Depletion of Tip60 from In Vivo Cardiomyocytes Increases Myocyte Density, Followed by Cardiac Dysfunction, Myocyte Fallout and Lethality.

Joseph B Fisher,Min-Su Kim,John Auchampach,John Lough,Audrey Horst,Tina Wan,Xiaolei Xu

doi:10.1371/journal.pone.0164855

Abstract

Tat-interactive protein 60 (Tip60), encoded by the Kat5 gene, is a member of the MYST family of acetyltransferases. Cancer biology studies have shown that Tip60 induces the DNA damage response, apoptosis, and cell-cycle inhibition. Although Tip60 is expressed in the myocardium, its role in cardiomyocytes (CMs) is unclear. Earlier studies here showed that application of cardiac stress to globally targeted Kat5+/—haploinsufficient mice resulted in inhibition of apoptosis and activation of the CM cell-cycle, despite only modest reduction of Tip60 protein levels. It was therefore of interest to ascertain the effects of specifically and substantially depleting Tip60 from CMs using Kat5LoxP/-;Myh6-Cre mice in the absence of stress. We report initial findings using this model, in which the effects of specifically depleting Tip60 protein from ventricular CMs, beginning at early neonatal stages, were assessed in 2–12 week-old mice. Although 5’-bromodeoxyuridine immunostaining indicated that CM proliferation was not altered at any of these stages, CM density was increased in 2 week-old ventricles, which persisted in 4 week-old hearts when TUNEL staining revealed inhibition of apoptosis. By week 4, levels of connexin-43 were depleted, and its patterning was dysmorphic, concomitant with an increase in cardiac hypertrophy marker expression and interstitial fibrosis. This was followed by systolic dysfunction at 8 weeks, after which extensive apoptosis and CM fallout occurred, followed by lethality as mice approached 12 weeks of age. In summary, chronic depletion of Tip60 from the ventricular myocardium beginning at early stages of neonatal heart development causes CM death after 8 weeks; hence, Tip60 protein has a crucial function in the heart.

Highlights

Tip60 (Tat-interactive protein 60 kD) is an acetylase protein, which in cancer cells has been shown to induce the DNA Damage Response (DDR) and apoptosis by respectively acetylating ATM [1,2] and p53 [3,4]
The initial objective of this study was to compare the effects of depleting Tip60 from CMs in the left ventricle of experimental mice having the Kat5LoxP/-;Myh6-Cre genotype, with control mice having the Kat5LoxP/+;Myh6-Cre genotype, at 4, 8 and 12 weeks of age
Depletion, experimental mice were bred to contain a null Kat5 allele instead of a second LoxPflanked allele, since only one Kat5 allele had to be recombined, and because our previous work had revealed that Kat5+/- heterozygous mice express Tip60 protein at near wild-type levels [9]

Summary

Introduction

Tip (Tat-interactive protein 60 kD) is an acetylase protein, which in cancer cells has been shown to induce the DNA Damage Response (DDR) and apoptosis by respectively acetylating ATM [1,2] and p53 [3,4]. Tip has been shown to inhibit the cell-cycle [5]. Global heterozygous Kat5+/- mice reproduce normally, exhibiting no apparent haploinsufficient phenotype under normal conditions [6]. This laboratory wishes to elucidate Tip60’s role in the adult heart, wherein it is relatively enriched [7]. In accord with Tip60’s role in cancer cells, both stressors reduced apoptosis while inducing re-entry of ventricular cardiomyocytes (CMs) into the cell-cycle, despite the retention of Tip protein at ~80% of wildtype levels [9]

Objectives

Methods

Results

Conclusion