 : e3647 Google Scholar Crossref Search ADS PubMed WorldCat Engler C , Youles M, Gruetzner R, Ehnert TM, Werner S, Jones JDG, Patron NJ, Marillonnet S ( 2014 ) A Golden Gate modular cloning toolbox for plants . ACS Synth Biol 3 : 839 – 843 Google Scholar Crossref Search ADS PubMed WorldCat Vasudevan R , Gale GAR, Schiavon AA, Puzorjov A, Malin J, Gillespie MD, Vavitsas K, Zulkower V, Wang B, Howe CJ, et al. ( 2019 ) CyanoGate: A modular cloning suite for engineering cyanobacteria based on the plant MoClo syntax . Plant Physiol 180 : 39 – 55 Google Scholar Crossref Search ADS PubMed WorldCat Weber E , Engler C, Gruetzner R, Werner S, Marillonnet S ( 2011 ) A modular cloning system for standardized assembly of multigene constructs . PLoS ONE 6 : e16765 Google Scholar Crossref Search ADS PubMed WorldCat Author notes 1 Author for contact: [email protected]. 2 Senior author. www.plantphysiol.org/cgi/doi/10.1104/pp.19.00296 © 2019 American Society of Plant Biologists. All Rights Reserved. This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model)
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