Abstract 3606: FHL2 Binds Calcineurin and Represses Pathologic Cardiac Growth
Introduction: FHL2 (four and a half LIM domain protein) is a protein of unknown function highly enriched in the myocardium. Whereas FHL2 knockout (FHL2−/−) mice are healthy, cardiac hypertrophy induced by chronic stimulation of beta-adrenergic receptors is amplified in FHL2−/−mice. The calcineurin-NFAT pathway is activated during isoproterenol-induced hypertrophy, and inhibition of calcineurin (CnA) is sufficient to block the hypertrophic response. We hypothesized FHL2 acts as brake on the calcineurin-NFAT pathway.
Methods and Results: To determine whether FHL2 regulates calcineurin signaling, wild type (WT) and FHL2−/− mice were treated with the beta-adrenergic agonist isoproterenol, and the expression of NFAT target genes was analyzed. Basal mRNA levels of RCAN1.4 and BNP were similar in control WT and FHL2−/− mice, but isoproterenol-induced expression of RCAN1.4 and BNP were each significantly elevated in FHL2−/− compared to WT. To analyze the direct effect of FHL2 on calcineurin-NFAT signaling, C2C12 myoblasts were transfected with a luciferase reporter construct driven by the promoters of either RCAN1 or IL-2 (NFAT target genes). Over-expression of FHL2 inhibited the activation of these promoters by CnA. Conversely, knockdown of FHL2 led to increased activation of these promoters by CnA. Similar results were observed in vivo, where a transgenic mouse carrying a cardiac-specific constitutively active calcineurin-A alpha transgene manifested higher levels of BNP in the FHL2−/− background as compared to WT background. Finally, an interaction between FHL2 and CnA was demonstrated by co-immunoprecipitation, and immunostaining revealed co-localization of these proteins predominantly to the sarcomere in adult cardiomyocytes.
Conclusions: In vitro and in vivo analyses demonstrate inhibition of the CnA pathway by FHL2 and suggest that FHL2 suppresses hypertrophic cardiac growth, at least in part, by inhibiting CnA signaling
This research has received full or partial funding support from the American Heart Association, South Central Affiliate (Arkansas, New Mexico, Oklahoma & Texas).