Songguang Yang

HD2C interacts with HDA6 and is involved in ABA and salt stress response in Arabidopsis

HD2 proteins are plant specific histone deacetylases. Four HD2 proteins, HD2A, HD2B, HD2C, and HD2D, have been identified in Arabidopsis. It was found that the expression of HD2A, HD2B, HD2C, and HD2D was repressed by ABA and NaCl. To investigate the function of HD2 proteins further, two HD2C T-DNA insertion lines of Arabidopsis, hd2c-1 and hd2c-3 were identified. Compared with wild-type plants, hd2c-1 and hd2c-3 plants displayed increased sensitivity to ABA and NaCl during germination and decreased tolerance to salt stress. These observations support a role of HD2C in the ABA and salt-stress response in Arabidopsis. Moreover, it was demonstrated that HD2C interacted physically with a RPD3-type histone deacetylase, HDA6, and bound to histone H3. The expression of ABA-responsive genes, ABI1 and ABI2, was increased in hda6, hd2c, and hda6/hd2c-1 double mutant plants, which was associated with increased histone H3K9K14 acetylation and decreased histone H3K9 dimethylation. Taken together, our results suggested that HD2C functionally associates with HDA6 and regulates gene expression through histone modifications.

PHYTOCHROME INTERACTING FACTOR3 Associates with the Histone Deacetylase HDA15 in Repression of Chlorophyll Biosynthesis and Photosynthesis in Etiolated Arabidopsis Seedlings

PIF3 recruits HDA15 to repress chlorophyll biosynthesis and photosynthesis gene expression in etiolated seedlings, suggesting that epigenetic regulation is critical for phytochrome- and PIF3-regulated light signal transduction. This study provides insight into the molecular mechanism underlying photomorphogenesis in plants. PHYTOCHROME INTERACTING FACTOR3 (PIF3) is a key basic helix-loop-helix transcription factor of Arabidopsis thaliana that negatively regulates light responses, repressing chlorophyll biosynthesis, photosynthesis, and photomorphogenesis in the dark. However, the mechanism for the PIF3-mediated transcription regulation remains largely unknown. In this study, we found that the REDUCED POTASSIUM DEPENDENCY3/HISTONE DEACETYLASE1-type histone deacetylase HDA15 directly interacted with PIF3 in vivo and in vitro. Genome-wide transcriptome analysis revealed that HDA15 acts mainly as a transcriptional repressor and negatively regulates chlorophyll biosynthesis and photosynthesis gene expression in etiolated seedlings. HDA15 and PIF3 cotarget to the genes involved in chlorophyll biosynthesis and photosynthesis in the dark and repress gene expression by decreasing the acetylation levels and RNA Polymerase II–associated transcription. The binding of HDA15 to the target genes depends on the presence of PIF3. In addition, PIF3 and HDA15 are dissociated from the target genes upon exposure to red light. Taken together, our results indicate that PIF3 associates with HDA15 to repress chlorophyll biosynthetic and photosynthetic genes in etiolated seedlings.

HISTONE DEACETYLASE19 interacts with HSL1 and participates in the repression of seed maturation genes in Arabidopsis seedlings.

HDA19 interacts with HSL1 and together they negatively regulate seed maturation gene expression in vegetative organs, suggesting that epigenetic regulation is critical for seed development. This study provides insight into the molecular mechanism underlying the gene regulation in seed development. The seed maturation genes are specifically and highly expressed during late embryogenesis. In this work, yeast two-hybrid, bimolecular fluorescence complementation, and coimmunoprecipitation assays revealed that HISTONE DEACETYLASE19 (HDA19) interacted with the HIGH-LEVEL EXPRESSION OF SUGAR-INDUCIBLE GENE2-LIKE1 (HSL1), and the zinc-finger CW [conserved Cys (C) and Trp (W) residues] domain of HSL1 was responsible for the interaction. Furthermore, we found that mutations in HDA19 resulted in the ectopic expression of seed maturation genes in seedlings, which was associated with increased levels of gene activation marks, such as Histone H3 acetylation (H3ac), Histone H4 acetylation (H4ac), and Histone H3 Lys 4 tri-methylation (H3K4me3), but decreased levels of the gene repression mark Histone H3 Lys 27 tri-methylation (H3K27me3) in the promoter and/or coding regions. In addition, elevated transcription of certain seed maturation genes was also found in the hsl1 mutant seedlings, which was also accompanied by the enrichment of gene activation marks but decreased levels of the gene repression mark. Chromatin immunoprecipitation assays showed that HDA19 could directly bind to the chromatin of the seed maturation genes. These results suggest that HDA19 and HSL1 may act together to repress seed maturation gene expression during germination. Further genetic analyses revealed that the homozygous hsl1 hda19 double mutants are embryonic lethal, suggesting that HDA19 and HSL1 may play a vital role during embryogenesis.

Involvement of Histone Modifications in Plant Abiotic Stress Responses

As sessile organisms, plants encounter various environmental stimuli including abiotic stresses during their lifecycle. To survive under adverse conditions, plants have evolved intricate mechanisms to perceive external signals and respond accordingly. Responses to various stresses largely depend on the plant capacity to modulate the transcriptome rapidly and specifically. A number of studies have shown that the molecular mechanisms driving the responses of plants to environmental stresses often depend on nucleosome histone post-translational modifications including histone acetylation, methylation, ubiquitination, and phosphorylation. The combined effects of these modifications play an essential role in the regulation of stress responsive gene expression. In this review, we highlight our current understanding of the epigenetic mechanisms of histone modifications and their roles in plant abiotic stress response.

Molecular characterization of a rice metal tolerance protein, OsMTP1

Rice (Oryza sativa L. ‘Nipponbare’) cDNA subtractive suppression hybridization (SSH) libraries constructed using cadmium (Cd)-treated seedling roots were screened to isolate Cd-responsive genes. A cDNA clone, encoding the rice homolog of Metal Tolerance Protein (OsMTP1), was induced by Cd treatment. Plant MTPs belong to cation diffusion facilitator (CDF) protein family, which are widespread in bacteria, fungi, plants, and animals. OsMTP1 heterologous expression in yeast mutants showed that OsMTP1 was able to complement the mutant strains’ hypersensitivity to Ni, Cd, and Zn, but not other metals including Co and Mn. OsMTP1 expression increased tolerance to Zn, Cd, and Ni in wild-type yeast BY4741 during the exponential growth phase. OsMTP1 fused to green fluorescent protein was localized in onion epidermal cell plasma membranes, consistent with an OsMTP1 function in heavy metal transporting. OsMTP1 dsRNAi mediated by transgenic assay in rice seedlings resulted in heavy metal sensitivity and changed the heavy metal accumulation in different organs of mature rice under low-concentration heavy metal stress. Taken together, our results show that OsMTP1 is a bivalent cation transporter localized in the cell membrane, which is necessary for efficient translocation of Zn, Cd and other heavy metals, and maintain ion homeostasis in plant.

The Arabidopsis SWI2/SNF2 Chromatin Remodeler BRAHMA Regulates Polycomb Function during Vegetative Development and Directly Activates the Flowering Repressor Gene SVP

The chromatin remodeler BRAHMA (BRM) is a Trithorax Group (TrxG) protein that antagonizes the functions of Polycomb Group (PcG) proteins in fly and mammals. Recent studies also implicate such a role for Arabidopsis (Arabidopsis thaliana) BRM but the molecular mechanisms underlying the antagonism are unclear. To understand the interplay between BRM and PcG during plant development, we performed a genome-wide analysis of trimethylated histone H3 lysine 27 (H3K27me3) in brm mutant seedlings by chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq). Increased H3K27me3 deposition at several hundred genes was observed in brm mutants and this increase was partially supressed by removal of the H3K27 methyltransferase CURLY LEAF (CLF) or SWINGER (SWN). ChIP experiments demonstrated that BRM directly binds to a subset of the genes and prevents the inappropriate association and/or activity of PcG proteins at these loci. Together, these results indicate a crucial role of BRM in restricting the inappropriate activity of PcG during plant development. The key flowering repressor gene SHORT VEGETATIVE PHASE (SVP) is such a BRM target. In brm mutants, elevated PcG occupancy at SVP accompanies a dramatic increase in H3K27me3 levels at this locus and a concomitant reduction of SVP expression. Further, our gain- and loss-of-function genetic evidence establishes that BRM controls flowering time by directly activating SVP expression. This work reveals a genome-wide functional interplay between BRM and PcG and provides new insights into the impacts of these proteins in plant growth and development.

Concerted genomic targeting of H3K27 demethylase REF6 and chromatin-remodeling ATPase BRM in Arabidopsis

SWI/SNF-type chromatin remodelers, such as BRAHMA (BRM), and H3K27 demethylases both have active roles in regulating gene expression at the chromatin level, but how they are recruited to specific genomic sites remains largely unknown. Here we show that RELATIVE OF EARLY FLOWERING 6 (REF6), a plant-unique H3K27 demethylase, targets genomic loci containing a CTCTGYTY motif via its zinc-finger (ZnF) domains and facilitates the recruitment of BRM. Genome-wide analyses showed that REF6 colocalizes with BRM at many genomic sites with the CTCTGYTY motif. Loss of REF6 results in decreased BRM occupancy at BRM–REF6 co-targets. Furthermore, REF6 directly binds to the CTCTGYTY motif in vitro, and deletion of the motif from a target gene renders it inaccessible to REF6 in vivo. Finally, we show that, when its ZnF domains are deleted, REF6 loses its genomic targeting ability. Thus, our work identifies a new genomic targeting mechanism for an H3K27 demethylase and demonstrates its key role in recruiting the BRM chromatin remodeler.

The Arabidopsis SWI2/SNF2 Chromatin Remodeling ATPase BRAHMA Targets Directly to PINs and Is Required for Root Stem Cell Niche Maintenance

The Arabidopsis SWI/SNF chromatin-remodeling ATPase BRAHMA acts to maintain the stem cell niche in roots by modulating auxin distribution via regulation of the expression of PINs. BRAHMA (BRM), a SWI/SNF chromatin remodeling ATPase, is essential for the transcriptional reprogramming associated with development and cell differentiation in Arabidopsis thaliana. In this study, we show that loss-of-function mutations in BRM led to defective maintenance of the root stem cell niche, decreased meristematic activity, and stunted root growth. Mutations of BRM affected auxin distribution by reducing local expression of several PIN-FORMED (PIN) genes in the stem cells and impaired the expression of the stem cell transcription factor genes PLETHORA (PLT1) and PLT2. Chromatin immunoprecipitation assays showed that BRM could directly target to the chromatin of PIN1, PIN2, PIN3, PIN4, and PIN7. In addition, genetic interaction assays indicate that PLTs acted downstream of BRM, and overexpression of PLT2 partially rescued the stem cell niche defect of brm mutants. Taken together, these results support the idea that BRM acts in the PLT pathway to maintain the root stem cell niche by altering the expression of PINs.

AtACDO1, an ABC1-like kinase gene, is involved in chlorophyll degradation and the response to photooxidative stress in Arabidopsis

ABC1 (activity of bc1 complex) is a newly discovered atypical kinase in plants. Here, it is reported that an ABC1 protein kinase-encoded gene, AtACDO1 (ABC1-like kinase related to chlorophyll degradation and oxidative stress), located in chloroplasts, was up-regulated by methyl viologen (MV) treatment. AtACDO1 RNAi (RNA interference) plants showed developmental defects, including yellow-green leaves and reduced contents of carotenoids and chlorophyll; the chlorophyll reduction was associated with a change in the numbers of chlorophyll-binding proteins of the photosynthetic complexes. Chlorophyllide (Chlide) a the first product of chlorophyll degradation, and pheophorbide a, a subsequent intermediate of Chlide a degradation, were increased in AtACDO1 RNAi plants. The AtACDO1 RNAi plants were more sensitive to high light and MV than wild-type plants. The AtACDO1 RNAi plants had lower transcript levels of the oxidative stress response genes FSD1, CSD1, CAT1, and UTG71C1 after MV treatment compared with wild-type or 35S::AtACDO1 plants. Taken together, the results suggest that the chloroplast AtACDO1 protein plays important roles in mediating chlorophyll degradation and maintaining the number of chlorophyll-binding photosynthetic thylakoid membranes, as well as in the photooxidative stress response.

Arabidopsis BREVIPEDICELLUS interacts with the SWI2/SNF2 chromatin remodeling ATPase BRAHMA to regulate KNAT2 and KNAT6 expression in control of inflorescence architecture.

BREVIPEDICELLUS (BP or KNAT1), a class-I KNOTTED1-like homeobox (KNOX) transcription factor in Arabidopsis thaliana, contributes to shaping the normal inflorescence architecture through negatively regulating other two class-I KNOX genes, KNAT2 and KNAT6. However, the molecular mechanism of BP-mediated transcription regulation remains unclear. In this study, we showed that BP directly interacts with the SWI2/SNF2 chromatin remodeling ATPase BRAHMA (BRM) both in vitro and in vivo. Loss-of-function BRM mutants displayed inflorescence architecture defects, with clustered inflorescences, horizontally orientated pedicels, and short pedicels and internodes, a phenotype similar to the bp mutants. Furthermore, the transcript levels of KNAT2 and KNAT6 were elevated in brm-3, bp-9 and brm-3 bp-9 double mutants. Increased histone H3 lysine 4 tri-methylation (H3K4me3) levels were detected in brm-3, bp-9 and brm-3 bp-9 double mutants. Moreover, BRM and BP co-target to KNAT2 and KNAT6 genes, and BP is required for the binding of BRM to KNAT2 and KNAT6. Taken together, our results indicate that BP interacts with the chromatin remodeling factor BRM to regulate the expression of KNAT2 and KNAT6 in control of inflorescence architecture.