Mitofusin 2 Regulates STIM1 Migration from the Ca2+ Store to the Plasma Membrane in Cells with Depolarized Mitochondria

Karthika Singaravelu,Charmaine Nelson,Daniel Bakowski,Olga Martins De Brito,Siaw-Wei Ng,Joseph Di Capite,Trevor Powell,Luca Scorrano,Anant B Parekh

doi:10.1074/jbc.m110.174029

Abstract

Store-operated Ca2+ channels in the plasma membrane (PM) are activated by the depletion of Ca2+ from the endoplasmic reticulum (ER) and constitute a widespread and highly conserved Ca2+ influx pathway. After store emptying, the ER Ca2+ sensor STIM1 forms multimers, which then migrate to ER-PM junctions where they activate the Ca2+ release-activated Ca2+ channel Orai1. Movement of an intracellular protein to such specialized sites where it gates an ion channel is without precedence, but the fundamental question of how STIM1 migrates remains unresolved. Here, we show that trafficking of STIM1 to ER-PM junctions and subsequent Ca2+ release-activated Ca2+ channel activity is impaired following mitochondrial depolarization. We identify the dynamin-related mitochondrial protein mitofusin 2, mutations of which causes the inherited neurodegenerative disease Charcot-Marie-Tooth IIa in humans, as an important component of this mechanism. Our results reveal a molecular mechanism whereby a mitochondrial fusion protein regulates protein trafficking across the endoplasmic reticulum and reveals a homeostatic mechanism whereby mitochondrial depolarization can inhibit store-operated Ca2+ entry, thereby reducing cellular Ca2+ overload.

Highlights

In eukaryotic cells, a variety of different agonists, including hormones, neurotransmitters, and growth factors, elicit cellular responses through a rise in cytoplasmic Ca2ϩ concentration [1]
Some of the effects of mitochondria arise from their ability to buffer cytoplasmic Ca2ϩ and reduce Ca2ϩ-dependent inactivation of CRAC channels, growing evidence suggests that they might have an additional role in regulating CRAC channels that is unrelated to their ability to take up Ca2ϩ, produce ATP, and generate reactive oxygen species [30, 34]
We show here that mitochondrial depolarization suppresses STIM1 puncta formation and subsequent Orai1-dependent CRAC currents, and these inhibitory effects can be partially overcome by overexpression of either STIM1 or a STIM1 mutant that occupies endoplasmic reticulum (ER)-plasma membrane (PM) junctions in nonstimulated cells with intact stores

Summary

Introduction

A variety of different agonists, including hormones, neurotransmitters, and growth factors, elicit cellular responses through a rise in cytoplasmic Ca2ϩ concentration [1]. Mouse embryonic fibroblasts lacking Mfn display loosened ER-mitochondria tethering and reduced rate of mitochondrial Ca2ϩ uptake following InsP3mediated Ca2ϩ release from the ER [36]. Whether this physical uncoupling impacts upon spatially more distal events is unclear. We show here that mitochondrial depolarization suppresses STIM1 puncta formation and subsequent Orai1-dependent CRAC currents, and these inhibitory effects can be partially overcome by overexpression of either STIM1 or a STIM1 mutant that occupies ER-PM junctions in nonstimulated cells with intact stores. In cells lacking Mfn, STIM1 puncta formation and CRAC channel activity were independent of mitochondrial status, and analysis of Mfn mutants revealed a major role for mitochondrially targeted Mfn. Our findings reveal Mfn as an important component in the mechanism whereby mitochondrial depolarization inhibits CRAC channel activity

Methods

Results

Conclusion