Protein Kinase G Stimulates Chaperone Mediated Autophagy in Cardiomyocytes

Protein quality control (PQC) systems are crucial for eliminating misfolded or damaged proteins, and their dysfunction contributes to heart failure pathogenesis. The carboxyl terminus of Hsc70-interacting protein (Chip), a co-chaperone with E3 ligase activity, plays a central role in cardiac PQC. Chaperone-mediated autophagy (CMA) is a selective degradation process mediated by the lysosome and involved internalization through lysosome-associated membrane protein 2A (Lamp2A). For CMA to occur, the Hsc70/Chip complex recognizes a KFERQ motif on a substrate protein, will then bind to the substrate protein, and translocate it to the Lamp2A receptor. We find Lamp2A expression is downregulated in the myocardium of human heart failure with either reduced or preserved ejection fraction (HFrEF, HFpEF), suggesting impaired CMA and a potential therapeutic opportunity. However, little is known about the mechanisms regulating CMA in the myocardium or if it is possible to pharmacologically stimulate CMA in disease conditions. Here we test if phosphorylation of Chip at serine 19 by protein kinase G (PKG) stimulates CMA in cardiomyocytes. Preliminary data indicate that PKG phosphorylation of Chip provides cardioprotective effects under ischemic stress. To investigate this further, we have generated CHIP mutants, including a phosphonull (S19A) and a phosphomimic (S19E), to dissect the role of PKG-mediated regulation of CMA to enhance cardiomyocyte CMA. We find phosphorylated Chip (S19E) is more localized at the lysosome than non-phosphorylated Chip (S19A), suggesting greater CMA activity. We confirm these findings using a BioID proximity labeling system to identify protein-protein interactions and that increasing CMA via PKG/Chip S19 phosphorylation protects cardiomyocytes from proteotoxic stress. Collectively our findings suggest PKG phosphorylation of Chip at serine 19 is a new mechanism regulating CMA in the heart; which has therapeutic potential for improving myocardial PQC and function. NHLBI HL169273, AHA 20TPA35500008. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.