Mechanism of Crosstalk between the LSD1 Demethylase and HDAC1 Deacetylase in the CoREST Complex

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Histone deacetylase OsHDA706 increases salt tolerance via H4K5/K8 deacetylation of OsPP2C49 in rice

High salt is a major environmental factor that threatens plant growth and development. Increasing evidence indicates that histone acetylation is involved in plant responses to various abiotic stress; however, the underlying epigenetic regulatory mechanisms remain poorly understood. In this study, we revealed that the histone deacetylase OsHDA706 epigenetically regulates the expression of salt stress response genes in rice (Oryza sativa L.). OsHDA706 localizes to the nucleus and cytoplasm and OsHDA706 expression is significantly induced under salt stress. Moreover, oshda706 mutants showed a higher sensitivity to salt stress than the wild-type. In vivo and in vitro enzymatic activity assays demonstrated that OsHDA706 specifically regulates the deacetylation of lysines 5 and 8 on histone H4 (H4K5 and H4K8). By combining chromatin immunoprecipitation and mRNA sequencing, we identified the clade A protein phosphatase 2 C gene, OsPP2C49, which is involved in the salt response as a direct target of H4K5 and H4K8 acetylation. We found that the expression of OsPP2C49 is induced in the oshda706 mutant under salt stress. Furthermore, the knockout of OsPP2C49 enhances plant tolerance to salt stress, while its overexpression has the opposite effect. Taken together, our results indicate that OsHDA706, a histone H4 deacetylase, participates in the salt stress response by regulating the expression of OsPP2C49 via H4K5 and H4K8 deacetylation.     

组蛋白去乙酰化酶3对外周CD4+ T细胞分化的影响

目的 探讨组蛋白去乙酰化酶3（HDAC3）对外周CD4⁺ T细胞分化及功能的调控作用。方法 采用CD4cre酶介导Hdac3杂合基因缺失小鼠（Hdac3^fl/flCD4^cre+/-）及其野生型正常对照小鼠（Hdac3^fl/fl，WT），流式细胞术检测HDAC3缺失对外周CD4⁺和CD8⁺ T细胞比例和数量的影响；在体外佛波酯（PMA）和离子霉素（Ionomycin）刺激条件下，流式细胞术检测HDAC3缺失对CD4⁺ T细胞中IFN-γ、IL-4和IL-17A的表达以及Tfh细胞产生的影响；采用ELISA检测HDAC3缺失对小鼠血清IFN-γ、IL-4和IL-17表达的影响；分选Hdac3^fl/flCD4^cre+/-和WT小鼠外周初始CD4⁺ T细胞，分别在Th1和Th2分化条件下培养，细胞内染色检测HDAC3缺失对Th1、Th2以及Th17相关细胞因子及其特异转录因子表达的影响；采用Microarray检测HDAC3缺失对CD4⁺ T细胞分化亚群相关基因表达的影响；采用链脲佐菌素（STZ）处理小鼠构建I型糖尿病（TIDM）疾病模型，检测HDAC3缺失对T1DM发病的影响。结果 与WT小鼠相比，Hdac3^fl/flCD4^cre+/-小鼠外周CD4⁺和CD8⁺ T细胞的比例和数量显著降低。Hdac3^fl/flCD4^cre+/-小鼠CD4⁺ T细胞及血清中IFN-γ的表达显著降低，而IL-4和IL-17A的表达显著增加，Tfh细胞比例也显著增加；HDAC3缺失抑制体外培养CD4⁺ T细胞向Th1分化但促进其向Th2分化；Microarray检测发现HDAC3缺失导致Th1型细胞谱系基因表达降低，而Th2、Th17以及Tfh细胞谱系基因表达增加；在STZ诱导条件下，HDAC3缺失抑制小鼠T1DM的发生和CD4⁺ T细胞向Th1分化。结论 HDAC3促进外周CD4⁺ T细胞向Th1细胞分化并加重T1DM的发生。     

Restoring Histone Deacetylase Activity by Waste Product Release. A View from Molecular Mechanics Simulations with Mammalian HDAC8

HDAC8 is a Zn^II‐based, single‐peptide mammalian histone deacetylase that is localized mainly in the cytoskeleton of smooth muscle cells, thus regulating muscle contractility. HDACs are also widely involved in cellular processes, ranging from cell differentiation to proliferation, senescence, and apoptosis; in particular, protecting a telomerase activator from ubiquitin‐mediated degradation. How HDACs can eliminate the hydrolytic reaction products, in order that the process of deacetylation of the acetyllysine moiety of histones can take place again, has long been debated in the scientific literature, without reaching any firm conclusion, however. This question is the subject of the present work, carried out along a theoretical line that is capable of describing the whole pathway followed by the acetate product (ACT). A model was built here on the crystal data for the Y306F‐mutated HDAC8 complex with a diacetylated peptide of the p53‐tumor‐suppressor class. That was followed by manually hydrolyzing the acetylated moiety bound to Zn^II and discharging the monoacetylated peptide product (MAP). The latter was replaced by a H₂O molecule bound to Zn^II, while ACT was left free in the reaction cage. This Zn^II cluster was DFT‐parameterized for the ff99SB force field without any further bias. As the result of random‐acceleration molecular dynamics (RAMD) simulations, egress of ACT from the reaction cage toward the aqueous environment can follow three pathways. Two of them utilize the channel for peptide (or histone) uptake and are preferred, if ACT leaves the reaction center before MAP (or the deacetylated histone). The third pathway, developing along the internal channel, is available to ACT even if MAP is still in place.     

Structural insights into recognition of acetylated histone ligands by the BRPF1 bromodomain

Bromodomain-PHD finger protein 1 (BRPF1) is part of the MOZ HAT complex and contains a unique combination of domains typically found in chromatin-associated factors, which include plant homeodomain (PHD) fingers, a bromodomain and a proline-tryptophan-tryptophan-proline (PWWP) domain. Bromodomains are conserved structural motifs generally known to recognize acetylated histones, and the BRPF1 bromodomain preferentially selects for H2AK5ac, H4K12ac and H3K14ac. We solved the X-ray crystal structures of the BRPF1 bromodomain in complex with the H2AK5ac and H4K12ac histone peptides. Site-directed mutagenesis on residues in the BRPF1 bromodomain-binding pocket was carried out to investigate the contribution of specific amino acids on ligand binding. Our results provide critical insights into the molecular mechanism of ligand binding by the BRPF1 bromodomain, and reveal that ordered water molecules are an essential component driving ligand recognition.     

Anti-tumor activity evaluation of novel tubulin and HDAC dual-targeting inhibitors

Histone deacetylases (HDACs) have proven to be promising targets for the development of anti-cancer drugs. In this study, we reported a series of novel chalcone based tubulin and HDAC dual-targeting inhibitors. Three compounds inhibited the activities of HDAC and tubulin polymerization simultaneously and displayed anti-proliferative activities toward eleven human tumor cell lines. Compound 8a remarkably induced growth inhibition, apoptosis and G2/M phase arrest of A549 tumor cells. Finally, the inhibitory activities of 8a against HDAC6 and tubulin were rationalized by molecular docking studies.     

The synergistic effects of valproic acid and fluvastatin on apoptosis induction in glioblastoma multiforme cell lines

Glioblastoma multiforme (GBM) is the most common primary central nervous system malignant tumor. It responds poorly to standard therapies, such as surgical resection, radiation therapy and chemotherapy. Many chemotherapeutic drugs are focused on apoptosis induction and radiation sensitivity. Inhibition of histone acetylation via histone deacetylase inhibitor (HDACI) is one such strategy. Statins (or 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors) are classical drugs used to lower cholesterol but also inhibitors of histone deacetylation activity. This study analyzes the combinatory effects of valproic acid (VPA) and fluvastatin on apoptosis induction in GBM8401 cells. The results show that they act synergistically in inducing γ-H2AX and apoptosis accompanied by higher acetylated histones H3 and H4. Downregulation of p53 occurred by VPA alone and fluvastatin alone, but not at their combined application; upregulation of p21 at the protein level was induced by each of the drugs alone and no further increase occurred at combined application. The drug BEZ235 inhibited phosphorylation of Akt and attenuated the level of γ-H2AX as well as cleaved PARP (cPARP) induced at combined application of VPA and fluvastatin. Induction of apoptosis within a 48 h incubation period was massive when measured as the subG1 peak (97%) and was detected after a 24 h incubation at low level when assayed with PE Annexin V. Synergistic apoptosis induction was demonstrated also after 24 h incubation by the appearance of cPARP. Partial silencing of p21 reduced cPARP as well as the percentage of apoptotic cells in the subG1 peak. However, partial silencing of p53 had no effect on apoptosis. Such findings offer a better understanding of the mechanism of action of HDACIs in combination with statins that may guide the development of a new combinatory reposition for the treatment of GBM.