The heart undergoes hypertrophic enlargement in response to both physiologic and pathophysiologic stimulation. Hypertrophy allows the myocardium to adapt to routine alterations in workload associated with developmental maturation, physiologic challenge, or injury. A finite number of intracellular signaling pathways have been implicated as regulators of the cardiac hypertrophic response including, mitogen-activated protein kinase (MAPK), protein kinase C (PKC), PI3K/Akt, and calcineurin-NFAT. Our previous studies have suggested a critical role for the calcium activated phosphatase calcineurin (PP2B) and its downstream transcriptional effector NFAT (nuclear factor of activated T-cells) in regulating the cardiac hypertrophic response. Calcineurin is an attractive candidate molecule for investigation as a hypertrophic transducer given its unique ability to dynamically respond to alterations in intracellular calcium handling. Indeed, data generated from both gain and loss of function approaches in genetically modified mouse models support the hypothesis that calcineurin functions as an important transducer of the cardiac hypertrophic response. Here we propose to investigate the downstream effectors of calcineurin in the heart and to investigate the role that calcineurin plays in regulating physiologic and/or adaptive growth responses of the myocardium. To accompolish this objective, the following specific aims and approaches are proposed: 1) To define calcineurin's role as a regulator of cardiac mass in response to physiologic stimuli. Specifically, CnBl-loxP targeted mice will undergo inducible Cre-mediated recombination in the heart to examine the hypothesis that calcineurin regulates the steady-state mass of the myocardium in response to physiologic stimuli. NFAT-dependent reporter mice will also be investigated to assess the hypothesis that calcineurin is a long-term regulator of hypertrophic growth. 2) To define the downstream effectors of calcineurin that mediate the hypertrophic growth of the myocardium. Both dominant negative NFAT and MEF2 inducible transgenic mice will be characterized to evaluate the hypothesis that these factors are necessary calcineurin transducers in the heart. 3) To characterize a novel class of NFAT interacting proteins (Nip-1 and Nip-2) in the heart. Yeast 2-hybrid screening has identified a novel class of cardiac-expressed NFAT inhibiting factors that will be further evaluated in vitro and in vivo.
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