RBCK1, a protein kinase C βI binding partner and its splice variant RBCK2: potential role in cardiac hypertrophy?

Abstract

The protein kinase C (PKC) family participates in signal transduction in a large number of mammalian cell types. Several PKC isozymes are expressed in rat cardiac myocytes. Many studies have shown that different PKC isozymes are involved in the modulation of diverse functions in the heart, including the developmental and pathological hypertrophic growth of cardiomyocytes.We are focused on understanding the individual role of each PKC isozyme and how they regulate cardiac functions. Our laboratory and others have demonstrated that activation of PKC beta plays a critical role in the development of cardiac hypertrophy. Recently, the PKC-binding partner designated RBCK1 for RBCC protein interacting with PKC 1, has been shown to interact specifically with PKCβI when coexpressed in cultured COS-7 cells. In addition, two different studies have established a correlation between the variation of RBCK1 mRNA levels and cardiac hypertrophy. However, whether the product protein mediate hypertrophy or whether the changes in its levels are the result of hypertrophy cannot be determined from such an analysis. Based on these results, the goal of our present study is therefore to determine if RBCK1 participates actively in the processes leading to cardiac hypertrophy and characterize the molecular mechanisms involved. Does RBCK1 bind PKCβI in cardiac myocytes and is their interaction altered during the induction of hypertrophy? Is the level of expression of PKCβI regulated by the level of expression of RBCK1? A splice variant of RBCK1 named RBCK2 has been cloned and characterized. Does RBCK2 also participate in the development of cardiac hypertrophy? To investigate these questions, adenoviralmediated gene transfer of wild-type RBCK1 and RBCK2 fused to the green fluorescent protein (GFP) was carried out in cultured neonatal rat ventricular cardiac myocytes. By identifying regulators and binding proteins of PKC, we may identify new means to modulate the progression of pathological hypertrophy of the heart. In this study, we showed the preferential interaction of RBCK1 with inactive PKCβI in vitro and the perinuclear localization of endogenous PKCβI and overexpressed RBCK1 in unstimulated cells. This result suggests that RBCK1 could be a binding partner for inactive PKCβI in rat neonatal cardiac myocytes. We also found an increase in the level of expression of endogenous PKCβI and RBCK1 proportional to the time in culture and the induction of hypertrophy with phenylephrine (PE). The level of expression of PKCβI is directly correlated to the level of expression of RBCK1. An isoform specific motility shift of PKCβI was also seen upon the overexpression of RBCK1-GFP, which could reflect a dephosphorylation state. Finally, there was an Nesiritide Improves Urine Output in Severely Ill Pediatric Patients Awaiting Heart Transplantation Without Severe Hypotension Ruchir Sehra, Karen R. Underwood2—Pediatrics/Pediatric Cardiology, Loma Linda University, Loma Linda, CA; Critical Care Medicine, Chlidren’s Hospital of Los Angeles, Los Angeles, CA