Mitochondrial Matrix Ca²⁺ Accumulation Regulates Cytosolic NAD⁺/NADH Metabolism, Protein Acetylation, and Sirtuin Expression

Abstract

Mitochondrial calcium uptake stimulates bioenergetics and drives energy production in metabolic tissue. It is unknown how a calcium-mediated acceleration in matrix bioenergetics would influence cellular metabolism in glycolytic cells that do not require mitochondria for ATP production. Using primary human endothelial cells (ECs), we discovered that repetitive cytosolic calcium signals (oscillations) chronically loaded into the mitochondrial matrix. Mitochondrial calcium loading in turn stimulated bioenergetics and a persistent elevation in NADH. Rather than serving as an impetus for mitochondrial ATP generation, matrix NADH rapidly transmitted to the cytosol to influence the activity and expression of cytosolic sirtuins, resulting in global changes in protein acetylation. In endothelial cells, the mitochondrion-driven reduction in both the cytosolic and mitochondrial NAD⁺/NADH ratio stimulated a compensatory increase in SIRT1 protein levels that had an anti-inflammatory effect. Our studies reveal the physiologic importance of mitochondrial bioenergetics in the metabolic regulation of sirtuins and cytosolic signaling cascades.

ACKNOWLEDGMENTS

We thank David Hockenbery and Daciana Margineantu for their assistance in the Seahorse assay measurements and Seahorse Bioscience for providing reagents and technical instruction. We also thank Gerda Breitwieser for the CaSR plasmid and technical advice.

This study was supported by HL094536 and a UW RRF award to B.J.H.