Novel Roles of GATA4/6 in the Postnatal Heart Identified through Temporally Controlled, Cardiomyocyte-Specific Gene Inactivation by Adeno-Associated Virus Delivery of Cre Recombinase.

Terence W Prendiville,Li Zhong,William T Pu,Da-Zhi Wang,Guangping Gao,Sean M Stevens,Pingzhu Zhou,Aibin He,Zhiqiang Lin,Nathan Vandusen,Haidong Guo,Jinghai Chen

doi:10.1371/journal.pone.0128105

Abstract

GATA4 and GATA6 are central cardiac transcriptional regulators. The postnatal, stage-specific function of the cardiac transcription factors GATA4 and GATA6 have not been evaluated. In part, this is because current Cre-loxP approaches to cardiac gene inactivation require time consuming and costly breeding of Cre-expressing and “floxed” mouse lines, often with limited control of the extent or timing of gene inactivation. We investigated the stage-specific functions of GATA4 and GATA6 in the postnatal heart by using adeno-associated virus serotype 9 to control the timing and extent of gene inactivation by Cre. Systemic delivery of recombinant, adeno-associated virus 9 (AAV9) expressing Cre from the cardiac specific Tnnt2 promoter was well tolerated and selectively and efficiently recombined floxed target genes in cardiomyocytes. AAV9:Tnnt2-Cre efficiently inactivated Gata4 and Gata6. Neonatal Gata4/6 inactivation caused severe, rapidly lethal systolic heart failure. In contrast, Gata4/6 inactivation in adult heart caused only mild systolic dysfunction but severe diastolic dysfunction. Reducing the dose of AAV9:Tnnt2-Cre generated mosaics in which scattered cardiomyocytes lacked Gata4/6. This mosaic knockout revealed that Gata4/6 are required cell autonomously for physiological cardiomyocyte growth. Our results define novel roles of GATA4 and GATA6 in the neonatal and adult heart. Furthermore, our data demonstrate that evaluation of gene function hinges on controlling the timing and extent of gene inactivation. AAV9:Tnnt2-Cre is a powerful tool for controlling these parameters.

Highlights

GATA4 and GATA6 are critical cardiac transcription factors with partial functional redundancy [1,2]
associated virus (AAV) serotype 9 (AAV9) efficiently delivers its cargo to post-mitotic cells including cardiomyocytes [13], and we show that associated virus 9 (AAV9)-mediated Cre delivery is a simple and effective means to control the timing and extent of cardiomyocyte gene inactivation
Acute loss of Gata4 and Gata6 in the perinatal period caused rapid loss of systolic function. This is a critical period in heart development, during which time cardiomyocytes adapt to substantially higher hemodynamic demands by undergoing numerous changes in their size, ultrastructure, sarcomere composition, and metabolism

Summary

Introduction

GATA4 and GATA6 are critical cardiac transcription factors with partial functional redundancy [1,2]. Later fetal inactivation of either GATA4 or GATA6 with Myh6-Cre ( known as MHCα-Cre) permitted embryonic survival [4,5] These mutant mice developed progressive dilated cardiomyopathy with severe systolic dysfunction and died in adulthood. These mice had attenuated cardiomyocyte hypertrophy in response to pressure overload, indicating that GATA4 and GATA6 are required for this pathological growth response. Inactivation of both Gata and Gata with Myh6-Cre was compatible with embryonic survival, and suggested an additive effect of combined inactivation of both GATA4 and GATA6 [2,4]. The extent to which the adult phenotypes reflect developmental roles of GATA4 or GATA6 or their function in the adult heart remain uncertain

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Conclusion