In the early developing embryo Gata4
were shown to be of critical importance for the establishment of the entire cardiac gene program, as loss of both factors, but not either alone, results in acardia.27
This result demonstrates the importance of GATA transcription factors as necessary regulators of cardiac differentiation specific gene expression, hence it was not entirely unexpected when GATA-4 was shown to be necessary for cardiac hypertrophy in the adult heart, as this process requires re-establishment of the fetal gene program. While almost nothing is known about the role of GATA-6 in regulating hypertrophy or differentiation specific gene expression in the adult heart, GATA-4 and GATA-6 are each capable of inducing hypertrophy when overexpressed in neonatal rat cardiomyocytes, suggesting for the first time that GATA-6 might function similar to GATA-4 in this respect.13, 28
Indeed, GATA-4 can functionally and physically interact with GATA-6 in activation of the ANF and BNP promoters.28
While GATA-4 and GATA-6 have been shown to positively regulate BNP and MYH promoters in cultured cardiomyocytes13, 28
, we actually observed upregulation of BNP and βMHC expression in the absence of GATA-6. It is likely that gene regulation in vivo is distinctly different from conditions that occur with transfection of minimal promoters and co-overexpression of GATA-4 and/or GATA-6 in cultured cells. Also, these minimal promoter constructs that were transfected into neonatal myocytes could also easily lack inhibitory GATA binding sites or other negative regulatory sites that can bind secondary transcription factors induced by GATA-4/6.
Here we showed for the first time, that in addition to GATA-4, GATA-6 protein expression is dramatically enhanced in mouse hearts subjected to pressure overload stimulation, which correlated with a dramatic increase in total GATA DNA binding activity. It was intriguing that deletion of Gata6 from the neonatal mouse heart, but not Gata4, severely reduced total GATA DNA binding activity assayed with a canonical GATA binding site from the αMHC promoter. This suggests that GATA-6 protein levels may be higher than GATA-4 in the postnatal heart, although the overall transcriptional potency of both proteins may differ such that GATA-4 may still be just as critical. In addition, we found a significant reduction in GATA-4 protein in the Gata6-deleted hearts, which could contribute to the reduced DNA binding activity. Intriguingly, this result hints at a positive regulatory relationship between GATA-6 and GATA-4 in the adult heart.
We modeled the observed increase in GATA-6 protein levels after pressure overload stimulation using a transgenic approach. The low and medium expressing line approximated this increase in endogenous GATA-6 with hypertrophy, suggesting that our transgenic approach closely model this known increase. Finally, and perhaps more importantly, deletion of Gata6 from the mouse heart significantly reduced the hypertrophic growth response, both after TAC and after stimulation with AngII/PE, together suggesting that GATA-6 functions as a necessary and sufficient mediator of adult cardiac hypertrophic growth.
That GATA-6 appears to be equally important to GATA-4 in regulating the cardiac hypertrophic response might suggest a degree of functional redundancy between these 2 transcription factors. Indeed, we previously demonstrated that Gata4+/− Gata6+/−
(double heterozygotes) are not viable and perish during mid-gestation, while single heterozygotes for either gene were viable.9
In the adult heart, the phenotype of reduced cardiac hypertrophy after 2 weeks of TAC was nearly identical between Gata4
heart-deleted mice, and each succumbed to heart failure and reductions in cardiac function to a roughly similar extent. Moreover, GATA-4 and GATA-6 overexpressing transgenic mice each developed similar levels of spontaneous hypertrophy at about the same time in middle adulthood.13
Finally, deletion of both Gata4
specifically from the heart resulted in spontaneous heart failure and death by the age of 16 weeks. While these collective observations suggest functional redundancy between GATA-4 and GATA-6, some differences were noted that might suggest unique functions for each. First, Gata4fl/flβMHC-cre
mice showed spontaneous deterioration of cardiac function with aging, but Gata6fl/flβMHC-cre
mice showed more preserved cardiac function up to one year of age as assessed by echocardiography (although invasive hemodynamics showed a trend towards reduced baseline function with Gata6
deletion). Similarly, we found a slight but significant reduction in heart size at baseline in Gata6
-deleted mice due to decreased cardiomyocyte content in the heart, while we did not observe this effect in Gata4
-deleted mice. However, Pu and colleagues did observe a decrease in myocyte proliferation in the hearts of Gata4
heart-specific hypomorphic mice (their targeted allele had partial expression), suggesting that there could still be functional similarity between GATA-4 and GATA-6 with respect to control of cell number in the heart.29
Thus, our working hypothesis is that GATA-4 and GATA-6 are roughly equivalent in effect in the heart, such that each of the 4 alleles functions in a dosage-dependent manner. However, it will be important to generate combinations of Gata4
allelic deletions, as well as conduct rescue approaches whereby the GATA-4 transgene is crossed into the Gata6
heart-specific deleted background, and vice versa, before we can unequivocally determine if these 2 factors are completely redundant in function in the heart or if some unique regulatory actions persist between them.
Novelty and significance
What is known?
- GATA-6 can induce hypertrophy in cultured cardiomyocytes
- Upon pressure overload, GATA binding activity increases in the murine heart
- GATA-4 has been shown to be both required and sufficient for cardiac hypertrophy in vivo.
What new information does this article contribute?
- We provide the first description of adult cardiac deletion of GATA-6 showing its requirement for both cardiac hypertrophy in response to various stimuli and for cardiac compensation
- We provide the first proof that cardiac-specific overexpression of GATA-6 is sufficient to induce cardiac hypertrophy in vivo
- We provide the first description of cardiac-specific deletion of both GATA-4 and GATA-6 from the adult heart, which results in spontaneous dilated cardiomyopathy.
This study was designed to evaluate the importance of GATA-6 in vivo for cardiac hypertrophy and compensation. We studied both GATA-6 overexpression and deletion as effectors of pressure overload induced hypertrophy. We also studied the role of GATA-6 deletion in neurohormonal induced hypertrophy. Finally, we also studied GATA-4 deletion as a comparison and both GATA-4 and GATA-6 deletion to begin to address the possible redundancy between these transcription factors. Our results show the requirement of GATA-6 for cardiac hypertrophy and compensation in response to pressure overload and neurohormonal stimulation. In addition, we show the both GATA-4 and GATA-6 are required to maintain normal cardiac function suggesting they may act redundantly.