RNA 5-Methylcytosine Modification Regulates Vegetative Development Associated with H3K27 Trimethylation in Arabidopsis.

Methylating RNA post-transcriptionally is emerging as a significant mechanism of gene regulation in eukaryotes. The crosstalk between RNA methylation and histone modification is critical for chromatin state and gene expression in mammals. However, it is not well understood mechanistically in plants. Here, the authors report a genome-wide correlation between RNA 5-cytosine methylation (m5 C) and histone 3 lysine27 trimethylation (H3K27me3) in Arabidopsis. The plant-specific Polycomb group (PcG) protein EMBRYONIC FLOWER1 (EMF1) plays dual roles as activators or repressors. Transcriptome-wide RNA m5 C profiling revealed that m5 C peaks are mostly enriched in chromatin regions that lacked H3K27me3 in both wild type and emf1 mutants. EMF1 repressed the expression of m5 C methyltransferase tRNA specific methyltransferase 4B (TRM4B) through H3K4me3, independent of PcG-mediated H3K27me3 mechanism. The 5-Cytosine methylation on targets is increased in emf1 mutants, thereby decreased the mRNA transcripts of photosynthesis and chloroplast genes. In addition, impairing EMF1 activity reduced H3K27me3 levels of PcG targets, such as starch genes, which are de-repressed in emf1 mutants. Both EMF1-mediated promotion and repression of gene activities via m5 C and H3K27me3 are required for normal vegetative growth. Collectively, t study reveals a previously undescribed epigenetic mechanism of RNA m5 C modifications and histone modifications to regulate gene expression in eukaryotes.

Trithorax Group Proteins Act Together with a Polycomb Group Protein to Maintain Chromatin Integrity for Epigenetic Silencing during Seed Germination in Arabidopsis.

Polycomb group (PcG) and trithorax group (trxG) proteins have been shown to act antagonistically to epigenetically regulate gene expression in eukaryotes. The trxG proteins counteract PcG-mediated floral repression in Arabidopsis, but their roles in other developmental processes are poorly understood. We investigated the interactions between the trxG genes, ARABIDOPSIS HOMOLOG OF TRITHORAX1 (ATX1) and ULTRAPETALA1 (ULT1), and the PcG gene EMBRYONIC FLOWER 1 (EMF1) during early development. Unexpectedly, we found that mutations in the trxG genes failed to rescue the early-flowering phenotype of emf1 mutants. Instead, emf1 atx1 ult1 seedlings showed a novel swollen root phenotype and massive deregulation of gene expression. Greater ectopic expression of seed master regulatory genes in emf1 atx1 ult1 triple than in emf1 single mutants indicates that PcG and trxG factors together repress seed gene expression after germination. Furthermore, we found that the widespread gene derepression is associated with reduced levels of H3K27me3, an epigenetic repressive mark of gene expression, and with globally altered chromatin organization. EMF1, ATX1, and ULT1 are able to bind the chromatin of seed genes and ULT1 can physically interact with ATX1 and EMF1, suggesting that the trxG and EMF1 proteins directly associate at target gene loci for EMF1-mediated gene silencing. Thus, while ATX1, ULT1, and EMF1 interact antagonistically to regulate flowering, they work together to maintain chromatin integrity and prevent precocious seed gene expression after germination.

PcG and trxG in plants - friends or foes.

The highly-conserved Polycomb group (PcG) and trithorax group (trxG) proteins play major roles in regulating gene expression and maintaining developmental states in many organisms. However, neither the recruitment of Polycomb repressive complexes (PRC) nor the mechanisms of PcG and trxG-mediated gene silencing and activation are well understood. Recent progress in Arabidopsis research challenges the dominant model of PRC2-dependent recruitment of PRC1 to target genes. Moreover, evidence indicates that diverse forms of PRC1, with shared components, are a common theme in plants and mammals. Although trxG is known to antagonize PcG, emerging data reveal that trxG can also repress gene expression, acting cooperatively with PcG. We discuss these recent findings and highlight the employment of diverse epigenetic mechanisms during development in plants and animals.

EMBRYONIC FLOWER1 and ULTRAPETALA1 Act Antagonistically on Arabidopsis Development and Stress Response.

Epigenetic regulation of gene expression is of fundamental importance for eukaryotic development. EMBRYONIC FLOWER1 (EMF1) is a plant-specific gene that participates in Polycomb group-mediated transcriptional repression of target genes such as the flower MADS box genes AGAMOUS, APETALA3, and PISTILLATA. Here, we investigated the molecular mechanism underlying the curly leaf and early flowering phenotypes caused by reducing EMF1 activity in the leaf primordia of LFYasEMF1 transgenic plants and propose a combined effect of multiple flower MADS box gene activities on these phenotypes. ULTRAPETALA1 (ULT1) functions as a trithorax group factor that counteracts Polycomb group action in Arabidopsis (Arabidopsis thaliana). Removing ULT1 activity rescues both the abnormal developmental phenotypes and most of the misregulated gene expression of LFYasEMF1 plants. Reducing EMF1 activity increases salt tolerance, an effect that is diminished by introducing the ult1-3 mutation into the LFYasEMF1 background. EMF1 is required for trimethylating lysine-27 on histone 3 (H3K27me3), and ULT1 associates with ARABIDOPSIS TRITHORAX1 (ATX1) for trimethylating lysine-3 on histone 4 (H3K4me3) at flower MADS box gene loci. Reducing EMF1 activity decreases H3K27me3 marks and increases H3K4me3 marks on target gene loci. Removing ULT1 activity has the opposite effect on the two histone marks. Removing both gene activities restores the active and repressive marks to near wild-type levels. Thus, ULT1 acts as an antirepressor that counteracts EMF1 action through modulation of histone marks on target genes. Our analysis indicates that, instead of acting as off and on switches, EMF1 and ULT1 mediate histone mark deposition and modulate transcriptional activities of the target genes.

Molecular and functional characterization of broccoli EMBRYONIC FLOWER 2 genes.

Polycomb group (PcG) proteins regulate major developmental processes in Arabidopsis. EMBRYONIC FLOWER 2 (EMF2), the VEFS domain-containing PcG gene, regulates diverse genetic pathways and is required for vegetative development and plant survival. Despite widespread EMF2-like sequences in plants, little is known about their function other than in Arabidopsis and rice. To study the role of EMF2 in broccoli (Brassica oleracea var. italica cv. Elegance) development, we identified two broccoli EMF2 (BoEMF2) genes with sequence homology to and a similar gene expression pattern to that in Arabidopsis (AtEMF2). Reducing their expression in broccoli resulted in aberrant phenotypes and gene expression patterns. BoEMF2 regulates genes involved in diverse developmental and stress programs similar to AtEMF2 in Arabidopsis. However, BoEMF2 differs from AtEMF2 in the regulation of flower organ identity, cell proliferation and elongation, and death-related genes, which may explain the distinct phenotypes. The expression of BoEMF2.1 in the Arabidopsis emf2 mutant (Rescued emf2) partially rescued the mutant phenotype and restored the gene expression pattern to that of the wild type. Many EMF2-mediated molecular and developmental functions are conserved in broccoli and Arabidopsis. Furthermore, the restored gene expression pattern in Rescued emf2 provides insights into the molecular basis of PcG-mediated growth and development.

The elongation defective1 mutant of Arabidopsis is impaired in the gene encoding a serine-rich secreted protein.

Coordinated cell growth and differentiation is crucial for the development of higher plants. Using the elongation defective 1-1 (eld1-1) mutant, we cloned the ELD1 gene, which encodes a serine-rich protein. Genes homologous to ELD1 can be found in plants, including Arabidopsis, rice, and tobacco, but not in other organisms. Using reverse genetics, we identified a new allele, eld1-2, which is phenotypically indistinguishable from eld1-1, but does not produce a detectable ELD1 transcript. The ELD1 gene sequence is the same as that of the KOBITO1 sequence. However, the kob1 mutants display weak phenotype relative to the two eld1 mutants, which are likely null alleles. KOB1 was reported to be a membrane protein involved in cellulose synthesis. However, based on ELD1-GFP localization in plasmolyzed cells, we found that ELD1 is localized to the cell wall/extracellular matrix, rather than the membrane. Thus, ELD1/KOB1 is a secreted protein involved in promoting cell growth. To investigate the relationship between ELD1 and Arabidopsis genes with high sequence similarity, we analyzed the possible subcellular location of their proteins as well as their amino acid sequence. The ELD1-related proteins in Arabidopsis were predicted to be localized to subcellular compartments different from that of ELD1. Thus, ELD1 is likely to be functionally distinct from related Arabidopsis genes. These results suggest that ELD1 is a single-copy gene which belongs to a small family of plant-specific genes with diverse function.