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Zeng, Tian; Holmer, Rens; Hontelez, Jan; Lintel‐Hekkert, Bas; Marufu, Lucky; Zeeuw, Thijs; Wu, Fangyuan; Schijlen, Elio; Bisseling, Ton; Limpens, Erik
doi: 10.1111/tpj.13908pmid: 29570877
Arbuscular mycorrhizal fungi form the most wide‐spread endosymbiosis with plants. There is very little host specificity in this interaction, however host preferences as well as varying symbiotic efficiencies have been observed. We hypothesize that secreted proteins (SPs) may act as fungal effectors to control symbiotic efficiency in a host‐dependent manner. Therefore, we studied whether arbuscular mycorrhizal (AM) fungi adjust their secretome in a host‐ and stage‐dependent manner to contribute to their extremely wide host range. We investigated the expression of SP‐encoding genes of Rhizophagus irregularis in three evolutionary distantly related plant species, Medicago truncatula, Nicotiana benthamiana and Allium schoenoprasum. In addition we used laser microdissection in combination with RNA‐seq to study SP expression at different stages of the interaction in Medicago. Our data indicate that most expressed SPs show roughly equal expression levels in the interaction with all three host plants. In addition, a subset shows significant differential expression depending on the host plant. Furthermore, SP expression is controlled locally in the hyphal network in response to host‐dependent cues. Overall, this study presents a comprehensive analysis of the R. irregularis secretome, which now offers a solid basis to direct functional studies on the role of fungal SPs in AM symbiosis.
Liu, Tzu‐Yin; Chou, Wen‐Chun; Chen, Wei‐Yuan; Chu, Ching‐Yi; Dai, Chen‐Yi; Wu, Pei‐Yu
doi: 10.1111/tpj.13874pmid: 29451720
Despite the great interest in identifying protein–protein interactions (PPIs) in biological systems, only a few attempts have been made at large‐scale PPI screening in planta. Unlike biochemical assays, bimolecular fluorescence complementation allows visualization of transient and weak PPIs in vivo at subcellular resolution. However, when the non‐fluorescent fragments are highly expressed, spontaneous and irreversible self‐assembly of the split halves can easily generate false positives. The recently developed tripartite split‐GFP system was shown to be a reliable PPI reporter in mammalian and yeast cells. In this study, we adapted this methodology, in combination with the β‐estradiol‐inducible expression cassette, for the detection of membrane PPIs in planta. Using a transient expression assay by agroinfiltration of Nicotiana benthamiana leaves, we demonstrate the utility of the tripartite split‐GFP association in plant cells and affirm that the tripartite split‐GFP system yields no spurious background signal even with abundant fusion proteins readily accessible to the compartments of interaction. By validating a few of the Arabidopsis PPIs, including the membrane PPIs implicated in phosphate homeostasis, we proved the fidelity of this assay for detection of PPIs in various cellular compartments in planta. Moreover, the technique combining the tripartite split‐GFP association and dual‐intein‐mediated cleavage of polyprotein precursor is feasible in stably transformed Arabidopsis plants. Our results provide a proof‐of‐concept implementation of the tripartite split‐GFP system as a potential tool for membrane PPI screens in planta.
Ogita, Nobuo; Okushima, Yoko; Tokizawa, Mutsutomo; Yamamoto, Yoshiharu Y.; Tanaka, Maho; Seki, Motoaki; Makita, Yuko; Matsui, Minami; Okamoto‐Yoshiyama, Kaoru; Sakamoto, Tomoaki; Kurata, Tetsuya; Hiruma, Kei; Saijo, Yusuke; Takahashi, Naoki; Umeda, Masaaki
Mao, Chanjuan; Ding, Jialin; Zhang, Bin; Xi, Dandan; Ming, Feng
doi: 10.1111/tpj.13867pmid: 29436050
Plant development and adaptation to environmental stresses are intimately associated with programmed cell death (PCD). Although some of the mechanisms regulating PCD [e.g., accumulation of reactive oxygen species (ROS)] are common among responses to different abiotic stresses, the pathways mediating salt‐induced PCD remain largely uncharacterized. Here we report that overexpression of OsNAC2, which encodes a plant‐specific transcription factor, promotes salt‐induced cell death accompanied by the loss of plasma membrane integrity, nuclear DNA fragmentation, and changes to caspase‐like activity. In OsNAC2‐knockdown lines, cell death was markedly decreased in response to severe salt stress. Additionally, OsNAC2 expression was enhanced in rice seedlings exposed to a high NaCl concentration. Moreover, the results of quantitative real‐time PCR, chromatin immunoprecipitation, dual‐luciferase, and yeast one‐hybrid assays indicated that OsNAC2 targeted genes that encoded an ROS scavenger (OsCOX11) and a caspase‐like protease (OsAP37). Furthermore, K+‐efflux channels (OsGORK and OsSKOR) were clearly activated by OsNAC2. Overall, our results suggested that OsNAC2 accelerates NaCl‐induced PCD and provide new insights into the mechanisms that affect ROS accumulation, plant caspase‐like activity, and K+ efflux.
Carqueijeiro, Inês; Dugé de Bernonville, Thomas; Lanoue, Arnaud; Dang, Thu‐Thuy; Teijaro, Christiana N; Paetz, Christian; Billet, Kevin; Mosquera, Angela; Oudin, Audrey; Besseau, Sébastien; Papon, Nicolas; Glévarec, Gaëlle; Atehortùa, Lucía; Clastre, Marc;
Herbst, Josephine; Girke, Annabel; Hajirezaei, Mohammad Reza; Hanke, Guy; Grimm, Bernhard
doi: 10.1111/tpj.13869pmid: 29443418
Chlorophyll is synthesized from activated glutamate in the tetrapyrrole biosynthesis pathway through at least 20 different enzymatic reactions. Among these, the MgProto monomethylester (MgProtoME) cyclase catalyzes the formation of a fifth isocyclic ring to tetrapyrroles to form protochlorophyllide. The enzyme consists of two proteins. The CHL27 protein is proposed to be the catalytic component, while LCAA/YCF54 likely acts as a scaffolding factor. In comparison to other reactions of chlorophyll biosynthesis, this enzymatic step lacks clear elucidation and it is hardly understood, how electrons are delivered for the NADPH‐dependent cyclization reaction. The present study intends to elucidate more precisely the role of LCAA/YCF54. Transgenic Arabidopsis lines with inactivated and overexpressed YCF54 reveal the mutual stability of YCF54 and CHL27. Among the YCF54‐interacting proteins, the plastidal ferredoxin‐NADPH reductase (FNR) was identified. We showed in N. tabacum and A. thaliana that a deficit of FNR1 or YCF54 caused MgProtoME accumulation, the substrate of the cyclase, and destabilization of the cyclase subunits. It is proposed that FNR serves as a potential donor for electrons required in the cyclase reaction and connects chlorophyll synthesis with photosynthetic activity.
Ben‐Tov, Daniela; Idan‐Molakandov, Anat; Hugger, Anat; Ben‐Shlush, Ilan; Günl, Markus; Yang, Bo; Usadel, Björn; Harpaz‐Saad, Smadar
doi: 10.1111/tpj.13871pmid: 29446495
The production of hydrophilic mucilage along the course of seed coat epidermal cell differentiation is a common adaptation in angiosperms. Previous studies have identified COBRA‐LIKE 2 (COBL2), a member of the COBRA‐LIKE gene family, as a novel component required for crystalline cellulose deposition in seed coat epidermal cells. In recent years, Arabidopsis seed coat epidermal cells (SCEs), also called mucilage secretory cells, have emerged as a powerful model system for the study of plant cell wall components biosynthesis, secretion, assembly and de muro modification. Despite accumulating data, the molecular mechanism of COBL function remains largely unknown. In the current research, we utilized genetic interactions to study the role of COBL2 as part of the protein network required for seed mucilage production. Using correlative phenotyping of structural and biochemical characteristics, unique features of the cobl2 extruded mucilage are revealed, including: ‘unraveled’ ray morphology, loss of primary cell wall ‘pyramidal’ organization, reduced Ruthenium red staining intensity of the adherent mucilage layer, and increased levels of the monosaccharides arabinose and galactose. Examination of the cobl2cesa5 double mutant provides insight into the interface between COBL function and cellulose deposition. Additionally, genetic interactions between cobl2 and fei1fei2 as well as between each of these mutants to mucilage‐modified 2 (mum2) suggest that COBL2 functions independently of the FEI‐SOS pathway. Altogether, the presented data place COBL2 within the complex protein network required for cell wall deposition in the context of seed mucilage and introduce new methodology expending the seed mucilage phenotyping toolbox.
Li, Ran; Fu, Daqi; Zhu, Benzhong; Luo, Yunbo; Zhu, Hongliang
doi: 10.1111/tpj.13872pmid: 29446503
With the development of high‐throughput sequencing, many long non‐coding RNAs (lncRNAs) have been found to play important roles in diverse biological processes. However, the biological functions of most plant lncRNAs are still unknown. We have previously discovered a tomato ripening‐related lncRNA, lncRNA1459. Here, we cloned the full‐length lncRNA1459, giving two transcript isoforms. In addition, lncRNA1459 exhibited a specific location in the nucleus. Furthermore, in order to fully identify the function of lncRNA1459 in tomato ripening, loss‐of‐function mutants of lncRNA1459 were developed using clustered regularly interspaced short palindromic repeats (CRISPR)/‐associated protein 9 (Cas9)‐induced genome editing technology. Compared with wild‐type fruits, the tomato ripening process was significantly repressed in lncRNA1459 mutants. Ethylene production and lycopene accumulation were largely repressed in lncRNA1459 mutants. Additionally, genes related to ethylene and carotenoid biosynthesis were distinctly downregulated in lncRNA1459 mutants compared with wild‐type fruits. Moreover, expression of numerous ripening‐related genes was changed significantly when lncRNA1459 was knocked out. Expression of potential tomato ripening‐related lncRNAs was also specifically changed after knocking out lncRNA1459. Taken together, these results provide insight into the role of lncRNA1459 in tomato fruit ripening.
Showing 1 to 10 of 14 Articles
doi: 10.1111/tpj.13866pmid: 29430765
In mammalian cells, the transcription factor p53 plays a crucial role in transmitting DNA damage signals to maintain genome integrity. However, in plants, orthologous genes for p53 and checkpoint proteins are absent. Instead, the plant‐specific transcription factor SUPPRESSOR OF GAMMA RESPONSE 1 (SOG1) controls most of the genes induced by gamma irradiation and promotes DNA repair, cell cycle arrest, and stem cell death. To date, the genes directly controlled by SOG1 remain largely unknown, limiting the understanding of DNA damage signaling in plants. Here, we conducted a microarray analysis and chromatin immunoprecipitation (ChIP)‐sequencing, and identified 146 Arabidopsis genes as direct targets of SOG1. By using ChIP‐sequencing data, we extracted the palindromic motif [CTT(N)7AAG] as a consensus SOG1‐binding sequence, which mediates target gene induction in response to DNA damage. Furthermore, DNA damage‐triggered phosphorylation of SOG1 is required for efficient binding to the SOG1‐binding sequence. Comparison between SOG1 and p53 target genes showed that both transcription factors control genes responsible for cell cycle regulation, such as CDK inhibitors, and DNA repair, whereas SOG1 preferentially targets genes involved in homologous recombination. We also found that defense‐related genes were enriched in the SOG1 target genes. Consistent with this finding, SOG1 is required for resistance against the hemi‐biotrophic fungus Colletotrichum higginsianum, suggesting that SOG1 has a unique function in controlling the immune response.
doi: 10.1111/tpj.13868pmid: 29438577
While the characterization of the biosynthetic pathway of monoterpene indole alkaloids (MIAs) in leaves of Catharanthus roseus is now reaching completion, only two enzymes from the root counterpart dedicated to tabersonine metabolism have been identified to date, namely tabersonine 19‐hydroxylase (T19H) and minovincine 19‐O‐acetyltransferase (MAT). Albeit the recombinant MAT catalyzes MIA acetylation at low efficiency in vitro, we demonstrated that MAT was inactive when expressed in yeast and in planta, suggesting an alternative function for this enzyme. Therefore, through transcriptomic analysis of periwinkle adventitious roots, several other BAHD acyltransferase candidates were identified based on the correlation of their expression profile with T19H and found to localize in small genomic clusters. Only one, named tabersonine derivative 19‐O‐acetyltransferase (TAT) was able to acetylate the 19‐hydroxytabersonine derivatives from roots, such as minovincinine and hörhammericine, following expression in yeast. Kinetic studies also showed that the recombinant TAT was specific for root MIAs and displayed an up to 200‐fold higher catalytic efficiency than MAT. In addition, gene expression analysis, protein subcellular localization and heterologous expression in Nicotiana benthamiana were in agreement with the prominent role of TAT in acetylation of root‐specific MIAs, thereby redefining the molecular determinants of the root MIA biosynthetic pathway. Finally, identification of TAT provided a convenient tool for metabolic engineering of MIAs in yeast enabling efficiently mixing different biosynthetic modules spatially separated in the whole plant. This combinatorial synthesis associating several enzymes from Catharanthus roseus resulted in the conversion of tabersonine in tailor‐made MIAs bearing both leaf and root‐type decorations.