CLAVATA3/EMBRYO SURROUNDING REGION-related Research Articles

The CLE33 peptide represses phloem differentiation via autocrine and paracrine signaling in Arabidopsis

Plant meristems require a constant supply of photoassimilates and hormones to the dividing meristematic cells. In the growing root, such supply is delivered by protophloem sieve elements. Due to its preeminent function for the root apical meristem, protophloem is the first tissue to differentiate. This process is regulated by a genetic circuit involving in one side the positive regulators DOF transcription factors, OCTOPUS (OPS) and BREVIX RADIX (BRX), and in the other side the negative regulators CLAVATA3/EMBRYO SURROUNDING REGION RELATED (CLE) peptides and their cognate receptors BARELY ANY MERISTEM (BAM) receptor-like kinases. brx and ops mutants harbor a discontinuous protophloem that can be fully rescued by mutation in BAM3, but is only partially rescued when all three known phloem-specific CLE genes, CLE25/26/45 are simultaneously mutated. Here we identify a CLE gene closely related to CLE45, named CLE33. We show that double mutant cle33cle45 fully suppresses brx and ops protophloem phenotype. CLE33 orthologs are found in basal angiosperms, monocots, and eudicots, and the gene duplication which gave rise to CLE45 in Arabidopsis and other Brassicaceae appears to be a recent event. We thus discovered previously unidentified Arabidopsis CLE gene that is an essential player in protophloem formation.

Plants Recruit Peptides and Micro RNAs to Regulate Nutrient Acquisition from Soil and Symbiosis.

Plants engage in symbiotic relationships with soil microorganisms to overcome nutrient limitations in their environment. Among the best studied endosymbiotic interactions in plants are those with arbuscular mycorrhizal (AM) fungi and N-fixing bacteria called rhizobia. The mechanisms regulating plant nutrient homeostasis and acquisition involve small mobile molecules such as peptides and micro RNAs (miRNAs). A large number of CLE (CLAVATA3/EMBRYO SURROUNDING REGION-RELATED) and CEP (C-TERMINALLY ENCODED PEPTIDE) peptide hormones as well as certain miRNAs have been reported to differentially respond to the availability of essential nutrients such as nitrogen (N) and phosphorus (P). Interestingly, a partially overlapping pool of these molecules is involved in plant responses to root colonization by rhizobia and AM fungi, as well as mineral nutrition. The crosstalk between root endosymbiosis and nutrient availability has been subject of intense investigations, and new insights in locally or systemically mobile molecules in nutrient- as well as symbiosis-related signaling continue to arise. Focusing on the key roles of peptides and miRNAs, we review the mechanisms that shape plant responses to nutrient limitation and regulate the establishment of symbiotic associations with beneficial soil microorganisms.

Characterization and response of two potato receptor-like kinases to cyst nematode infection

ABSTRACT Plant-parasitic cyst nematodes (Heterodera and Globodera spp.) secrete CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) effector proteins, which act as ligand mimics of plant CLE peptides to promote successful nematode infection. Previous studies of the Arabidopsis-beet cyst nematode (BCN; H. schachtii) pathosystem showed that Arabidopsis CLE receptors including CLAVATA1 (CLV1), CLV2, and RECEPTOR-LIKE PROTEIN KINASE 2 (RPK2) are required for BCN CLE signaling. Studies further revealed that nematode CLE signaling through GmCLV2 and StCLV2, an Arabidopsis CLV2 orthologue from soybean (Glycines max) and potato (Solanum tuberosum), respectively, is required for the soybean cyst nematode (SCN; H. glycines) and the potato cyst nematode (PCN; G. rostochiensis) to induce disease in their respective host plant. In this study, we identified and characterized two additional potato receptors, StRPK2 and StCLV1, homologues of Arabidopsis RPK2 and CLV1, for a role in PCN parasitism. Using promoter-reporter lines we showed that both StRPK2 and StCLV1 are expressed in the potato root but vary in their spatial expression patterns. Interestingly, StRPK2 but not StCLV1 was found to be expressed and upregulated at PCN infection sites. Nematode infection assays on StRPK2-knockdown lines revealed a decrease in nematode infection. Collectively, our results suggest that parallel CLE signaling pathways involving StCLV2 and StRPK2 are important for PCN parasitism and that manipulation of nematode CLE signaling may represent a viable means to engineer nematode resistance in crop plants including potato.

Genome-wide identification and comparative analysis of CLE family in rapeseed and its diploid progenitors.

Crop genomics and breeding CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) proteins belong to a small peptide family in plants. During plant development, CLE gene family members play a pivotal role in regulating cell-to-cell communication and stem cell maintenance. However, the evolutionary process and functional importance of CLEs are unclear in Brassicaceae. In this study, a total of 70 BnCLEs were identified in Brassica napus (2n = 4x = 38, AnCn): 32 from the An subgenome, 36 from the Cn subgenome, and 2 from the unanchored subgenome. Meanwhile, 29 BrCLE and 32 BoCLE genes were explored in Brassica rapa (2n = 2x = 20, Ar) and Brassica oleracea (2n = 2x = 18, Co). Phylogenetic analysis revealed that 163 CLEs derived from three Brassica species and Arabidopsis thaliana can be divided into seven subfamilies. Homology and synteny analyses indicated whole-genome triplication (WGT) and segmental duplication may be the major contributors to the expansion of CLE family. In addition, RNA-seq and qPCR analysis indicated that 19 and 16 BnCLEs were more highly expressed in immature seeds and roots than in other tissues. Some CLE gene pairs exhibited different expression patterns in the same tissue, which indicated possible functional divergence. Furthermore, genetic variations and regional association mapping analysis indicated that 12 BnCLEs were potential genes for regulating important agronomic traits. This study provided valuable information to understand the molecular evolution and biological function of CLEs in B. napus and its diploid progenitors, which will be helpful for genetic improvement of high-yield breeding in B. napus.

Spatial range, temporal span, and promiscuity of CLE-RLK signaling.

CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) signaling through receptor-like kinases (RLKs) regulates developmental transitions and responses to biotic and abiotic inputs by communicating the physiological state of cells and tissues. CLE peptides have varying signaling ranges, which can be defined as the distance between the source, i.e., the cells or tissue that secrete the peptide, and their destination, i.e., cells or tissue where the RLKs that bind the peptide and/or respond are expressed. Case-by-case analysis substantiates that CLE signaling is predominantly autocrine or paracrine, and rarely endocrine. Furthermore, upon CLE reception, the ensuing signaling responses extend from cellular to tissue, organ and whole organism level as the downstream signal gets amplified. CLE-RLK-mediated effects on tissue proliferation and differentiation, or on subsequent primordia and organ development have been widely studied. However, studying how CLE-RLK regulates different stages of proliferation and differentiation at cellular level can offer additional insights into these processes. Notably, CLE-RLK signaling also mediates diverse non-developmental effects, which are less often observed; however, this could be due to biased experimental approaches. In general, CLEs and RLKs, owing to the sequence or structural similarity, are prone to promiscuous interactions at least under experimental conditions in which they are studied. Importantly, there are regulatory mechanisms that suppress CLE-RLK cross-talk in vivo, thereby eliminating the pressure for co-evolving binding specificity. Alternatively, promiscuity in signaling may also offer evolutionary advantages and enable different CLEs to work in combination to activate or switch off different RLK signaling pathways.

Identification of Diverse Stress-Responsive Xylem Sap Peptides in Soybean.

Increasing evidence has revealed that plant secretory peptides are involved in the long-distance signaling pathways that help to regulate plant development and signal stress responses. In this study, we purified small peptides from soybean (Glycine max) xylem sap via o-chlorophenol extraction and conducted an in-depth peptidomic analysis using a mass spectrometry (MS) and bioinformatics approach. We successfully identified 14 post-translationally modified peptide groups belonging to the peptide families CEP (C-terminally encoded peptides), CLE (CLAVATA3/embryo surrounding region-related), PSY (plant peptides containing tyrosine sulfation), and XAP (xylem sap-associated peptides). Quantitative PCR (qPCR) analysis showed unique tissue expression patterns among the peptide-encoding genes. Further qPCR analysis of some of the peptide-encoding genes showed differential stress-response profiles toward various abiotic stress factors. Targeted MS-based quantification of the nitrogen deficiency-responsive peptides, GmXAP6a and GmCEP-XSP1, demonstrated upregulation of peptide translocation in xylem sap under nitrogen-deficiency stress. Quantitative proteomic analysis of GmCEP-XSP1 overexpression in hairy soybean roots revealed that GmCEP-XSP1 significantly impacts stress response-related proteins. This study provides new insights that root-to-shoot peptide signaling plays important roles in regulating plant stress-response mechanisms.

Control of Root Stem Cell Differentiation and Lateral Root Emergence by CLE16/17 Peptides in Arabidopsis.

Secreted peptide-mediated cell-to-cell communication plays a crucial role in the development of multicellular organisms. A large number of secreted peptides have been predicated by bioinformatic approaches in plants. However, only a few of them have been functionally characterized. In this study, we show that two CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) peptides CLE16/17 are required for both stem cell differentiation and lateral root (LR) emergence in Arabidopsis. We further demonstrate that the CLE16/17 peptides act through the CLAVATA1-ARABIDOPSIS CRINKLY4 (CLV1-ACR4) protein kinase complex in columella stem cell (CSC) differentiation, but not in LR emergence. Furthermore, we show that CLE16/17 promote LR emergence probably via activating the expression of HAESA/HAESA-LIKE2 (HAE/HSL2) required for cell wall remodeling. Collectively, our results reveal a CLV1-ACR4-dependent and -independent dual-function of the CLE16/17 peptides in root development.

Peptide Effectors in Phytonematode Parasitism and Beyond

Peptide signaling is an emerging paradigm in molecular plant-microbe interactions with vast implications for our understanding of plant-nematode interactions and beyond. Plant-like peptide hormones, first discovered in cyst nematodes, are now recognized as an important class of peptide effectors mediating several different types of pathogenic and symbiotic interactions. Here, we summarize what has been learned about nematode-secreted CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) peptide effectors since the last comprehensive review on this topic a decade ago. We also highlight new discoveries of a diverse array of peptide effectors that go beyond the CLE peptide effector family in not only phytonematodes but in organisms beyond the phylum Nematoda.

Root-specific CLE3 expression is required for WRKY33 activation in Arabidopsis shoots.

This study focused on the role of CLE1-7 peptides as defense mediators, and showed that root-expressed CLE3 functions as a systemic signal to regulate defense-related gene expression in shoots. In the natural environment, plants employ diverse signaling molecules including peptides to defend themselves against various pathogen attacks. In this study, we investigated whether CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) genes (CLE1-7) respond to biotic stimuli. CLE3 showed significant up-regulation upon treatment with flg22, Pep2, and salicylic acid (SA). Quantitative real-time PCR (qRT-PCR) analysis revealed that CLE3 expression is regulated by the NON-EXPRESSOR OF PR GENES1 (NPR1)-dependent SA signaling and flg22-FLAGELLIN-SENSITIVE 2 (FLS2) signaling pathways. We demonstrated that SA-induced up-regulation of CLE3 in roots was required for activation of WRKY33, a gene involved in the regulation of systemic acquired resistance (SAR), in shoots, suggesting that CLE3 functions as a root-derived signal that regulates the expression of defense-related genes in shoots. Microarray analysis of transgenic Arabidopsis lines overexpressing CLE3 under the control of a β-estradiol-inducible promoter revealed that root-confined CLE3 overexpression affected gene expression in both roots and shoots. Comparison of CLE2- and CLE3-induced genes indicated that CLE2 and CLE3 peptides target a few common but largely distinct downstream genes. These results suggest that root-derived CLE3 is involved in the regulation of systemic rather than local immune responses. Our study also sheds light on the potential role of CLE peptides in long-distance regulation of plant immunity.

Genome-wide identification and functional characterization of the MdCLE peptide family in apple (Malus × domestica)

The CLAVATA3/EMBRYO SURROUNDING REGION-related (CLE) peptides are critical for stem cell homeostasis in plant shoot and root apical meristem. Although CLE genes have been reported in numerous plants, there is limited information for apple. Here, twenty-five MdCLE genes were identified from apple genome (Apple Genome V1.0 predicted peptides). Analysis of chromosomal location showed that the 25 MdCLE genes were located on 12 of 17 apple chromosomes. Genetic structure analysis showed that 21 of 25 the MdCLE genes were intron-free. Expression patterns showed that most of the MdCLE genes showed higher expression levels in leaves and root compared with the other tissues. In addition, expression analysis demonstrated that MdCLE genes had different gene expression patterns for abiotic stress treatment, suggesting their potential roles in acclimation of apple to adverse environments. The 25 MdCLE genes encoded 19 different CLE peptides, and they were divided into two groups depending on their effects on the inhibition of Arabidopsis root growth. Our results suggest that MdCLE genes may have an important but redundant function in regulating plant growth and development, and this work provides valuable references for further investigation of the biological functions of MdCLE genes.

Peptide Signaling Pathways Regulate Plant Vascular Development.

Plant small peptides, including CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) and Epidermal Patterning Factor-Like (EPFL) peptides, play pivotal roles in coordinating developmental processes through cell-cell communication. Recent studies have revealed that the phloem-derived CLE peptides, CLE41/44 and CLE42, promote (pro-)cambial cell proliferation and inhibit xylem cell differentiation. The endodermis-derived EPFL peptides, EPFL4 and EPFL6, modulate vascular development in the stem. Further, several other peptide ligands CLE9, CLE10, and CLE45 play crucial roles in regulating vascular development in the root. The peptide signaling pathways interact with each other and crosstalk with plant hormone signals. In this mini-review, we summtarize the recent advances on peptides function in vascular development and discuss future perspectives for the research of the CLE and EPFL peptides.

An Evolutionarily Conserved Coreceptor Gene Is Essential for CLAVATA Signaling in Marchantia polymorpha

Growth and development of land plants are controlled by CLAVATA3/EMBRYO SURROUNDING REGION-related (CLE) family of peptide hormones. In contrast to the genetic diversity of CLE family in flowering plants, the liverwort Marchantia polymorpha possesses a minimal set of CLE, MpCLE1(TDIF homolog), and MpCLE2 (CLV3 homolog). MpCLE1 and MpCLE2 peptides exert distinct function at the apical meristem of M. polymorpha gametophyte via specific receptors, MpTDIF RECEPTOR (MpTDR) and MpCLAVATA1 (MpCLV1), respectively, both belonging to the subclass XI of leucine-rich repeat receptor-like kinases (LRR-RLKs). Biochemical and genetic studies in Arabidopsis have shown that TDR/PXY family and CLV1/BAM family recognize the CLE peptide ligand in a heterodimeric complex with a member of subclass-II coreceptors. Here we show that three LRR-RLK genes of M. polymorpha are classified into subclass II, representing three distinct subgroups evolutionarily conserved in land plants. To address the involvement of subclass-II coreceptors in M. polymorpha CLE signaling, we performed molecular genetic analysis on one of them, MpCLAVATA3 INSENSITIVE RECEPTOR KINASE (MpCIK). Two knockout alleles for MpCIK formed narrow apical meristems marked by promMpYUC2:GUS marker, which were not expanded by MpCLE2 peptide treatment, phenocopying Mpclv1. Loss of sensitivity to MpCLE2 peptide was also observed in gemma cup formation in both Mpclv1 and Mpcik. Biochemical analysis using a Nicotiana benthamiana transient expression system revealed weak association between MpCIK and MpCLV1, as well as MpCIK and MpTDR. While MpCIK may also participate in MpCLE1 signaling, our data show that the conserved CLV3-CLV1-CIK module functions in M. polymorpha, controlling meristem activity for development and organ formation for asexual reproduction.

Genome-wide identification of CLE gene family and their potential roles in bolting and fruit bearing in cucumber (Cucumis sativus L.)

BackgroundSignal peptides are essential for plant growth and development. In plants, biological processes including cell-cell communication, cellular proliferation and differentiation, cellular determination of self-incompatibility, and defensive responses, all depend heavily on peptide-signaling networks such as CLE (CLAVATA3/Embryo surrounding region-related). The CLEs are indispensable in different periods of plant growth and development, especially in maintaining the balance between proliferation and differentiation of stem cells in various meristematic tissues.The working system of CLE genes in cucumber, an important economical vegetable (Cucumis sativus L.), has not been fully studied yet. The distributional patterns of chromosome-level genome assembly in cucumber provide a fundamental basis for a genome-wide comparative analysis of CLE genes in such plants.ResultsA total of 26 individual CLE genes were identified in Chinese long ‘9930’ cucumber, the majority of which belong to unstable short alkaline and hydrophilic peptides. A comparative analysis showed a close relationship in the development of CLE genes among Arabidopsis thaliana, melon, and cucumber. Half of the exon-intron structures of all CsCLEs genes are single-exon genes, and motif 1, a typical CLE domain near the C-terminal functioning in signal pathways, is found in all cucumber CLE proteins but CsCLE9. The analysis of CREs (Cis-Regulatory Elements) in the upstream region of the 26 cucumber CLE genes indicates a possible relationship between CsCLE genes and certain functions of hormone response elements. Cucumber resulted closely related to Arabidopsis and melon, having seven and 15 orthologous CLE genes in Arabidopsis and melon, respectively. Additionally, the calculative analysis of a pair of orthologous genes in cucumber showed that as a part of the evolutionary process, CLE genes are undergoing a positive selection process which leads to functional differentiation. The specific expression of these genes was vigorous at the growth and development period and tissues. Cucumber gene CLV3 was overexpressed in Arabidopsis, more than half of the transformed plants in T1 generation showed the phenomena of obvious weakness of the development of growing point, no bolting, and a decreased ability of plant growth. Only two bolted strains showed that either the pod did not develop or the pod was short, and its development was significantly inferior to that in the wild type.ConclusionsIn this study, 26 CLE genes were identified in Chinese long ‘9930’ cucumber genome. The CLE genes were mainly composed of alkaline hydrophilic unstable proteins. The genes of the CLE family were divided into seven classes, and shared close relationships with their homologs in Arabidopsis and melon. The specific expression of these genes was evaluated in different periods of growth and tissue development, and CLV3, which the representative gene of the family, was overexpressed in Arabidopsis, suggesting that it has a role in bolting and fruit bearing in cucumber.

Molecular Dialog between Root and Shoot via Regulatory Peptides and Its Role in Systemic Control of Plant Development

Plant developmental processes are very flexible and highly depend on environmental factors. This is largely due to the existence of regulatory mechanisms that systemically control development on the whole plant level. In plants, regulatory peptides produced in the roots have been identified that are activated in response to different factors influencing root system, such as variation in the level of macronutrients (first of all, nitrogen and phosphorus) in the soil, influence of symbiotic microorganisms (soil rhizobial bacteria and arbuscular mycorrhiza fungi), and water deficiency. Among the systemically acting peptides, the most thoroughly investigated are CLE (CLAVATA3/EMBRYO SURROUNDING REGION-related) and CEP (C-TERMINALLY ENCODED PEPTIDES) peptides that are capable of travelling through the xylem from the roots to the shoot and triggering responses via binding to specific receptors operating in the phloem of the leaf. This review focuses on the role of these two groups of peptides in molecular dialog between the root and shoot.

Induction of Multichotomous Branching by CLAVATA Peptide in Marchantia polymorpha

A key innovation in land plants was the evolution of meristems with stem cells possessing multiple cutting faces (division planes) from which three-dimensional growth is derived in both haploid (gametophyte) and diploid (sporophyte) generations [1-3]. Within each meristem exists a pool of stem cells that must be maintained at a relatively constant size for development to occur appropriately [4-6]. In flowering plants, stem cells of the diploid generation are maintained by CLAVATA3/EMBRYO SURROUNDING REGION-related (CLE) peptide signaling [7, 8]. In the liverwort Marchantia polymorpha, the haploid body undergoes dichotomous branching, an ancestral characteristic of growth derived from the meristem, in which two equivalent body axes are developed via stem cell division, regulated by unknown molecular mechanisms. We show here that in M.polymorpha, treatment with MpCLE2/CLAVATA3 (CLV3) peptide resulted in the accumulation of undifferentiated cells, marked by MpYUC2 expression, in the apical meristem. Removal of MpCLE2 peptide resulted in multichotomous branching from the accumulated cells. Genetic analysis demonstrated that the CLAVATA1 (MpCLV1) receptor, but not the WUSCHEL-related HOMEOBOX (MpWOX) transcription factor, is responsible for MpCLE2 peptide signaling. In the apical meristem, MpCLV1 was expressed broadly in the central region, including the MpYUC2-positive area, whereas MpCLE2 was expressed in a largely complementary manner compared to MpYUC2, suggesting MpCLE2 mediates local cell-to-cell communication. CLV3/CLE peptide, a negative regulator of diploid stem cells in flowering plants, acts as a haploid stem cell-promoting signal in M.polymorpha, implicating a critical role for this pathway in the evolution of body plan in land plants.

Ginsenosides regulate adventitious root formation in Panax ginseng via a CLE45–WOX11 regulatory module

Adventitious root branching is vital to plant growth and regeneration, but the regulation of this process remains unclear. We therefore investigated how ginsenosides regulate adventitious root branching in Panax ginseng. Cell proliferation and adventitious root branching were decreased in the presence of ginsenoside Rb1 and a high concentration of ginsenoside Re, but increased when treating with a low concentration of Re. Moreover, the exogenous application of a synthetic dodeca-amino acid peptide that has a CLAVATA3/EMBRYO SURROUNDING REGION-related (CLE) motif corresponding to PgCLE45 retarded root growth in both ginseng and Arabidopsis. The root Re levels and the expression of the DDS, CYP716A47, and CYP716A53 genes that encode enzymes involved in ginsenoside synthesis were decreased in the presence of PgCLE45. The expression profiles of PgWOX and PgCLE genes were determined to further investigate the CLE-WOX signaling pathway. The levels of PgWOX11 transcripts showed an inverse pattern to PgCLE45 transcripts. Using yeast one-hybrid assay, EMSA, and ChIP assay, we showed that PgWOX11 bound to the PgCLE45 promoter, which contained the HD motif. Transient expression assay showed that PgWOX11 induced the expression of PgCLE45 in adventitious roots, while PgCLE45 suppressed the expression of PgWOX11. These results suggest that there is a negative feedback regulation between PgCLE45 and PgWOX11. Taken together, these data show that ginsenosides regulate adventitious root branching via a novel PgCLE45-PgWOX11 regulatory loop, providing a potential mechanism for the regulation of adventitious root branching.

Recent Advances in Arabidopsis CLE Peptide Signaling

Like communities of people, communities of cells must continuously communicate to thrive. Polypeptide signaling molecules that act as mobile ligands are widely used by eukaryotic organisms to transmit information between cells to coordinate developmental processes and responses to environmental cues. In plants, the CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) genes encode a large family of extracellular signaling peptides that stimulate receptor-mediated signal transduction cascades to modulate diverse developmental and physiological processes. This review highlights the emerging roles of Arabidopsisthaliana CLE peptide signaling pathways in shoot stem cell homeostasis and root xylem development, as well as in root protophloem cell differentiation, vascular cambium activity, and stomatal formation and closure.

Control of proliferation in the haploid meristem by CLE peptide signaling in Marchantia polymorpha.

The homeostasis of meristems in flowering plants is maintained by cell-to-cell communication via CLE (CLAVATA3/EMBRYO SURROUNDING REGION-related) peptide hormones. In contrast, cell signals that regulate meristem activity remains elusive in bryophytes that maintain apical meristems in the gametophyte (haploid) body and undergo a gametophyte-dominant life cycle. We here show that MpCLE1 confines the proliferative activity of gametophytic meristem and affects the overall size of gametangiophores (reproductive organs) in Marchantia polymorpha, which is in sharp contrast with the meristem-promoting function of its ortholog TDIF/CLE41/CLE44 in Arabidopsis vascular meristems. Expression analysis suggests that MpCLE1 and its receptor gene MpTDR are expressed in distinct patterns across the apical meristem. These data suggest that local CLE peptide signaling may have had a role in regulating cell proliferation in the shoot meristem in the ancestral land plant and acts in both sporophytic and gametophytic meristems of extant plants.

Comprehensive Analyses of Nitric Oxide-Induced Plant Stem Cell-Related Genes in Arabidopsis thaliana.

Plant stem cells are pluripotent cells that have diverse applications in regenerative biology and medicine. However, their roles in plant growth and disease resistance are often overlooked. Using high-throughput RNA-seq data, we identified approximately 20 stem cell-related differentially expressed genes (DEGs) that were responsive to the nitric oxide (NO) donor S-nitrosocysteine (CySNO) after six hours of infiltration. Among these DEGs, the highest number of positive correlations (R ≥ 0.8) was observed for CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) 12. Gene ontology (GO) terms for molecular function showed DEGs associated with signal transduction and receptor activity. A promoter study of these DEGs showed the presence of cis-acting elements that are involved in growth as well as the regulation of abiotic and biotic stress. Phylogenetic analysis of the Arabidopsis stem cell-related genes and their common orthologs in rice, soybean, poplar, and tomato suggested that most soybean stem cell-related genes were grouped with the Arabidopsis CLE type of stem cell genes, while the rice stem cell-related genes were grouped with the Arabidopsis receptor-like proteins. The functional genomic-based characterization of the role of stem cell DEGs showed that under control conditions, the clv1 mutant showed a similar phenotype to that of the wild-type (WT) plants; however, under CySNO-mediated nitrosative stress, clv1 showed increased shoot and root length compared to WT. Furthermore, the inoculation of clv1 with virulent Pst DC3000 showed a resistant phenotype with fewer pathogens growing at early time points. The qRT-PCR validation and correlation with the RNA-seq data showed a Pearson correlation coefficient of >0.8, indicating the significantly high reliability of the RNA-seq analysis.

Comprehensive Analyses of Nitric Oxide-Induced Plant Stem Cell-Related Genes in Arabidopsis thaliana

Plant stem cells are pluripotent cells that have diverse applications in regenerative biology and medicine. However, their roles in plant growth and disease resistance are often overlooked. Using high-throughput RNA-seq data, we identified approximately 20 stem cell-related differentially expressed genes (DEGs) that were responsive to the nitric oxide (NO) donor S-nitrosocysteine (CySNO) after six hours of infiltration. Among these DEGs, the highest number of positive correlations (R ≥ 0.8) was observed for CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) 12. Gene ontology (GO) terms for molecular function showed DEGs associated with signal transduction and receptor activity. A promoter study of these DEGs showed the presence of cis-acting elements that are involved in growth as well as the regulation of abiotic and biotic stress. Phylogenetic analysis of the Arabidopsis stem cell-related genes and their common orthologs in rice, soybean, poplar, and tomato suggested that most soybean stem cell-related genes were grouped with the Arabidopsis CLE type of stem cell genes, while the rice stem cell-related genes were grouped with the Arabidopsis receptor-like proteins. The functional genomic-based characterization of the role of stem cell DEGs showed that under control conditions, the clv1 mutant showed a similar phenotype to that of the wild-type (WT) plants; however, under CySNO-mediated nitrosative stress, clv1 showed increased shoot and root length compared to WT. Furthermore, the inoculation of clv1 with virulent Pst DC3000 showed a resistant phenotype with fewer pathogens growing at early time points. The qRT-PCR validation and correlation with the RNA-seq data showed a Pearson correlation coefficient of >0.8, indicating the significantly high reliability of the RNA-seq analysis.