Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, PC353

Poster Communications

Arterial acetylation of histones is increased by a broad class I/II lysine deacetylase (KDAC) inhibitor but not by a specific KDAC8 antagonist.

A. Chen1, M. Karolczak-Bayatti1, M. Sweeney1, S. Ulrich2, N. G. Europe-Finner1, M. J. Taggart1

1. Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom. 2. Department of Chemistry, Ithaca College, Ithaca, New York, United States.


Acetylation is increasingly recognised as a post-translational modification with a wide array of potential protein targets to rival that of phosphorylation. Originally associated with the modification of nuclear histones it is becoming apparent that many non-nuclear proteins are targets for specific lysine residue acetylation by lysine acetyltransferases (KATs formerly known as HATs) and lysine deacetylases (KDACs formerly known as HDACs) (1,2). Indeed, class I KDACs (1, 2, 3, 8 and 11) have been suggested to be localised in the nucleus yet in cultured vascular smooth muscle cells KDAC8 was found to bind to alpha smooth muscle actin and determine cell motility (3). The aim of this study, therefore, was to investigate in intact arterial smooth muscle the effect of a classI/II KDAC inhibitor, trichostatin A (TSA), in comparison to that of a specific KDAC8 inhibitor (Compound 2), on acetylation of nuclear histones. Aorta tissue isolated from Wistar rats, or rat A7r5 cells to enable a comparison between differentiated smooth muscle tissue and cultured cells, were incubated in DMEM with either TSA (3.3 µM in ethanol), Compound 2 (200 µM in DMSO) or appropriate control for 0, 1, 3 or 24 hours. Protein homogenates were analysed by western blotting for histone acetylation using an anti-acetylated H3 antibody. Densitometry data were analysed with one-way ANOVA and Bonferroni post-hoc tests, values are means ± sem. TSA induced significant (P<0.05) elevation of histone acetylation at all timepoints in both aorta tissue and cells. For example, aorta tissue: control 0 hr, 3.68±2.50 denitometric a.u., TSA 13.90±2.52 a.u., 20.6±2.52 a.u. and 27.3±2.42 a.u. at 1, 3 and 24 hours respectively (n=3). In contrast, treatment with Compound 2 was without effect on histone acetylation even though it was efficacious in inhibiting KDAC8 activity in an in vitro fluorimetric assay. Vehicle controls had no effect. Immunofluorescent staining of aorta tissue clarified a predominantly nuclear localisation of KDAC1 yet an almost entirely non-nuclear expression pattern of KDAC8. These data support the notion that KDAC8, in location and function, is distinct from other class I KDACs and is therefore unlikely to directly influence gene expression by chromatin modification. The non-nuclear protein target(s) of KDAC8 in arterial smooth muscle, and the functional consequences of such interactions, remain to be determined.

Where applicable, experiments conform with Society ethical requirements