Autophagic Burden on Calcium Signaling in Sympathetic Activation

Abstract

Signaling via calcium plays essential roles in all aspects of cell functions. Macroautophagy is a catabolic process that regulates important aspects of cell metabolism. While many studies have indicated that calcium is involved at various steps in the regulation of autophagic flux, the impact of autophagic flux on calcium signaling is unknown. We conceptually hypothesized that an “autophagic burden” exists in which enhanced requirement of the cell to process autophagic cargos would exert negative impact on calcium signaling. To test this hypothesis, we utilized an “non‐stimulation” approach to increase the “autophagic burden” by heterologously expressing in cells the wildtype microtubule‐associated proteins 1A/1B light chain 3B (LC3) or a mutant version in which the C‐terminal glycine responsible for ATG4‐mediated cleavage for LC3 lipidation has been removed (LC3DG) and examined the impact on adrenergic receptor‐mediated calcium signaling. HEK293 cells produced oscillatory intracellular calcium signals in response to agonists of b adrenoceptors (isoproterenol), α1A adrenoceptor (A61603), or a combination thereof (10 mM norepinephrine, (NE)). Cells expressing a fusion between the RFP mKate2 and wildtype LC3 (mKate2‐LC3) responded to 10 mM NE with substantial reductions in the number of intracellular calcium oscillations in the physiological ranges of 300 – 600 nM, 600 – 900 nM, or 900 – 1200 nM, as compared to the non‐expressing cells examined in the same microscopic fields, recognized by absence of mKate2 fluorescence. In contrast, cells expressing the mKate2‐LC3DG produced the same number of oscillatory calcium signals compared to non‐expressing cells in the same microscopic fields across the same ranges of cytoplasmic calcium stimulated by NE. The data suggest that an “autophagic burden” exists that may dampen the effects of acute sympathetic stimulation significantly via reduction in adrenoceptor‐mediated calcium signals.Support or Funding InformationIOER Grant 091707 to Q‐KT