Changes in cardiac Nav1.5 expression, function, and acetylation by pan-histone deacetylase  inhibitors

Histone deacetylase (HDAC) inhibitors are small molecule anticancer therapeutics that exhibit limiting cardiotoxicities including QT interval prolongation and life-threatening cardiac arrhythmias. Because the molecular mechanisms for HDAC inhibitor-induced cardiotoxicity are poorly understood, we performed whole cell patch voltage-clamp experiments to measure cardiac sodium currents ( INa) from wild-type neonatal mouse ventricular or human-induced pluripotent stem cell-derived cardiomyocytes treated with trichostatin A (TSA), vorinostat (VOR), or romidepsin (FK228). All three pan-HDAC inhibitors dose dependently decreased peak INa density and shifted the voltage activation curve 3- to 8-mV positive. Increases in late INa were not observed despite a moderate slowing of the inactivation rate at low activating potentials (<−40 mV). Scn5a mRNA levels were not significantly altered but NaV1.5 protein levels were significantly reduced. Immunoprecipitation with anti-NaV1.5 and Western blotting with anti-acetyl-lysine antibodies indicated that NaV1.5 acetylation is increased in vivo after HDAC inhibition. FK228 inhibited total cardiac HDAC activity with two apparent IC50s of 5 nM and 1.75 μM, consistent with previous findings with TSA and VOR. FK228 also decreased ventricular gap junction conductance ( gj), again consistent with previous findings. We conclude that pan-HDAC inhibition reduces cardiac INa density and NaV1.5 protein levels without affecting late INa amplitude and, thus, probably does not contribute to the reported QT interval prolongation and arrhythmias associated with pan-HDAC inhibitor therapies. Conversely, reductions in gj may enhance the occurrence of triggered activity by limiting electrotonic inhibition and, combined with reduced INa, slow myocardial conduction and increase vulnerability to reentrant arrhythmias.

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Regulation of intestinal serotonin transporter expression via epigenetic mechanisms: role of HDAC2

AJP Cell Physiology ◽

10.1152/ajpcell.00361.2012 ◽

2013 ◽

Vol 304 (4) ◽

pp. C334-C341 ◽

Cited By ~ 21

Author(s):

Ravinder K. Gill ◽

Anoop Kumar ◽

Pooja Malhotra ◽

Daniel Maher ◽

Varsha Singh ◽

...

Keyword(s):

Hdac Inhibitors ◽

In Vivo Model ◽

Mrna Levels ◽

Epigenetic Mechanisms ◽

Hdac Inhibition ◽

Exon 2 ◽

Protein Levels ◽

Histone Hyperacetylation

The serotonin (5-HT) transporter (SERT) facilitates clearance of extracellular 5-HT by its uptake and internalization. Decreased expression of SERT and consequent high 5-HT levels have been implicated in various diarrheal disorders. Thus, appropriate regulation of SERT is critical for maintenance of 5-HT homeostasis in health and disease. Previous studies demonstrated that SERT is regulated via posttranslational and transcriptional mechanisms. However, the role of epigenetic mechanisms in SERT regulation is not known. Current studies investigated the effects of histone deacetylase (HDAC) inhibition on SERT expression and delineated the mechanisms. Treatment of Caco-2 cells with the pan-HDAC inhibitors butyrate (5 mM) and trichostatin (TSA, 1 μM) decreased SERT mRNA and protein levels. Butyrate- or TSA-induced decrease in SERT was associated with decreased activity of human SERT (hSERT) promoter 1 (upstream of exon 1a), but not hSERT promoter 2 (upstream of exon 2). Butyrate + TSA did not show an additive effect on SERT expression, indicating that mechanisms involving histone hyperacetylation may be involved. Chromatin immunoprecipitation assays demonstrated enrichment of the hSERT promoter 1 (flanking nt −250/+2) with tetra-acetylated histone H3 or H4, which was increased (∼3-fold) by butyrate. Interestingly, specific inhibition of HDAC2 (but not HDAC1) utilizing small interfering RNA decreased SERT mRNA and protein levels. The decrease in SERT expression by HDAC inhibition was recapitulated in an in vivo model . SERT mRNA levels were decreased in the ileum and colon of mice fed pectin (increased availability of butyrate) compared with controls fed a fiber-free diet (∼50–60%). Our results identify a novel role of HDAC2 as a regulator of SERT gene expression in intestinal epithelial cells.

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Acetylation of PML Plays a Key Role in Histone Deacetylase Inhibitor-Mediated Apoptosis through Enhanced PML Sumoylation.

Blood ◽

10.1182/blood.v110.11.4164.4164 ◽

2007 ◽

Vol 110 (11) ◽

pp. 4164-4164

Author(s):

Fumihiko Hayakawa ◽

Issay Kitabayashi ◽

Pier P. Pandolfi ◽

Tomoki Naoe

Keyword(s):

Cancer Cells ◽

Histone Deacetylase ◽

Anticancer Agents ◽

Posttranslational Modification ◽

Hdac Inhibitor ◽

Trichostatin A ◽

Hdac Inhibitors ◽

Antibody Array

Abstract The promyelocytic leukemia (PML) protein is a potent tumor suppressor and proapoptotic factor, and is functionally regulated by posttranslational modification such as phosphorylation, sumoylation, and ubiquitination. Histone deacetylase (HDAC) inhibitors are a promising class of targeted anticancer agents and induce apoptosis to cancer cells. In addition to their effects on histones, HDAC inhibitors increase the acetylation level of several non-histone proteins such as transcription factors, which are important for their effects to cancer cells. However, the mechanism of HDAC inhibitor-induced apoptocis is largely unknown. We report here a novel posttranscriptional modification, acetylation, of PML. By the screening using antibody array, we identified PML as a new acetylation target of Trichostatin A (TSA), a HDAC inhibitor. PML acetylation was enhanced by coexpression of p300 or treatment with TSA. We also showed that increased PML acetylation was associated with increased sumoylation of PML in vitro and in vivo. PML involvement in TSA-induced apoptosis was demonstrated by PML knocking down in Hela cells or PML overexpression in PML−/− MEF cells. Furthermore, PML with acetylation-defective mutation showed disability to mediate the apoptosis, suggesting the importance of PML acetylation for it. Our work provides new insights into the PML regulation by posttranslational modification, and new information about the therapeutic mechanism of HDAC inhibitors.

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Activation of the Growth-Differentiation Factor 11 Gene by the Histone Deacetylase (HDAC) Inhibitor Trichostatin A and Repression by HDAC3

Molecular and Cellular Biology ◽

10.1128/mcb.24.12.5106-5118.2004 ◽

2004 ◽

Vol 24 (12) ◽

pp. 5106-5118 ◽

Cited By ~ 64

Author(s):

Xiaohong Zhang ◽

Walker Wharton ◽

Zhigang Yuan ◽

Shih-Chang Tsai ◽

Nancy Olashaw ◽

...

Keyword(s):

Histone Deacetylase ◽

Anticancer Agents ◽

Hdac Inhibitor ◽

Transforming Growth Factor ◽

Trichostatin A ◽

Expression Profiles ◽

Hdac Inhibitors ◽

Gene Expression Profiles ◽

Differentiation Factor

ABSTRACT Histone deacetylase (HDAC) inhibitors inhibit the proliferation of transformed cells in vitro, restrain tumor growth in animals, and are currently being actively exploited as potential anticancer agents. To identify gene targets of the HDAC inhibitor trichostatin A (TSA), we compared the gene expression profiles of BALB/c-3T3 cells treated with or without TSA. Our results show that TSA up-regulates the expression of the gene encoding growth-differentiation factor 11 (Gdf11), a transforming growth factor β family member that inhibits cell proliferation. Detailed analyses indicated that TSA activates the gdf11 promoter through a conserved CCAAT box element. A comprehensive survey of human HDACs revealed that HDAC3 is necessary and sufficient for the repression of gdf11 promoter activity. Chromatin immunoprecipitation assays showed that treatment of cells with TSA or silencing of HDAC3 expression by small interfering RNA causes the hyperacetylation of Lys-9 in histone H3 on the gdf11 promoter. Together, our results provide a new model in which HDAC inhibitors reverse abnormal cell growth by inactivation of HDAC3, which in turn leads to the derepression of gdf11 expression.

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