Histone deacetylase inhibition reduces cardiac connexin43 expression and gap junction communication

Qin Xu,Richard D Veenstra,Xianming Lin,Laura Andrews,Paul D Lampe,Dakshesh Patel

doi:10.3389/fphar.2013.00044

Qin Xu, Richard D Veenstra + Show 4 more

Open Access

https://doi.org/10.3389/fphar.2013.00044

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Abstract

Histone deacetylase inhibitors (HDACIs) are being investigated as novel therapies for cancer, inflammation, neurodegeneration, and heart failure. The effects of HDACIs on the functional expression of cardiac gap junctions (GJs) are essentially unknown. The purpose of this study was to determine the effects of trichostatin A (TSA) and vorinostat (VOR) on functional GJ expression in ventricular cardiomyocytes. The effects of HDAC inhibition on connexin43 (Cx43) expression and functional GJ assembly were examined in primary cultured neonatal mouse ventricular myocytes. TSA and VOR reduced Cx43 mRNA, protein expression, and immunolocalized Cx43 GJ plaque area within ventricular myocyte monolayer cultures in a dose-dependent manner. Chromatin immunoprecipitation experiments revealed altered protein interactions with the Cx43 promoter. VOR also altered the phosphorylation state of several key regulatory Cx43 phospho-serine sites. Patch clamp analysis revealed reduced electrical coupling between isolated ventricular myocyte pairs, altered transjunctional voltage-dependent inactivation kinetics, and steady state junctional conductance inactivation and recovery relationships. Single GJ channel conductance was reduced to 54 pS only by maximum inhibitory doses of TSA (≥ 100 nM). These two hydroxamate pan-HDACIs exert multiple levels of regulation on ventricular GJ communication by altering Cx43 expression, GJ area, post-translational modifications (e.g., phosphorylation, acetylation), gating, and channel conductance. Although a 50% downregulation of Cx43 GJ communication alone may not be sufficient to slow ventricular conduction or induce arrhythmias, the development of class-selective HDACIs may help avoid the potential negative cardiovascular effects of pan-HDACI.

Highlights

Histone acetyltransferases (HATs) and deacetylases (HDACs) regulate the nuclear protein acetylation–deacetylation cycle that modulates gene expression by altering chromatin condensation and transcription factor [e.g., MEF2, p53] activities (Kouzarides, 2000; Yang and Grégoire, 2007; Yang and Seto, 2008; Haberland et al, 2009)
HDAC EXPRESSION AND INHIBITION IN CULTURED VENTRICULAR MYOCYTES The relative expression of all 11 HDACs in neonatal mouse ventricular myocyte cultures was examined by RT-PCR
The www.frontiersin.org same relative HDAC mRNA expression pattern was found in atrial cardiomyocyte cultures wherein the Cx40 mRNA level was 38.4 ± 5.9%, consistent with the contribution of Cx40 to atrial myocardial gap junctions (GJs) (Figure 1A; Lin et al, 2010)

Summary

Introduction

Histone acetyltransferases (HATs) and deacetylases (HDACs) regulate the nuclear protein acetylation–deacetylation cycle that modulates gene expression by altering chromatin condensation and transcription factor [e.g., MEF2 (myocyte enhancer factor-2), p53] activities (Kouzarides, 2000; Yang and Grégoire, 2007; Yang and Seto, 2008; Haberland et al, 2009). The class IIa HDACs (HDAC4, 5, 7, and 9) possess amino-terminal MEF2 and 14-3-3 protein binding motifs and shuttle between the nucleus and cytosol in a phosphorylationdependent manner (McKinsey et al, 2000; Zhou et al, 2000; Little et al, 2007). The function of the class IIb HDAC10 is poorly understood

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