Abstract 3989: Novel Role of CHIP E3 Ubiquitin Ligase, C-terminus of Hsp70-interacting Protein, on Myocyte Apoptosis via ERK5-mediated Destabilization of Inducible Camp Early Repressor (ICER)
Apoptosis plays a critical role in cardiac dysfunction after myocardial infarction (MI), and emerging evidence suggested the critical role of ICER in this process. Previously, we found that PKA activity stabilized ICER at protein level. In contrast, insulin growth factor-1 (IGF-1)-mediated activation of ERK5 de-stabilized ICER and inhibited apoptosis. In this study, we investigated the mechanism by which ERK5 activation can regulate ICER stability. The de-stabilization of ICER induced by ERK5 activation was inhibited by MG132, proeasome inhibitor, suggesting the involvement of ubiquitin proteasome system. After extensive survey, we found that IGF-1 increased CHIP association with ERK5 as a candidate ubiquitination (Ub) enzyme downstream of ERK5 in cardiomyocytes. In vitro run-off assay showed that CHIP de-stabilized ICER, and enzyme dead form of CHIP and MG132 inhibited this de-stabilization, indicating that ICER is a novel substrate of CHIP. IGF-1 and adenovirus containing constitutive active form of MEK5 (Ad-CA-MEK5α; specific ERK5 activator) increased CHIP Ub activity using GST-ICER as a substrate, and depletion of ERK5 by siRNA inhibited this CHIP activation, supporting the key role of ERK5 on CHIP Ub activation. IGF-1 and Ad-CA-MEK5α decreased isoproterenol-mediated ICER induction and myocyte apoptosis (% of TUNEL positive/total cells, vehicle vs IGF-1: 8.2±0.8 vs 3.5±1.1 and Ad-LacZ vs Ad-CA-MEK5α: 8.8±1.2 vs 3.2±1.4, p<0.01), but the depletion of CHIP using siRNA abolished this inhibition (IGF-1 + si-CHIP: 7.4±0.2, Ad-CA-MEK5α + si-CHIP: 7.7±0.8, p<0.05). These data strongly suggest the functional consequence of ERK5-CHIP Ub activation on ICER de-stabilization and subsequent myocyte apoptosis. Previously, we reported the protective role of CA-MEK5α on cardiac dysfunction and apoptosis after MI via inhibiting ICER induction using cardiac specific CA-MEK5α transgenic (Tg) mice. We found that CHIP Ub activity significantly was increased in CA-MEK5α-Tg mice (fold increase vs wild type mice: 2.5±0.7, p<0.05), also suggesting the critical role of ERK5-CHIP Ub activation on ICER expression and subsequent apoptosis in vivo. The novel cardio-protective mechanism by ERK5-CHIP will provide a new therapeutic strategy for heart failure.
This research has received full or partial funding support from the American Heart Association, National Center.