Intracellular localization of histone deacetylase HDA6 in plants

Journal of Plant Research - Histone modification is an important epigenetic mechanism in eukaryotes. Histone acetyltransferase and deacetylase regulate histone acetylation levels antagonistically,...     

Regulation of flowering time by the histone deacetylase HDA5 in Arabidopsis

The acetylation level of histones on lysine residues regulated by histone acetyltransferases and histone deacetylases plays an important but under‐studied role in the control of gene expression in plants. With the aim of characterizing the Arabidopsis RPD3/HDA1 family histone deacetylase HDA5, we present evidence showing that HDA5 displays deacetylase activity. Mutants defective in the expression of HDA5 displayed a late‐flowering phenotype. Expression of the flowering repressor genes FLC and MAF1 was up‐regulated in hda5 mutants. Furthermore, the gene activation markers, histone H3 acetylation and H3K4 trimethylation on FLC and MAF1 chromatin were increased in hda5‐1 mutants. Chromatin immunoprecipitation analysis showed that HDA5 binds to the chromatin of FLC and MAF1. Bimolecular fluorescence complementation assays and co‐immunoprecipitation assays showed that HDA5 interacts with FVE, FLD and HDA6, indicating that these proteins are present in a protein complex involved in the regulation of flowering time. Comparing gene expression profiles of hda5 and hda6 mutants by RNA‐seq revealed that HDA5 and HDA6 co‐regulate gene expression in multiple development processes and pathways.     

The histone deacetylase inhibitor sodium butyrate inhibits baculovirus-mediated transgene expression in Sf9 cells

Recent studies have indicated that histone deacetylase inhibitors (HDACis) could enhance and prolong expression of exogenous genes delivered by various viral vehicles in mammalian cells, including baculovirus vectors. In this study, the effects of HDACis on expression of a baculovirus-mediated eGFP reporter gene under control of baculovirus late promoter p10 in Sf9 cells were evaluated. It was found that sodium butyrate (NaBu) decreased the expression level of the target gene driven by p10 promoter by four to fivefold. Moreover, addition of NaBu increased DNaseI-sensitivity of transgene p10 promoter region and did not influence viral DNA replication. FACS assay has shown that both NaBu and fluorodeoxyuridine (FdUrd) blocked Sf9 cells at G1 phase and inhibited the target gene expression. Another HDACi, trichostatin, had little effects on both cell cycle and Ac-p10-eGFP expression, strongly suggesting that cell cycle arrest accounts for the mechanisms by which NaBu inhibits Ac-p10-eGFP expression. The inhibiting effects of NaBu on baculovirus transgene expression in Sf9 cells are promoter specific since the enhancement of NaBu on transgene expression in insect and mammalian cells are mediated by baculovirus harboring a murine cytomegalovirus (mCMV) immediate early promoter. This study was aimed at improving the productivity of the recombinant proteins and providing a better understanding of the epigenetic regulation of baculovirus gene expression.     

Arabidopsis HDA6 is required for freezing tolerance

The control of energy homeostasis within the hypothalamus is under the regulated control of homeostatic hormones, nutrients and the expression of neuropeptides that alter feeding behavior. Elevated levels of palmitate, a predominant saturated fatty acid in diet and fatty acid biosynthesis, alter cellular function. For instance, a key mechanism involved in the development of insulin resistance is lipotoxicity, through increased circulating saturated fatty acids. Although many studies have begun to determine the underlying mechanisms of lipotoxicity in peripheral tissues, little is known about the effects of excess lipids in the brain. To determine these mechanisms we used an immortalized, clonal, hypothalamic cell line, mHypoE-44, to demonstrate that palmitate directly alters the expression of molecular clock components, by increasing Bmal1 and Clock, or by decreasing Per2, and Rev-erbα, their mRNA levels and altering their rhythmic period within individual neurons. We found that these neurons endogenously express the orexigenic neuropeptides NPY and AgRP, thus we determined that palmitate administration alters the mRNA expression of these neuropeptides as well. Palmitate treatment causes a significant increase in NPY mRNA levels and significantly alters the phase of rhythmic expression. We explored the link between AMPK and the expression of neuropeptide Y using the AMPK inhibitor compound C and the AMP analog AICAR. AMPK inhibition decreased NPY mRNA. AICAR also elevated basal NPY, but prevented the palmitate-mediated increase in NPY mRNA levels. We postulate that this palmitate-mediated increase in NPY and AgRP synthesis may initiate a detrimental positive feedback loop leading to increased energy consumption.     

Tissue-dependent enhancement of transgene expression by introns of replacement histone H3 genes of Arabidopsis

Intron-bearing replacement histone H3 genes in Arabidopsis and other plants are highly and constitutively expressed. We demonstrate that the introns located within the 5-untranslated regions (5-UTR) of the two Arabidopsis replacement H3 genes will abolish the cell cycle dependence of an endogenous histone H4 promoter. We demonstrate that these introns, functionally combined with their endogenous promoters, could produce the high and constitutive expression of the replacement H3 genes observed in planta. They strongly increase gene expression whatever the promoter, from the strong 35S CaMV promoter to complete and resected promoters of cell cycle-dependent and replacement histone genes. Quantitative analysis of the extent of reporter gene enhancement in different parts of developing transgenic plantlets, ranging from 2-fold to 70-fold, supports the notion that trans-acting factors are responsible for this effect. Such factors appear most abundant in roots.     

Altered levels of histone deacetylase OsHDT1 affect differential gene expression patterns in hybrid rice

Hybrids between different inbred varieties display novel patterns of gene expression resulted from parental variation in allelic nucleotide sequences. To study the function of chromatin regulators in hybrid gene expression, the histone deacetylase gene OsHDT1 whose expression displayed a circadian rhythm was over-expressed or inactivated by RNAi in an elite rice parent. Increased OsHDT1 expression did not affect plant growth in the parent but led to early flowering in the hybrid. Nonadditive up-regulation of key flowering time genes was found to be related to flowering time of the hybrid. Over-expression of OsHDT1 repressed the nonadditive expression of the key flowering repressors in the hybrid (i.e. OsGI and Hd1) inducing early flowering. Analysis of histone acetylation suggested that OsHDT1 over-expression might promote deacetylation on OsGI and Hd1 chromatin during the peak expression phase. High throughput differential gene expression analysis revealed that altered OsHDT1 levels affected nonadditive expression of many genes in the hybrid. These data demonstrate that nonadditive gene expression was involved in flowering time control in the hybrid rice and that OsHDT1 level was important for nonadditive or differential expression of many genes including the flowering time genes, suggesting that OsHDT1 may be involved in epigenetic control of parental genome interaction for differential gene expression.     

Identification of Arabidopsis rat mutants

Limited knowledge currently exists regarding the roles of plant genes and proteins in the Agrobacterium tumefaciens-mediated transformation process. To understand the host contribution to transformation, we carried out root-based transformation assays to identify Arabidopsis mutants that are resistant to Agrobacterium transformation (rat mutants). To date, we have identified 126 rat mutants by screening libraries of T-DNA insertion mutants and by using various “reverse genetic” approaches. These mutants disrupt expression of genes of numerous categories, including chromatin structural and remodeling genes, and genes encoding proteins implicated in nuclear targeting, cell wall structure and metabolism, cytoskeleton structure and function, and signal transduction. Here, we present an update on the identification and characterization of these rat mutants.     

Epigenetic interplay of histone modifications and DNA methylation mediated by HDA6

One of the most fundamental questions in the control of gene expression is how epigenetic patterns of DNA methylation and histone modifications are established. Our recent studies demonstrate that histone deacetylase HDA6 integrates DNA methylation and histone modifications in gene silencing by interacting with DNA methyltransferase MET1 and histone demethylase FLD, suggesting that regulatory crosstalk between histone modifications and DNA methylation could be mediated by the interaction of various epigenetic modification proteins.