108 - Tuning Muscle Energy Homeostasis: The Role of Nix Durig Contractile Activity

Abstract

Exercise is instrumental in reversing derangements to muscle metabolism, such as insulin-resistance during type 2 diabetes. Mitochondrial quality control is important for efficient use of metabolic fuels in muscle, but the mechanism remains unclear. In an in vitro model of skeletal muscle, we observed an increase in the protein Nix contraction; this protein mediates the specific removal of dysfunctional mitochondria (mitophagy). Therefore, we hypothesize that Nix is elevated in contracting skeletal muscle, increasing the removal of dysfunctional mitochondria, to aid muscle energy homeostasis. Nix-mediated mitophagy was investigated in an in vitro model of skeletal muscle (C2C12 myotubes) where contraction was induced by electrical pulses for 1 hour. Levels of Nix and mitochondrial-biogenesis proteins were determined by immunoblotting. Next, fluorescent biosensors were used to assess levels of mitochondrial and nuclear calcium, and mitophagy. Following contraction, we found elevated levels of Nix (1.9-fold), and mitochondrial-biogenesis proteins PGC-1α (2.5-fold) and NRF2 (1.4-fold). Next, we examined mitochondrial clearance by assessing mitochondrial acidification, which increased (∼1.4-fold) following contraction as would be expected once mitochondria enter the acidic lysosome. Finally, we observed an increase in nuclear-calcium (∼2-fold) and a small but significant rise in mitochondrial calcium, suggesting changes in gene expression and elevated metabolic output, respectively. Together, these data uncover the potential contribution of Nix to muscle metabolism and the efficient use of metabolic fuels that is disrupted by the onset of insulin-resistance in diabetes.