Patrick M McLendon1, Gregory Davis1, James Gulick1, Sonia R Singh1, Na Xu1, Nathan Salomonis1, Jeffery D Molkentin1, Jeffrey Robbins2. 1. From the Division of Molecular Cardiovascular Biology, Heart Institute, Cincinnati Children's Hospital Medical Center, OH (P.M.M., G.D., J.G., S.R.S., N.X., J.D.M., J.R.); Division of Biomedical Informatics, Cincinnati Children's Hospital, OH (N.S.); and UES, Inc, Dayton, OH (P.M.M.). 2. From the Division of Molecular Cardiovascular Biology, Heart Institute, Cincinnati Children's Hospital Medical Center, OH (P.M.M., G.D., J.G., S.R.S., N.X., J.D.M., J.R.); Division of Biomedical Informatics, Cincinnati Children's Hospital, OH (N.S.); and UES, Inc, Dayton, OH (P.M.M.). Jeff.Robbins@cchmc.org pmclendon@ues.com.
Abstract
RATIONALE: Postmitotic cells, such as cardiomyocytes, seem to be particularly susceptible to proteotoxic stimuli, and large, proteinaceous deposits are characteristic of the desmin-related cardiomyopathies and crystallin cardiomyopathic diseases. Increased activity of protein clearance pathways in the cardiomyocyte, such as proteasomal degradation and autophagy, has proven to be beneficial in maintaining cellular and cardiac function in the face of multiple proteotoxic insults, holding open the possibility of targeting these processes for the development of effective therapeutics. OBJECTIVE: Here, we undertake an unbiased, total genome screen for RNA transcripts and their protein products that affect aggregate accumulations in the cardiomyocytes. METHODS AND RESULTS: Primary mouse cardiomyocytes that accumulate aggregates as a result of a mutant CryAB (αB-crystallin) causative for human desmin-related cardiomyopathy were used for a total genome-wide screen to identify gene products that affected aggregate formation. We infected cardiomyocytes using a short hairpin RNA lentivirus library in which the mouse genome was represented. The screen identified multiple candidates in many cell signaling pathways that were able to mediate significant decreases in aggregate levels. CONCLUSIONS: Subsequent validation of one of these candidates, Jak1 (Janus kinase 1), a tyrosine kinase of the nonreceptor type, confirmed the usefulness of this approach in identifying previously unsuspected players in proteotoxic processes.
RATIONALE: Postmitotic cells, such as cardiomyocytes, seem to be particularly susceptible to proteotoxic stimuli, and large, proteinaceous deposits are characteristic of the desmin-related cardiomyopathies and crystallin cardiomyopathic diseases. Increased activity of protein clearance pathways in the cardiomyocyte, such as proteasomal degradation and autophagy, has proven to be beneficial in maintaining cellular and cardiac function in the face of multiple proteotoxic insults, holding open the possibility of targeting these processes for the development of effective therapeutics. OBJECTIVE: Here, we undertake an unbiased, total genome screen for RNA transcripts and their protein products that affect aggregate accumulations in the cardiomyocytes. METHODS AND RESULTS: Primary mouse cardiomyocytes that accumulate aggregates as a result of a mutant CryAB (αB-crystallin) causative for humandesmin-related cardiomyopathy were used for a total genome-wide screen to identify gene products that affected aggregate formation. We infected cardiomyocytes using a short hairpin RNA lentivirus library in which the mouse genome was represented. The screen identified multiple candidates in many cell signaling pathways that were able to mediate significant decreases in aggregate levels. CONCLUSIONS: Subsequent validation of one of these candidates, Jak1 (Janus kinase 1), a tyrosine kinase of the nonreceptor type, confirmed the usefulness of this approach in identifying previously unsuspected players in proteotoxic processes.
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Authors: Sonia R Singh; Moritz Meyer-Jens; Erda Alizoti; W Clark Bacon; Gregory Davis; Hanna Osinska; James Gulick; Silke Reischmann-Düsener; Ellen Orthey; Patrick M McLendon; Jeffery D Molkentin; Saskia Schlossarek; Jeffrey Robbins; Lucie Carrier Journal: Autophagy Date: 2020-12-27 Impact factor: 16.016