Abstract 632: MTOR-Proteasome Imbalance Upon Deletion of Pras40 Inhibits Cardiac Growth but Results in Cardiac Failure

Abstract

Background: The mammalian target of Rapamycin complex 1 (mTORC1) increases cell size by initiating translation as well as by inhibiting catabolic functions such as proteolysis or autophagy. We have previously proposed Proline-rich Akt substrate 1 (Pras40) as a cardioprotective endogenous inhibitor of mTOR-dependent protein synthesis during pathologic growth. Pras40 is released from mTORC1 during growth, but other interactions are largely unknown. In a proteomic screen we have now found a novel interaction of Pras40 with the 26S proteasome during hypertrophy. We hypothesize that Pras40 directly links the mTOR-dependent protein synthesis network and the proteasomal degradation machinery to balance both processes. Methods and Results: To determine the growth-dependent Pras40-interactome, we identified binding partners by proximity-dependent biotin labeling using APEX2 and subsequent mass-spec. Interestingly, aside from interactions with the mTOR-dependent translation machinery, we find Pras40 binding to subunits of the 26S proteasome only during growth. To test consequences of Pras40 deletion on cardiac function in vivo , we generated cardiomycyte-specific knock-out mice, that we subjected to pathologic hypertrophy (TAC). Conversely to Pras40 overexpression, we find growth significantly blunted in KO animals, function reduced and fibrosis elevated. mTORC1 signaling as well as autophagy and proteasomal function are severely disturbed in KO animals. Mechanistically, chymotrypsin-like 26S proteasome activity is blunted in KO hearts as well as isolated cardiomyocytes from KO animals, whereas overexpression shows reciprocal effects in myocytes. Disturbed proteasomal function in KO mice leads to severe alterations in metabolic functions highlighting the importance of both intact mTORC1 signaling and proper proteasomal maintenance during cardiac stress. Conclusion: In this study we find a novel mechanism how mTOR and proteasomal function are linked in the diseased heart. We provide evidence that Pras40 links anabolic protein synthesis and catabolic proteolysis in the heart: At rest, Pras40 binds and inhibits mTOR, but when released during pathologic growth, Pras40 directly interacts with the 26S proteasome and modulates its activity.