Abstract
Mitochondria are highly dynamic organelles constantly undergoing fusion and fission. Ca2+ regulates many aspects of mitochondrial physiology by modulating the activity of several mitochondrial proteins. We previously showed that inhibition of constitutive IP3R-mediated Ca2+ transfer to the mitochondria leads to a metabolic cellular stress and eventually cell death. Here, we show that the decline of mitochondrial function generated by a lack of Ca2+ transfer induces a DRP-1 independent mitochondrial fragmentation that at an early time is mediated by an increase in the NAD+/NADH ratio and activation of SIRT1. Subsequently, AMPK predominates and drives the fragmentation. SIRT1 activation leads to the deacetylation of cortactin, favoring actin polymerization, and mitochondrial fragmentation. Knockdown of cortactin or inhibition of actin polymerization prevents fragmentation. These data reveal SIRT1 as a new player in the regulation of mitochondrial fragmentation induced by metabolic/bioenergetic stress through regulating the actin cytoskeleton.
Highlights
Mitochondria are a highly dynamic interconnected network of organelles that catalyze the synthesis of most cellular energy in the form of ATP, play a vital role in metal ion homeostasis, regulate several types of cell death and work as a hub for the biosynthesis of building blocks necessary for the generation of amino acids, lipids, and nucleotides (Ahn and Metallo, 2015; Spinelli and Haigis, 2018; Ward and Cloonan, 2019)
We show that the bioenergetic/metabolic stress caused by lack of Ca2+ transfer from the endoplasmic reticulum (ER) to the mitochondria induces mitochondrial fragmentation through a mechanism mediated by the action of SIRT1 on cortactin at an early time point, which is followed by the activation of AMPK acting through dynamin-related protein1 (Drp1) and SIRT1 upon a sustained bioenergetic/metabolic stress
As mitochondrial function is related to its morphology, we inhibited spontaneous IP3R-mediated Ca2+ signals in HeLa cells with the specific IP3R inhibitor Xestospongin B (XeB, 5 μM) for 1 h (Figure 1A) and mitochondrial morphology was determined by immunolabeling of the outer mitochondrial membrane (OMM) protein TOMM20 with confocal microscopy
Summary
Mitochondria are a highly dynamic interconnected network of organelles that catalyze the synthesis of most cellular energy in the form of ATP, play a vital role in metal ion homeostasis, regulate several types of cell death and work as a hub for the biosynthesis of building blocks necessary for the generation of amino acids, lipids, and nucleotides (Ahn and Metallo, 2015; Spinelli and Haigis, 2018; Ward and Cloonan, 2019). Physiological stimulation with InsP3-generating agonists generates an increase of cytosolic Ca2+ followed by a mitochondrial Ca2+ increase that induces a rise in the levels of NADPH due to a change in the pyruvate dehydrogenase (PDH) activity (Rizzuto et al, 1998; Robb-Gaspers et al, 1998). Corroborating this result, we found that in unstimulated conditions, the constitutive IP3Rmediated Ca2+ release is fundamental to maintain the optimal activity of PDH, energy levels and cellular homeostasis. Inhibition of the Ca2+ transfer to mitochondria by inhibition of either the IP3R or MCU causes a bioenergetic crisis that activates AMPK and autophagy as a part of a survival response (Cardenas et al, 2010)
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