Probing the elusive catalytic activity of vertebrate class IIa histone deacetylases

It has been widely debated whether class IIa HDACs have catalytic deacetylase activity, and whether this plays any part in controlling gene expression. Herein, it has been demonstrated that class IIa HDACs isolated from mammalian cells are contaminated with other deacetylases, but can be prepared cleanly in Escherichia coli. These bacteria preparations have weak but measurable deacetylase activity. The low efficiency can be restored either by: mutation of an active site histidine to tyrosine, or by the use of a non-acetylated lysine substrate, allowing the development of assays to identify class IIa HDAC inhibitors.     

Multiple histone deacetylases and the CREB-binding protein regulate pre-mRNA 3'-end processing

Trichostatin A (TSA), a specific inhibitor of histone deacetylases (HDACs), induces acetylation of various non-histone proteins such as p53 and alpha-tubulin. We purified several acetylated proteins by the affinity to an anti-acetylated lysine (AcLys) antibody from cells treated with TSA and identified them by mass spectrometry. Here we report on acetylation of CFIm25, a component of mammalian cleavage factor Im (CF Im), and poly(A) polymerase (PAP), a polyadenylating enzyme for the pre-mRNA 3'-end. The residues acetylated in these proteins were mapped onto the regions required for interaction with each other. Whereas CBP acetylated these proteins, HDAC1, HDAC3, HDAC10, SIRT1, and SIRT2 were involved in in vivo deacetylation. Acetylation of the CFIm25 occurred depending on the cleavage factor complex formation. Importantly, the interaction between PAP and CF Im complex was decreased by acetylation. We also demonstrated that acetylation of PAP inhibited the nuclear localization of PAP by inhibiting the binding to the importin alpha/beta complex. These results suggest that CBP and HDACs regulate the 3'-end processing machinery and modulate the localization of PAP through the acetylation and deacetylation cycle.     

Ⅱa型组蛋白脱乙酰基酶在心血管系统中的作用

组蛋白脱乙酰基酶(histone deacetylases, HDACs)具有转录抑制功能,近年来发现其参与心血管系统的分化发育,调控多种刺激诱发的心肌肥大及其相关基因表达.本文就有关Ⅱa型HDACs在心血管系统中作用的研究进展作一综述.     

Factors affecting the substrate specificity of histone deacetylases

Histone deacetylases (HDACs) catalyze the deacetylation of epsilon-acetyl-lysine residues within the N-terminal tail of core histones and thereby mediate changes in the chromatin structure and regulate gene expression in eukaryotic cells. So far, surprisingly little is known about the substrate specificities of different HDACs. Here, we prepared a library of fluorogenic tripeptidic substrates of the general format Ac-P(-2)-P(-1)-Lys(Ac)-MCA (P(-1), P(-2)=all amino acids except cysteine) and measured their HDAC-dependent conversion in a standard fluorogenic HDAC assay. Different HDAC subtypes can be ranked according to their substrate selectivity: HDAH > HDAC8 > HDAC1 > HDAC3 > HDAC6. HDAC1, HDAC3, and HDAC6 exhibit a similar specificity profile, whereas both HDAC8 and HDAH have rather distinct profiles. Furthermore, it was shown that second-site modification (e.g., phosphorylation) of substrate sequences as well as corepressor binding can modulate the selectivity of enzymatic substrate conversion.     

Targeting histone deacetylases for the treatment of disease

The 'histone code' is a well-established hypothesis describing the idea that specific patterns of post-translational modifications to histones act like a molecular 'code' recognized and used by non-histone proteins to regulate specific chromatin functions. One modification, which has received significant attention, is that of histone acetylation. The enzymes that regulate this modification are described as lysine acetyltransferases or KATs, and histone deacetylases or HDACs. Due to their conserved catalytic domain HDACs have been actively targeted as a therapeutic target. The pro-inflammatory environment is increasingly being recognized as a critical element for both degenerative diseases and cancer. The present review will discuss the current knowledge surrounding the clinical potential and current development of histone deacetylases for the treatment of diseases for which a pro-inflammatory environment plays important roles, and the molecular mechanisms by which such inhibitors may play important functions in modulating the pro-inflammatory environment.     

Kaempferol,a new nutrition-derived pan-inhibitor of human histone deacetylases

Kaempferol is a natural polyphenol belonging to the group of flavonoids. Different biological functions like inhibition of oxidative stress in plants or animal cells and apoptosis induction have been directly associated with kaempferol. The underlying mechanisms are only partially understood. Here we report for the first time that kaempferol has a distinct epigenetic activity by inhibition of histone deacetylases (HDACs). In silico docking analysis revealed that it fits into the binding pocket of HDAC2, 4, 7 or 8 and thereby binds to the zinc ion of the catalytic center. Further in vitro profiling of all conserved human HDACs of class I, II and IV showed that kaempferol inhibited all tested HDACs. In clinical oncology, HDAC inhibitors are currently under investigation as new anticancer compounds. Therefore, we studied the effect of kaempferol on human-derived hepatoma cell lines HepG2 and Hep3B as well as on HCT-116 colon cancer cells and found that it induces hyperacetylation of histone complex H3. Furthermore, kaempferol mediated a prominent reduction of cell viability and proliferation rate. Interestingly, toxicity assays revealed signs of relevant cellular toxicity in primary human hepatocytes only starting at 50 μM as well as in an in vivo chicken embryotoxicity assay at 200 μM.In conclusion, the identification of a novel broad inhibitory capacity of the natural compound kaempferol for human-derived HDAC enzymes opens up the perspective for clinical application in both tumor prevention and therapy. Moreover, kaempferol may serve as a novel lead structure for chemical optimization of pharmacokinetics, pharmacology or inhibitory activities.     

Novel inhibitors of human histone deacetylases: Design,synthesis and bioactivity of 3-alkenoylcoumarines

Histone deacetylases (HDACs) are well-established, promising targets for anticancer therapy due to their critical role in cancer development. Accordingly, an increasing number of HDAC inhibitors displaying cytotoxic effects against cancer cells have been reported. Among them, a large panel of chemical structures was described including coumarin-containing molecules. In this study, we described synthesis and biological activity of new coumarin-based derivatives as HDAC inhibitors. Among eight derivatives, three compounds showed HDAC inhibitory activities and antitumor activities against leukemia cell lines without affecting the viability of peripheral blood mononuclear cells from healthy donors.