Important recent advances derived from experimental models of cardiac hypertrophy suggest that distinct signaling pathways regulate compensated hypertrophy and decompensated heart failure. Ras is a small GTP binding protein that signals through multiple effectors, including the Raf-MEK-ERK pathway, PI3 kinase (PI3K), guanine exchange factors for Ral (RalGEFs) and cJun N-terminal kinase (JNK). In transgenic mice, V12Ras-mediated hypertrophy is associated with JNK activation and depressed cardiac function. However, certain Ras-dependent signaling cascades, including ERK and PI3K, activate compensatory hypertrophic responses that preserve contractile activity. To identify novel Ras effectors we screened a human heart cDNA library using Ras as bait and isolated Ral guanine nucleotide exchange factor-like factor (RIf). To examine the functional consequence of the Ras-RIf interaction, we expressed V12Ras in concert with RIf in neonatal ventricular rat myocytes (NRVM). Expression of RIf suppressed V12Ras-induced JNK activity and ANF expression but not other Ras effectors. RIf also inhibited sorbitol-induced JNK activation indicating that RIf expression negatively regulates JNK activation in response to multiple stimuli. These results demonstrate that RIf suppresses an effector pathway suggested to mediate depressed function of myocardial cells. We will test the hypothesis that RIf negatively regulates JNK activation and that function inhibits the development of hypertrophic cardiomyopathy in vivo. The signaling pathways that couple Ras to JNK activation are not known but may involve MEKK1 (MAPWERK kinase kinase 1), a Ras binding protein that preferentially activates JNK.
In aim #1, we will define the role of MEKK1-JKK-JNK in RIf-mediated inhibition of V12Ras- and stress- signaling.
In aims #2 and #3, the pathophysiological implications of RIf function will be investigated in transgenic mice that overexpress RIf. We will test the potential protective role of RIf on the pathogenesis of cardiac hypertrophy in response to pressure overload, isoproterenol infusion and V12Ras overexpression and investigate the signaling cascades involved. Elucidation of the mechanisms by which RIf regulates hypertrophic signaling pathways may lead the development of novel therapeutic approaches to prevent cardiac dysfunction.
