Novel players of mitochondrial communication with lysosome and endoplasmic reticulum

Abstract

Background: Mitochondria fulfil several functions that are essential for cellular homeostasis. This organelle forms a dynamic network, whose morphology is controlled by a set of GTPases dynamin-like proteins at the outer mitochondrial membrane (OMM) and inner mitochondrial membrane (IMM). In the IMM, the fusion process is mediated by OPA1. The activity of this pro-fusion protein is regulated, at least in part, by the mitochondrial metallo-protease OMA1. Loss of OMA1 prevents the processing of OPA1 and cell death induced by different stress conditions, indicating that OMA1 has mainly a pro-apoptotic function. Mitochondria interacts physically and functionally with other organelles, including lysosomes and endoplasmic reticulum (ER). Several studies in yeast and mammalian cells shown that vacuole/lysosomal dysfunction impairs mitochondrial homeostasis, via an unknown mechanism. Similarly, mitochondrial dysfunction impair lysosomal homeostasis, showing the bi-directional communication and the necessity to prevent damage of at least of one of these organelles during mitochondria/lysosome dysfunction. Mitochondria-ER communication plays an important role in cardiac pathophysiology. Norepinephrine (NE) treatment induces mitochondrial fragmentation and reduces mitochondria-ER communication, leading to the activation of calcineurin, a pro-hypertrophic cytosolic protein. Angiotensin-(1-9) is a nine amino peptide, member of the non-canonical renin-angiotensin system, with anti-hypertrophic properties in-vitro and in-vivo. However, the mechanism of action of angiotensin-(1-9) remains unknown. Hypothesis: “OMA1 prevents mitochondrial dysfunction and cell death induced by loss of lysosomal acidification” and “angiotensin-(1-9) treatment prevents the loss mitochondria-ER communication during cardiomyocyte hypertrophy”. Aims: To study the effect of loss of lysosomal acidification on mitochondrial homeostasis and the effect of angiotensin-(1-9) treatment over mitochondria-ER communication. Results: First, to address the effect of loss of lysosomal acidification over mitochondrial homeostasis, mouse embryonic fibroblast (MEF) wild type (WT) and OMA1 knockout (OKO) cells were treated with the V-ATPase inhibitor Bafilomycin A1 (Baf A1). Loss of lysosomal acidification induces cell death in WT, but more strongly in OKO cells, suggesting that OMA1 was required for cell viability under these experimental conditions. Nevertheless, the inhibition of early steps of autophagy did not affect cell viability, indicating that the effect of Baf A1 over cell death was independent of autophagy. Baf A1 reduced oxygen consumption rate (OCR) and glutamine incorporation into the tricarboxylic acid cycle (TCA) cycle in WT and OKO cells. OMA1 loss increased glutamine the incorporation into the TCA cycle. Loss of lysosomal acidification reduced iron availability, inducing an iron starvation response and reduction of iron-sulfur cluster binding proteins. Iron supplementation rescues respiration, iron-sulfur cluster binding protein levels and cell viability, indicating that iron is the main mediator of lysosome-mitochondria communication. Second, to address the effect of angiotensin-(1-9) over mitochondria-ER communication, neonatal rat cardiomyocytes were treated with NE in presence or absence of angiotensin-(1-9). This peptide prevented cardiomyocyte hypertrophy, mitochondrial fragmentation and loss of mitochondria-ER communication. Interestingly, angiotensin-(1-9) treatment by itself stimulated mitochondrial elongation and increased mitochondrial calcium buffer capacity. Finally, angiotensin-(1-9) treatment prevented the activation of Calcineurin/NFAT pathway triggered by NE, suggesting that this peptide regulates cytosolic calcium signalling. Conclusions. OMA1 plays a pro-survival role in conditions of V-ATPase inhibition and regulates glutamine incorporation into TCA cycle. Loss of lysosomal acidification impairs mitochondrial homeostasis by reducing iron availability, suggesting that iron plays a key role in lysosome-mitochondria communication. Angiotensin-(1-9) treatment prevents cardiomyocyte hypertrophy, mitochondrial fragmentation and loss of mitochondria-ER communication induced by NE. Angiotensin-(1-9) anti-hypertrophic mechanism involves the regulation of cytosolic calcium signalling, probably by keeping mitochondria-ER functional communication