Qingzhe Zhai

Role of the Arabidopsis thaliana NAC transcription factors ANAC019 and ANAC055 in regulating jasmonic acid-signaled defense responses.

Jasmonic acid (JA) is an important phytohormone that regulates plant defense responses against herbivore attack, pathogen infection and mechanical wounding. In this report, we provided biochemical and genetic evidence to show that the Arabidopsis thaliana NAC family proteins ANAC019 and ANAC055 might function as transcription activators to regulate JA-induced expression of defense genes. The role of the two NAC genes in JA signaling was examined with the anac019 anac055 double mutant and with transgenic plants overexpressing ANAC019 or ANAC055. The anac019 anac055 double mutant plants showed attenuated JA-induced VEGETATIVE STORAGE PROTEIN1 (VSP1) and LIPOXYGENASE2 (LOX2) expression, whereas transgenic plants overexpressing the two NAC genes showed enhanced JA-induced VSP1 and LOX2 expression. That the JA-induced expression of the two NAC genes depends on the function of COI1 and AtMYC2, together with the finding that overexpression of ANAC019 partially rescued the JA-related phenotype of the atmyc2-2 mutant, has led us to a hypothesis that the two NAC proteins act downstream of AtMYC2 to regulate JA-signaled defense responses. Further evidence to substantiate this idea comes from the observation that the response of the anac019 anac055 double mutant to a necrotrophic fungus showed high similarity to that of the atmyc2-2 mutant.

The Basic Helix-Loop-Helix Transcription Factor MYC2 Directly Represses PLETHORA Expression during Jasmonate-Mediated Modulation of the Root Stem Cell Niche in Arabidopsis

This study investigates the mechanisms underlying jasmonate-induced inhibition of primary root growth. Jasmonate inhibits the expression of two AP2-domain transcription factors, PLETHORA1 and 2, in a MYC2-dependent fashion. MYC2 is suggested to integrate the jasmonate and auxin pathways during the maintenance of the root stem cell niche. The root stem cell niche, which in the Arabidopsis thaliana root meristem is an area of four mitotically inactive quiescent cells (QCs) and the surrounding mitotically active stem cells, is critical for root development and growth. We report here that during jasmonate-induced inhibition of primary root growth, jasmonate reduces root meristem activity and leads to irregular QC division and columella stem cell differentiation. Consistently, jasmonate reduces the expression levels of the AP2-domain transcription factors PLETHORA1 (PLT1) and PLT2, which form a developmentally instructive protein gradient and mediate auxin-induced regulation of stem cell niche maintenance. Not surprisingly, the effects of jasmonate on root stem cell niche maintenance and PLT expression require the functioning of MYC2/JASMONATE INSENSITIVE1, a basic helix-loop-helix transcription factor that involves versatile aspects of jasmonate-regulated gene expression. Gel shift and chromatin immunoprecipitation experiments reveal that MYC2 directly binds the promoters of PLT1 and PLT2 and represses their expression. We propose that MYC2-mediated repression of PLT expression integrates jasmonate action into the auxin pathway in regulating root meristem activity and stem cell niche maintenance. This study illustrates a molecular framework for jasmonate-induced inhibition of root growth through interaction with the growth regulator auxin.

The Arabidopsis Mediator Subunit MED25 Differentially Regulates Jasmonate and Abscisic Acid Signaling through Interacting with the MYC2 and ABI5 Transcription Factors

This work describes that the MED25 subunit of the Arabidopsis thaliana Mediator complex positively regulates JA-mediated gene expression through interacting with the transcription factor MYC2. It also describes that MED25 negatively regulates ABA-mediated gene expression through interacting with the transcription factor ABI5. Transcriptional regulation plays a central role in plant hormone signaling. At the core of transcriptional regulation is the Mediator, an evolutionarily conserved, multisubunit complex that serves as a bridge between gene-specific transcription factors and the RNA polymerase machinery to regulate transcription. Here, we report the action mechanisms of the MEDIATOR25 (MED25) subunit of the Arabidopsis thaliana Mediator in regulating jasmonate- and abscisic acid (ABA)–triggered gene transcription. We show that during jasmonate signaling, MED25 physically associates with the basic helix-loop-helix transcription factor MYC2 in promoter regions of MYC2 target genes and exerts a positive effect on MYC2-regulated gene transcription. We also show that MED25 physically associates with the basic Leu zipper transcription factor ABA-INSENSITIVE5 (ABI5) in promoter regions of ABI5 target genes and shows a negative effect on ABI5-regulated gene transcription. Our results reveal that underlying the distinct effects of MED25 on jasmonate and ABA signaling, the interaction mechanisms of MED25 with MYC2 and ABI5 are different. These results highlight that the MED25 subunit of the Arabidopsis Mediator regulates a wide range of signaling pathways through selectively interacting with specific transcription factors.

The Arabidopsis RING Finger E3 Ligase RHA2a Is a Novel Positive Regulator of Abscisic Acid Signaling during Seed Germination and Early Seedling Development

The phytohormone abscisic acid (ABA) is well known for its regulatory roles in integrating environmental constraints with the developmental programs of plants. Here, we characterize the biological function of the Arabidopsis (Arabidopsis thaliana) RING-H2 protein RHA2a in ABA signaling. The rha2a mutant is less sensitive to ABA than the wild type during seed germination and early seedling development, whereas transgenic plants overexpressing RHA2a are hypersensitive, indicating that RHA2a positively regulates ABA-mediated control of seed germination and early seedling development. Double mutant analyses of rha2a with several known ABA-insensitive mutants suggest that the action of RHA2a in ABA signaling is independent of that of the transcription factors ABI3, ABI4, and ABI5. We provide evidence showing that RHA2a also positively regulates plant responses to salt and osmotic stresses during seed germination and early seedling development. RHA2a is a functional E3 ubiquitin ligase, and its conserved RING domain is likely important for the biological function of RHA2a in ABA signaling. Together, these results suggest that the E3 ligase RHA2a is an important regulator of ABA signaling during seed germination and early seedling development.Jasmonate- and ABA-mediated signalings are involved in the activation of defense responses of plants to biotic and abiotic stresses. Accumulating evidence has suggested the existence of comprehensive synergistic or antagonistic cross-talks between these two signaling pathways. However, relatively little is known about how these cross-talks are executed at the molecular level. Our recent works have implied that, ANAC019 and ANAC055, two highly related NAC family transcription factors in Arabidopsis, may play a dual role in regulating jasmonate response and ABA response.

Interaction between MYC2 and ETHYLENE INSENSITIVE3 Modulates Antagonism between Jasmonate and Ethylene Signaling in Arabidopsis

The authors reveal a mechanism underlying jasmonate (JA) and ethylene (ET) antagonism: Interaction between the JA-activated transcription factor MYC2 and the ET-stabilized transcription factor EIN3, reciprocally repressing their transcriptional activity, modulates the antagonistic actions of JA and ET in regulating apical hook curvature, wound-responsive gene expression, and defense against insect attack. Plants have evolved sophisticated mechanisms for integration of endogenous and exogenous signals to adapt to the changing environment. Both the phytohormones jasmonate (JA) and ethylene (ET) regulate plant growth, development, and defense. In addition to synergistic regulation of root hair development and resistance to necrotrophic fungi, JA and ET act antagonistically to regulate gene expression, apical hook curvature, and plant defense against insect attack. However, the molecular mechanism for such antagonism between JA and ET signaling remains unclear. Here, we demonstrate that interaction between the JA-activated transcription factor MYC2 and the ET-stabilized transcription factor ETHYLENE-INSENSITIVE3 (EIN3) modulates JA and ET signaling antagonism in Arabidopsis thaliana. MYC2 interacts with EIN3 to attenuate the transcriptional activity of EIN3 and repress ET-enhanced apical hook curvature. Conversely, EIN3 interacts with and represses MYC2 to inhibit JA-induced expression of wound-responsive genes and herbivory-inducible genes and to attenuate JA-regulated plant defense against generalist herbivores. Coordinated regulation of plant responses in both antagonistic and synergistic manners would help plants adapt to fluctuating environments.

Arabidopsis Tyrosylprotein Sulfotransferase Acts in the Auxin/PLETHORA Pathway in Regulating Postembryonic Maintenance of the Root Stem Cell Niche

Arabidopsis tyrosylprotein sulfotransferase (TPST) maintains the postembryonic root stem cell niche by regulating basal- and auxin-induced expression of the PLETHORA stem cell transcription factor. TPST-mediated activation of a group of sulfated peptides known as root meristem growth factors provides a link between the phytohormone auxin and PLETHORA in root stem cell niche maintenance. Recent identification of the Arabidopsis thaliana tyrosylprotein sulfotransferase (TPST) and a group of Tyr-sulfated peptides known as root meristem growth factors (RGFs) highlights the importance of protein Tyr sulfation in plant growth and development. Here, we report the action mechanism of TPST in maintenance of the root stem cell niche, which in the Arabidopsis root meristem is an area of four mitotically inactive quiescent cells plus the surrounding mitotically active stem cells. Mutation of TPST leads to defective maintenance of the root stem cell niche, decreased meristematic activity, and stunted root growth. We show that TPST expression is positively regulated by auxin and that mutation of this gene affects auxin distribution by reducing local expression levels of several PIN genes and auxin biosynthetic genes in the stem cell niche region. We also show that mutation of TPST impairs basal- and auxin-induced expression of the PLETHORA (PLT) stem cell transcription factor genes and that overexpression of PLT2 rescues the root meristem defects of the loss-of-function mutant of TPST. Together, these results support that TPST acts to maintain root stem cell niche by regulating basal- and auxin-induced expression of PLT1 and PLT2. TPST-dependent sulfation of RGFs provides a link between auxin and PLTs in regulating root stem cell niche maintenance.

Phosphorylation-Coupled Proteolysis of the Transcription Factor MYC2 Is Important for Jasmonate-Signaled Plant Immunity

As a master regulator of jasmonic acid (JA)–signaled plant immune responses, the basic helix-loop-helix (bHLH) Leu zipper transcription factor MYC2 differentially regulates different subsets of JA–responsive genes through distinct mechanisms. However, how MYC2 itself is regulated at the protein level remains unknown. Here, we show that proteolysis of MYC2 plays a positive role in regulating the transcription of its target genes. We discovered a 12-amino-acid element in the transcription activation domain (TAD) of MYC2 that is required for both the proteolysis and the transcriptional activity of MYC2. Interestingly, MYC2 phosphorylation at residue Thr328, which facilitates its turnover, is also required for the MYC2 function to regulate gene transcription. Together, these results reveal that phosphorylation-coupled turnover of MYC2 stimulates its transcription activity. Our results exemplify that, as with animals, plants employ an “activation by destruction” mechanism to fine-tune their transcriptome to adapt to their ever-changing environment.

PIF4 and PIF5 Transcription Factors Link Blue Light and Auxin to Regulate the Phototropic Response in Arabidopsis

This work reveals that two key transcription factors of light signaling, PHYTOCHROME INTERACTING FACTOR4 (PIF4) and PIF5, negatively regulate Arabidopsis phototropism through directly activating the expression of the auxin response repressors IAA19 and IAA29. The study identifies a key step of phototropic signaling in Arabidopsis by showing that the growth factors PIF4 and PIF5 link light and auxin. Both blue light (BL) and auxin are essential for phototropism in Arabidopsis thaliana. However, the mechanisms by which light is molecularly linked to auxin during phototropism remain elusive. Here, we report that PHYTOCHROME INTERACTING FACTOR4 (PIF4) and PIF5 act downstream of the BL sensor PHOTOTROPIN1 (PHOT1) to negatively modulate phototropism in Arabidopsis. We also reveal that PIF4 and PIF5 negatively regulate auxin signaling. Furthermore, we demonstrate that PIF4 directly activates the expression of the AUXIN/INDOLE-3-ACETIC ACID (IAA) genes IAA19 and IAA29 by binding to the G-box (CACGTG) motifs in their promoters. Our genetic assays demonstrate that IAA19 and IAA29, which physically interact with AUXIN RESPONSE FACTOR7 (ARF7), are sufficient for PIF4 to negatively regulate auxin signaling and phototropism. This study identifies a key step of phototropic signaling in Arabidopsis by showing that PIF4 and PIF5 link light and auxin.

Closely Related NAC Transcription Factors of Tomato Differentially Regulate Stomatal Closure and Reopening during Pathogen Attack

This work reports the distinct mechanisms of two homologous NAC proteins of tomato, JA2 and JA2L, in regulating Pseudomonas syringae pv tomato DC3000–induced stomatal movement. Whereas JA2 acts in abscisic acid (ABA)–mediated stomatal closure by promoting ABA biosynthesis, JA2L functions in jasmonate/coronatine–mediated stomatal reopening by suppressing salicylic acid accumulation. To restrict pathogen entry, plants close stomata as an integral part of innate immunity. To counteract this defense, Pseudomonas syringae pv tomato produces coronatine (COR), which mimics jasmonic acid (JA), to reopen stomata for bacterial entry. It is believed that abscisic acid (ABA) plays a central role in regulating bacteria-triggered stomatal closure and that stomatal reopening requires the JA/COR pathway, but the downstream signaling events remain unclear. We studied the stomatal immunity of tomato (Solanum lycopersicum) and report here the distinct roles of two homologous NAC (for NAM, ATAF1,2, and CUC2) transcription factors, JA2 (for jasmonic acid2) and JA2L (for JA2-like), in regulating pathogen-triggered stomatal movement. ABA activates JA2 expression, and genetic manipulation of JA2 revealed its positive role in ABA-mediated stomatal closure. We show that JA2 exerts this effect by regulating the expression of an ABA biosynthetic gene. By contrast, JA and COR activate JA2L expression, and genetic manipulation of JA2L revealed its positive role in JA/COR-mediated stomatal reopening. We show that JA2L executes this effect by regulating the expression of genes involved in the metabolism of salicylic acid. Thus, these closely related NAC proteins differentially regulate pathogen-induced stomatal closure and reopening through distinct mechanisms.

Bestatin, an Inhibitor of Aminopeptidases, Provides a Chemical Genetics Approach to Dissect Jasmonate Signaling in Arabidopsis

Bestatin, a potent inhibitor of some aminopeptidases, was shown previously to be a powerful inducer of wound-response genes in tomato (Lycopersicon esculentum). Here, we present several lines of evidence showing that bestatin specifically activates jasmonic acid (JA) signaling in plants. First, bestatin specifically activates the expression of JA-inducible genes in tomato and Arabidopsis (Arabidopsis thaliana). Second, the induction of JA-responsive genes by bestatin requires the COI1-dependent JA-signaling pathway, but does not depend strictly on JA biosynthesis. Third, microarray analysis using Arabidopsis whole-genome chip demonstrates that the gene expression profile of bestatin-treated plants is similar to that of JA-treated plants. Fourth, bestatin promotes a series of JA-related developmental phenotypes. Taken together, the unique action mode of bestatin in regulating JA-signaled processes leads us to the hypothesis that bestatin exerts its effects through the modulation of some key regulators in JA signaling. We have employed bestatin as an experimental tool to dissect JA signaling through a chemical genetic screening, which yielded a collection of Arabidopsis bestatin-resistant (ber) mutants that are insensitive to the inhibitory effects of bestatin on root elongation. Further characterization efforts demonstrate that some ber mutants are defective in various JA-induced responses, which allowed us to classify the ber mutants into three phenotypic groups: JA-insensitive ber mutants, JA-hypersensitive ber mutants, and mutants insensitive to bestatin but showing normal response to JA. Genetic and phenotypic analyses of the ber mutants with altered JA responses indicate that we have identified several novel loci involved in JA signaling.