Wenkun Zhou

Arabidopsis ASA1 Is Important for Jasmonate-Mediated Regulation of Auxin Biosynthesis and Transport during Lateral Root Formation

Plant roots show an impressive degree of plasticity in adapting their branching patterns to ever-changing growth conditions. An important mechanism underlying this adaptation ability is the interaction between hormonal and developmental signals. Here, we analyze the interaction of jasmonate with auxin to regulate lateral root (LR) formation through characterization of an Arabidopsis thaliana mutant, jasmonate-induced defective lateral root1 (jdl1/asa1-1). We demonstrate that, whereas exogenous jasmonate promotes LR formation in wild-type plants, it represses LR formation in jdl1/asa1-1. JDL1 encodes the auxin biosynthetic gene ANTHRANILATE SYNTHASE α1 (ASA1), which is required for jasmonate-induced auxin biosynthesis. Jasmonate elevates local auxin accumulation in the basal meristem of wild-type roots but reduces local auxin accumulation in the basal meristem of mutant roots, suggesting that, in addition to activating ASA1-dependent auxin biosynthesis, jasmonate also affects auxin transport. Indeed, jasmonate modifies the expression of auxin transport genes in an ASA1-dependent manner. We further provide evidence showing that the action mechanism of jasmonate to regulate LR formation through ASA1 differs from that of ethylene. Our results highlight the importance of ASA1 in jasmonate-induced auxin biosynthesis and reveal a role for jasmonate in the attenuation of auxin transport in the root and the fine-tuning of local auxin distribution in the root basal meristem.

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.

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.

Jasmonate modulates endocytosis and plasma membrane accumulation of the Arabidopsis PIN2 protein.

The subcellular distribution of the PIN-FORMED (PIN) family of auxin transporters plays a critical role in auxin gradient-mediated developmental processes, including lateral root formation and gravitropic growth. Here, we report two distinct aspects of CORONATINE INSENSITIVE 1 (COI1)- and AUXIN RESISTANT 1 (AXR1)-dependent methyl jasmonate (MeJA) effects on PIN2 subcellular distribution: at lower concentration (5 μM), MeJA inhibits PIN2 endocytosis, whereas, at higher concentration (50 μM), MeJA reduces PIN2 accumulation in the plasma membrane. We show that mutations of ASA1 (ANTHRANILATE SYNTHASE a1) and the TIR1/AFBs (TRANSPORT INHIBITOR RESPONSE 1/AUXIN-SIGNALING F-BOX PROTEINs) auxin receptor genes impair the inhibitory effect of 5 μM MeJA on PIN2 endocytosis, suggesting that a lower concentration of jasmonate inhibits PIN2 endocytosis through interaction with the auxin pathway. In contrast, mutations of ASA1 and the TIR1/AFBs auxin receptor genes enhance, rather than impair, the reduction effect of 50 μM MeJA on the plasma membrane accumulation of PIN2, suggesting that this action of jasmonate is independent of the auxin pathway. In addition to the MeJA effects on PIN2 endocytosis and plasma membrane residence, we also show that MeJA alters lateral auxin redistribution on gravi-stimulation, and therefore impairs the root gravitropic response. Our results highlight the importance of jasmonate-auxin interaction in the coordination of plant growth and the adaptation response.

Plastid-Localized Glutathione Reductase2–Regulated Glutathione Redox Status Is Essential for Arabidopsis Root Apical Meristem Maintenance

Plastid-localized glutathione reductase2 regulates root growth and root meristem maintenance through modulation of its GSH redox status, resulting in regulation of downstream effectors of the auxin/PLETHORA pathway, as well as of some auxin/ PLETHORA-independent pathways. This function differs from that of glutathione reductase1, which is not essential for plant development. Glutathione is involved in thiol redox signaling and acts as a major redox buffer against reactive oxygen species, helping to maintain a reducing environment in vivo. Glutathione reductase (GR) catalyzes the reduction of glutathione disulfide (GSSG) into reduced glutathione (GSH). The Arabidopsis thaliana genome encodes two GRs: GR1 and GR2. Whereas the cytosolic/peroxisomal GR1 is not crucial for plant development, we show here that the plastid-localized GR2 is essential for root growth and root apical meristem (RAM) maintenance. We identify a GR2 mutant, miao, that displays strong inhibition of root growth and severe defects in the RAM, with GR activity being reduced to ∼50%. miao accumulates high levels of GSSG and exhibits increased glutathione oxidation. The exogenous application of GSH or the thiol-reducing agent DTT can rescue the root phenotype of miao, demonstrating that the RAM defects in miao are triggered by glutathione oxidation. Our in silico analysis of public microarray data shows that auxin and glutathione redox signaling generally act independently at the transcriptional level. We propose that glutathione redox status is essential for RAM maintenance through both auxin/PLETHORA (PLT)-dependent and auxin/PLT-independent redox signaling pathways.

Suppression of Photosynthetic Gene Expression in Roots Is Required for Sustained Root Growth under Phosphate Deficiency.

Functional disruption of tyrosylprotein sulfotransferase in Arabidopsis activates photosynthetic gene expression in roots and enhances inhibition of root growth under phosphate deficiency. Plants cope with inorganic phosphate (Pi) deficiencies in their environment by adjusting their developmental programs and metabolic activities. For Arabidopsis (Arabidopsis thaliana), the developmental responses include the inhibition of primary root growth and the enhanced formation of lateral roots and root hairs. Pi deficiency also inhibits photosynthesis by suppressing the expression of photosynthetic genes. Early studies showed that photosynthetic gene expression was also suppressed in Pi-deficient roots, a nonphotosynthetic organ; however, the biological relevance of this phenomenon remains unknown. In this work, we characterized an Arabidopsis mutant, hypersensitive to Pi starvation7 (hps7), that is hypersensitive to Pi deficiency; the hypersensitivity includes an increased inhibition of root growth. HPS7 encodes a tyrosylprotein sulfotransferase. Accumulation of HPS7 proteins in root tips is enhanced by Pi deficiency. Comparative RNA sequencing analyses indicated that the expression of many photosynthetic genes is activated in roots of hps7. Under Pi deficiency, the expression of photosynthetic genes in hps7 is further increased, which leads to enhanced accumulation of chlorophyll, starch, and sucrose. Pi-deficient hps7 roots also produce a high level of reactive oxygen species. Previous research showed that the overexpression of GOLDEN-like (GLK) transcription factors in transgenic Arabidopsis activates photosynthesis in roots. The GLK overexpressing (GLK OX) lines also exhibit increased inhibition of root growth under Pi deficiency. The increased inhibition of root growth in hps7 and GLK OX lines by Pi deficiency was completely reversed by growing the plants in the dark. Based on these results, we propose that suppression of photosynthetic gene expression is required for sustained root growth under Pi deficiency.

In vivo FRET–FLIM reveals cell-type-specific protein interactions in Arabidopsis roots

During multicellular development, specification of distinct cell fates is often regulated by the same transcription factors operating differently in distinct cis-regulatory modules, either through different protein complexes, conformational modification of protein complexes, or combinations of both. Direct visualization of different transcription factor complex states guiding specific gene expression programs has been challenging. Here we use in vivo FRET–FLIM (Förster resonance energy transfer measured by fluorescence lifetime microscopy) to reveal spatial partitioning of protein interactions in relation to specification of cell fate. We show that, in Arabidopsis roots, three fully functional fluorescently tagged cell fate regulators establish cell-type-specific interactions at endogenous expression levels and can form higher order complexes. We reveal that cell-type-specific in vivo FRET–FLIM distributions reflect conformational changes of these complexes to differentially regulate target genes and specify distinct cell fates.

Mediator subunit MED31 is required for radial patterning of Arabidopsis roots

Significance SHORTROOT (SHR) and SCARECROW (SCR) orchestrate a transcriptional program that is essential for ground tissue patterning. The regulation of the transcriptional output of SHR/SCR remains unclear. We show that MED31, a subunit of the plant Mediator coactivator complex, bridges the communication between SHR/SCR and the RNA polymerase II general transcriptional machinery. MED31 and SHR bind to the same protein domain of SCR, and the relative abundance of MED31 and SHR determines the dynamic formation of a MED31/SCR/SHR ternary complex. We studied the role of the MED31/SCR/SHR ternary complex in controlling the spatiotemporal expression of CYCLIND6;1 during ground tissue patterning. This study provides insight into the mechanisms by which master transcriptional regulators control organ patterning. Stem cell specification in multicellular organisms relies on the precise spatiotemporal control of RNA polymerase II (Pol II)-dependent gene transcription, in which the evolutionarily conserved Mediator coactivator complex plays an essential role. In Arabidopsis thaliana, SHORTROOT (SHR) and SCARECROW (SCR) orchestrate a transcriptional program that determines the fate and asymmetrical divisions of stem cells generating the root ground tissue. The mechanism by which SHR/SCR relays context-specific regulatory signals to the Pol II general transcription machinery is unknown. Here, we report the role of Mediator in controlling the spatiotemporal transcriptional output of SHR/SCR during asymmetrical division of stem cells and ground tissue patterning. The Mediator subunit MED31 interacted with SCR but not SHR. Reduction of MED31 disrupted the spatiotemporal activation of CYCLIND6;1 (CYCD6;1), leading to defective asymmetrical division of stem cells generating ground tissue. MED31 was recruited to the promoter of CYCD6;1 in an SCR-dependent manner. MED31 was involved in the formation of a dynamic MED31/SCR/SHR ternary complex through the interface protein SCR. We demonstrate that the relative protein abundance of MED31 and SHR in different cell types regulates the dynamic formation of the ternary complex, which provides a tunable switch to strictly control the spatiotemporal transcriptional output. This study provides valuable clues to understand the mechanism by which master transcriptional regulators control organ patterning.

Transmission electron microscopy study of fine a-axis-oriented grains as flux-pinning centers in YBa2Cu3O7-x/ZrO2(Y) thin film

Abstract YBa 2 Cu 3 O 7-x /ZrO 2 (Y) thin film with main c-axis-orientation produced by DC magnetron sputtering was observed by transmission electron microscopy. A high density of a -axis-oriented grains was found embedding in the thin film. The angular dependence of J c and microstructure study show that the boundaries of a -axis-oriented grains and c -axis-oriented grains are effective flux pinning centers in the film.