Trichome Formation In Arabidopsis Research Articles

Regulation of capsule spine formation in castor.

Castor (Ricinus communis L.) is a dicotyledonous oilseed crop that can have either spineless or spiny capsules. Spines are protuberant structures that differ from thorns or prickles. The developmental regulatory mechanisms governing spine formation in castor or other plants have remained largely unknown. Herein, using map-based cloning in 2 independent F2 populations, F2-LYY5/DL01 and F2-LYY9/DL01, we identified the RcMYB106 (myb domain protein 106) transcription factor as a key regulator of capsule spine development in castor. Haplotype analyses demonstrated that either a 4,353-bp deletion in the promoter or a single nucleotide polymorphism leading to a premature stop codon in the RcMYB106 gene could cause the spineless capsule phenotype in castor. Results of our experiments indicated that RcMYB106 might target the downstream gene RcWIN1 (WAX INDUCER1), which encodes an ethylene response factor known to be involved in trichome formation in Arabidopsis (Arabidopsis thaliana) to control capsule spine development in castor. This hypothesis, however, remains to be further tested. Nevertheless, our study reveals a potential molecular regulatory mechanism underlying the spine capsule trait in a nonmodel plant species.

Involvement of ABA Responsive SVB Genes in the Regulation of Trichome Formation in Arabidopsis.

Trichome formation in Arabidopsis is regulated by several key regulators, and plants hormones such as gibberellin, salicylic acid, jasmonic acid and cytokinins have been shown to regulate trichome formation by affecting the transcription or activities of the key regulators. We report here the identification of two abscisic acid (ABA) responsive genes, SMALLER TRICHOMES WITH VARIABLE BRANCHES (SVB) and SVB2 as trichome formation regulator genes in Arabidopsis. The expression levels of SVB and SVB2 were increased in response to ABA treatment, their expression levels were reduced in the ABA biosynthesis mutant aba1-5, and they have similar expression pattern. In addition to the trichome defects reported previously for the svb single mutant, we found that even though the trichome numbers were largely unaffected in both the svb and svb2 single mutants generate by using CRISPR/Cas9 gene editing, the trichome numbers were greatly reduced in the svb svb2 double mutants. On the other hand, trichome numbers were increased in SVB or SVB2 overexpression plants. RT-PCR results show that the expression of the trichome formation key regulator gene ENHANCER OF GLABRA3 (EGL3) was affected in the svb svb2 double mutants. Our results suggest that SVB and SVB2 are ABA responsive genes, and SVB and SVB2 function redundantly to regulate trichome formation in Arabidopsis.

Overexpression of VvPPR1, a DYW-type PPR protein in grape, affects the phenotype of Arabidopsis thaliana leaves

Pentatricopeptide repeat (PPR) proteins play important roles in plant growth and development. However, little is known about their functions in the leaf morphogenesis of Jingxiu grape (Vitis vinifera L.). Here, we explored the function of VvPPR1, which encodes a DYW-type PPR protein in grape. We showed that VvPPR1 is involved in the regulation of leaf rolling, anthocyanin accumulation, and trichome formation in Arabidopsis thaliana. Analysis of structural characteristics showed that VvPPR1 is a DYW-type PPR gene in the PLS subfamily consisting of 15 PPR motifs. The N-terminal had a targeted chloroplast site, and the C-terminal had a DYW domain. Quantitative PCR analysis revealed that the expression level of VvPPR1 was highest in grape leaves. Subcellular localization revealed that VvPPR1 is localized in the cytoplasm and chloroplast. VvPPR1-overexpressing plants had rolled leaves, high degrees of anthocyanin accumulation, and longer trichomes. The expression levels of genes related to these phenotypes were either significantly up-regulated or down-regulated. These results demonstrate that VvPPR1 is involved in leaf rolling, anthocyanin accumulation, and trichome formation in Arabidopsis; more generally, our findings indicate that VvPPR1 could be a target for improving the cultivation of horticultural crops.

Nucleoredoxin2 (NRX2) Promotes Jasmonate-Mediated Trichome Formation in Arabidopsis

Thioredoxin (Trx) proteins are essential for the maintenance of cellular redox balance through thiol/disulfide exchange modification. In Arabidopsis, the Trx superfamily consists of multiple protein isotypes distributed in most cellular compartments. Although the functions of chloroplastic and cytosolic Trxs have been investigated in plants, the physiological role of nuclear Trx proteins remains elusive. Nucleoredoxin (NRX) is a nuclear Trx first identified in eukaryotic organisms. Arabidopsis possesses two NRX genes (AtNRX1 and AtNRX2), and the function of AtNRX2 has not been elucidated to date. In this study, we characterized the function of AtNRX2 using the atnrx2 knockout mutant, based on its comparison with the atnrx1 mutant. In atnrx2 knockout mutant plants, trichome number was significantly reduced compared with the wild type (WT; Col-0) and the atnrx1 mutant. In response to JA induction of trichome, trichome formation was markedly diminished in the atnrx2 mutant. In addition, expression levels of genes involved in trichome formation were reduced in the atnrx2 mutant compared with the WT and atnrx1 mutant. Overall, our results suggest that AtNRX2 plays a physiological role in JA-mediated trichome formation in Arabidopsis.

Single-repeat R3 MYB transcription factors from Platanus acerifolia negatively regulate trichome formation in Arabidopsis.

Four R3 MYB genes were cloned and identified from Platanus acerifolia and analysed according to endogenous gene expression profiles, protein-protein interaction patterns, phenotypic effects and related gene expression profiles in transgenic Arabidopsis, suggesting that London plane R3 MYB genes inhibit trichome formation in Arabidopsis. The CPC-like MYB transcription factors including CAPRICE (CPC), TRIPTYCHON (TRY), ENHANCER OF TRY AND CPC 1, 2 and 3 (ETC1, ETC2 and ETC3), TRICHOMELESS1 (TCL1) and TRICHOMELESS2(TCL2) play important roles in controlling trichome patterning in Arabidopsis. In this study, four sequences homologous with the Arabidopsis CPC family were identified from London plane and named PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3. Over-expression of PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3 in Arabidopsis resulted in glabrous phenotypes. In addition, expression of endogenous GL2, GL1, MYB23, TTG2 and a set of R3 MYB-encoding genes was markedly reduced. Furthermore, the protein products of PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3 were shown to interact with PaGL3 in yeast two-hybrid assays. Together, these results likely suggest that the mechanisms of trichome regulation in London plane have similarities with those in Arabidopsis.

Genetic evidence suggests that GIS functions downstream of TCL1 to regulate trichome formation in Arabidopsis

BackgroundTrichome formation in Arabidopsis is regulated by a MBW complex formed by MYB, bHLH and WD40 transcriptional factors, which can activate GLABRA2 (GL2) and the R3 MYB transcription factor genes. GL2 promotes trichome formation, whereas R3 MYBs are able to block the formation of the MBW complex. It has been reported that the C2H2 transcription factor GIS (GLABROUS INFLORESCENCE STEMS) functions upstream of the MBW activator complex to regulate trichome formation, and that the expression of TCL1 is not regulated by the MBW complex. However, gis and the R3 MYB gene mutant tcl1 (trichomeless 1) have opposite inflorescence trichome phenotypes, but their relationship in regulating trichome formation remained unknown.ResultsBy generating and characterization of the gis tcl1 double mutant, we found that trichome formation in the gis tcl1double and the tcl1 single mutants were largely indistinguishable, but the trichome formation in the 35S:TCL1/gis transgenic plant was similar to that in the gis mutant. By using quantitative RT-PCR analysis, we showed that expression level of GIS was increased in the triple mutant tcl1 try cpc, but the expression level of TCL1 was not affected in the gis mutant. On the other hand, trichome morphology in both gis tcl1 and 35S:TCL1/gis plants was similar to that in the gis mutant.ConclusionsIn summary, our results indicate that GIS may work downstream of TCL1 to regulate trichome formation, and GIS has a dominant role in controlling trichome morphology.

NTL8 Regulates Trichome Formation in Arabidopsis by Directly Activating R3 MYB Genes TRY and TCL1.

The NAM, ATAF1/2, and CUC (NAC) are plant-specific transcription factors that regulate multiple aspects of plant growth and development and plant response to environmental stimuli. We report here the identification of NTM1-LIKE8 (NTL8), a membrane-associated NAC transcription factor, as a novel regulator of trichome formation in Arabidopsis (Arabidopsis thaliana). From an activation-tagged Arabidopsis population, we identified a dominant, gain-of-function mutant with glabrous inflorescence stem. By using plasmid rescue and RT-PCR analyses, we found that NTL8 was tagged; thus, the mutant was named ntl8-1 Dominant (ntl8-1D). Recapitulation experiment further confirmed that the phenotype observed in the ntl8-1D mutant was caused by elevated expression of NTL8 Quantitative RT-PCR results showed that the expression level of the single-repeat R3 MYB genes TRIPTYCHON (TRY) and TRICHOMELESS1 (TCL1) was elevated in the ntl8-1D mutant. Genetic analyses demonstrated that NTL8 acts upstream of TRY and TCL1 in the regulation of trichome formation. When recruited to the promoter region of the reporter gene Gal4:GUS by a fused GAL4 DNA-binding domain, NTL8 activated the expression of the reporter gene. Chromatin immunoprecipitation results indicated that TRY and TCL1 are direct targets of NTL8. However, NTL8 did not interact with SQUAMOSA PROMOTER BINDING PROTEIN LIKE9, another transcription factor that regulates the expression of TRY and TCL1, in yeast and plant cells. Taken together, our results suggest that NTL8 negatively regulates trichome formation in Arabidopsis by directly activating the expression of TRY and TCL1.

Genome-wide identification of GLABRA3 downstream genes for anthocyanin biosynthesis and trichome formation in Arabidopsis

GLABRA3 (GL3), a bHLH transcription factor, has previously proved to be involved in anthocyanin biosynthesis and trichome formation in Arabidopsis, however, its downstream targeted genes are still largely unknown. Here, we found that GL3 was widely present in Arabidopsis vegetative and reproductive organs. New downstream targeted genes of GL3 for anthocyanin biosynthesis and trichome formation were identified in young shoots and expanding true leaves by RNA sequencing. GL3-mediated gene expression was tissue specific in the two biological processes. This study provides new clues to further understand the GL3-mediated regulatory network of anthocyanin biosynthesis and trichome formation in Arabidopsis.

MYB and HD-ZIP IV homologs related to trichome formation are involved in epidermal bladder cell development in the halophyte Mesembryanthemum crystallinum L.

The common ice plant, Mesembryanthemum crystallinum L., a halophytic new functional leafy vegetable crop, develops epidermal bladder cells (EBCs) on the surfaces of its aerial organs. Our previous studies of the physiological characteristics of the wild-type and the EBC-less mutant indicated that EBCs sequester salt and maintain ion homeostasis within photosynthetically active tissues. The EBC has been thought to be a modified trichome; however, molecular mechanisms governing EBC development in the common ice plant have not been fully understood. Here, we have analyzed the steady-state mRNA abundance of nineteen cotton fiber-related gene homologs and eight Arabidopsis trichome development-related genes, and found that a MYB transcription factor homolog (McMYB2) and a GLABRA2-like gene (McC4HDZ) were preferentially expressed in wild-type plants, whereas a putative TRIPTYCHON (McTRY)- and CAPRICE-like gene (McCPC) were preferentially expressed in the EBC-mutant. The full-length cDNA sequences of these homologs were determined, and constructs containing McC4HDZ and McMYB2 were introduced into an Arabidopsis trichome-less mutant and wild-type plants. Overexpression of McMYB2 in wild-type Arabidopsis increased trichome number, associated with activation of the trichome development-related gene, GLABRA2 (GL2). Moreover, overexpression of McC4HDZ partially complemented trichome development in the trichome-less mutant of gl2-1, and resulted in increased trichome number in wild-type Arabidopsis, associated with the upregulation of key trichome-positive regulators GLABRA1 (GL1) and TRANSPARENT TESTA GLABRA1 (TTG1). These results suggest that McMYB2 and McC4HDZ could be functional in Arabidopsis trichome formation, implying that EBCs of the common ice plant and trichomes of Arabidopsis may share some molecular mechanisms in their development.

A single amino acid substitution in the R3 domain of GLABRA1 leads to inhibition of trichome formation in Arabidopsis without affecting its interaction with GLABRA3.

GLABRA1 (GL1) is an R2R3 MYB transcription factor that regulates trichome formation in Arabidopsis by interacting with the bHLH transcription factor GLABRA3 (GL3) or ENHANCER OF GL3 (EGL3). The conserved [D/E]L×2 [R/K]×3L×6L×3R amino acid signature in the R3 domain of MYB proteins has been shown to be required for the interaction of MYBs with R/B-like bHLH transcription factors. By using genetic and molecular analyses, we show that the glabrous phenotype in the nph4-1 mutant is caused by a single nucleotide mutation in the GL1 gene, generating a Ser to Phe substitution (S92F) in the conserved [D/E]L×2[R/K]×3L×6L×3R amino acid signature of GL1. Activation of the integrated GL2p:GUS reporter gene in protoplasts by cotransfection of GL1 and GL3 or EGL3 was abolished by this GL1-S92F substitution. However, GL1-S92F interacted successfully with GL3 or EGL3 in protoplast transfection assays. Unlike VPGL1GL3, the fusion protein VPGL1-S92FGL3 failed to activate the integrated GL2p:GUS reporter gene in transfected protoplasts. These results suggested that the S92 in the conserved [D/E]L×2 [R/K]×3L×6L×3R amino acid signature of GL1 is not essential for the interaction of GL1 and GL3, but may play a role in the binding of GL1 to the promoters of its target genes.

GLABROUS INFLORESCENCE STEMS3 (GIS3) regulates trichome initiation and development in Arabidopsis.

Arabidopsis trichome formation is an excellent model for studying various aspects of plant cell development and cell differentiation. Our previous works have demonstrated that several C2H2 zinc finger proteins, including GIS, GIS2, ZFP5, ZFP6 and ZFP8, control trichome cell development through GA and cytokinin signalling in Arabidopsis. We identified a novel C2H2 zinc finger protein, GLABROUS INFLORESCENCE STEMS 3 (GIS3), which is a key factor in regulating trichome development in Arabidopsis. In comparison with wild-type plants, loss-of-function of GIS3 mutants exhibited a significantly decreased number of trichomes in cauline leaves, lateral branches, sepals of flowers, and main stems. Overexpression of GIS3 resulted in increased trichome densities in sepal, cauline leaves, lateral branches, main inflorescence stems and in the appearance of ectopic trichomes on carpels. The molecular and genetic analyses show that GIS3 acts upstream of GIS, GIS2, ZFP8 and the key trichome initiation factors, GL1 and GL3. Steroid-inducible gene expression analyses and chromatin immunoprecipitation (ChIP) experiments suggest that GIS and GIS2 are the direct target genes of GIS3.

Control of trichome formation in Arabidopsis by poplar single-repeat R3 MYB transcription factors.

In Arabidopsis, trichome formation is regulated by the interplay of R3 MYBs and several others transcription factors including the WD40-repeat protein TRANSPARENT TESTA GLABRA1 (TTG1), the R2R3 MYB transcription factor GLABRA1 (GL1), the bHLH transcription factor GLABRA3 (GL3) or ENHANCER OF GLABRA3 (EGL3), and the homeodomain protein GLABRA2 (GL2). R3 MYBs including TRICHOMELESS1 (TCL1), TCL2, TRYPTICHON (TRY), CAPRICE (CPC), ENHANCER OF TRY AND CPC1 (ETC1), ETC2 and ETC3 negatively regulate trichome formation by competing with GL1 for binding GL3 or EGL3, thus blocking the formation of TTG1–GL3/EGL3–GL1, an activator complex required for the activation of the trichome positive regulator gene GL2. However, it is largely unknown if R3 MYBs in other plant species especially woody plants have similar functions. By BLASTing the Populus trichocarpa protein database using the entire amino acid sequence of TCL1, an Arabidopsis R3 MYB transcription factor, we identified a total of eight R3 MYB transcription factor genes in poplar, namely P. trichocarpa TRICHOMELESS1 through 8 (PtrTCL1–PtrTCL8). The amino acid signature required for interacting with bHLH transcription factors and the amino acids required for cell-to-cell movement of R3 MYBs are not fully conserved in all PtrTCLs. When tested in Arabidopsis protoplasts, however, all PtrTCLs interacted with GL3. Expressing each of the eight PtrTCL genes in Arabidopsis resulted in either glabrous phenotypes or plants with reduced trichome numbers, and expression levels of GL2 in all transgenic plants tested were greatly reduced. Expression of PtrTCL1 under the control of TCL1 native promoter almost completely complemented the mutant phenotype of tcl. In contrast, expression of PtrTCL1 under the control of TRY native promoter in the try mutant, or under the control of CPC native promoter in the cpc mutant resulted in glabrous phenotypes, suggesting that PtrTCL1 functions similarly to TCL1, but not TRY and CPC.

A full-length R-like basic-helix-loop-helix transcription factor is required for anthocyanin upregulation whereas the N-terminal region regulates epidermal hair formation

Earlier studies have shown that the Lc gene of maize, a member of the R gene family that encode basic-helix-loop-helix (bHLH) transcription factors, is involved with anthocyanin production and trichome formation in Arabidopsis. We previously reported that the N-terminus of R protein interacts with CAPRICE (CPC), a regulatory protein, in triggering epidermal hair differentiation in Arabidopsis. In this study, we investigated the roles of full-length R, the N-terminal region of R (RN) and the C-terminal region of R (RC) in epidermal cell differentiation and anthocyanin production. We found that the N-terminal region was responsible for leaf trichome and root hair differentiation, whereas full-length R was required for anthocyanin upregulation. Yeast two-hybrid analysis showed that the C-terminal region was the binding site for the formation of homo- or hetero-dimers of the R-like bHLH transcription factor. To stimulate anthocyanin production, full-length R is required.

Two-dimensional patterning by a trapping/depletion mechanism: the role of TTG1 and GL3 in Arabidopsis trichome formation.

Trichome patterning in Arabidopsis serves as a model system to study how single cells are selected within a field of initially equivalent cells. Current models explain this pattern by an activator–inhibitor feedback loop. Here, we report that also a newly discovered mechanism is involved by which patterning is governed by the removal of the trichome-promoting factor TRANSPARENT TESTA GLABRA1 (TTG1) from non-trichome cells. We demonstrate by clonal analysis and misexpression studies that Arabidopsis TTG1 can act non-cell-autonomously and by microinjection experiments that TTG1 protein moves between cells. While TTG1 is expressed ubiquitously, TTG1–YFP protein accumulates in trichomes and is depleted in the surrounding cells. TTG1–YFP depletion depends on GLABRA3 (GL3), suggesting that the depletion is governed by a trapping mechanism. To study the potential of the observed trapping/depletion mechanism, we formulated a mathematical model enabling us to evaluate the relevance of each parameter and to identify parameters explaining the paradoxical genetic finding that strong ttg1 alleles are glabrous, while weak alleles exhibit trichome clusters.

Trichomes: different regulatory networks lead to convergent structures

Sometimes, proteins, biological structures or even organisms have similar functions and appearances but have evolved through widely divergent pathways. There is experimental evidence to suggest that different developmental pathways have converged to produce similar outgrowths of the aerial plant epidermis, referred to as trichomes. The emerging picture suggests that trichomes in Arabidopsis thaliana and, perhaps, in cotton develop through a transcriptional regulatory network that differs from those regulating trichome formation in Antirrhinum and Solanaceous species. Several lines of evidence suggest that the duplication of a gene controlling anthocyanin production and subsequent divergence might be the major force driving trichome formation in Arabidopsis, whereas the multicellular trichomes of Antirrhinum and Solanaceous species appear to have a different regulatory origin.

Spatial regulation of trichome formation inArabidopsis thaliana

Trichomes in Arabidopsis are single-celled hairs that develop from epidermal cells regularly distributed on leaves, sepals and stems. Trichome formation in Arabidopsis is emerging as a genetic and molecular model system for the analysis of the spatial regulation of cell type specification in plants. In this review we focus on two aspects of trichome initiation: the regulation of organ and tissue layer specificity and the establishment of an epidermal spacing pattern.

Pattern Formation and Cell Differentiation: Trichomes in Arabidopsis as a Genetic Model System

Arabidopsis trichomes are single-celled hairs that originate from epidermal cells and are distributed regularly on most aerial body parts. During the last decade, trichome formation in Arabidopsis has been established as a genetic and molecular model system to study various general developmental and cellular mechanisms. This review summarizes progress in the investigation of several aspects of trichome development: the spatial regulation of cell fate determination, the regulation of cell differentiation in response to exogenous signals and plant hormones, and the regulation of endoreplication, cell growth, and cell morphogenesis.