Reconstitution of the mitochondrial calcium uniporter in yeast

E. Kovács, Yasemin Sancak, Kimberli J. Kamer, Molly Plovanich, Ashwini Jambhekar, Robert J. Huber, Michael A. Myre, Michael D. Blower, Vamsi K. Mootha

Research output: Contribution to journalArticle

76 Citations (Scopus)

Abstract

The mitochondrial calcium uniporter is a highly selective calcium channel distributed broadly across eukaryotes but absent in the yeast Saccharomyces cerevisiae. The molecular components of the human uniporter holocomplex (uniplex) have been identified recently. The uniplex consists of three membrane-spanning subunits-mitochondrial calcium uniporter (MCU), its paralog MCUb, and essential MCU regulator (EMRE)- and two soluble regulatory components-MICU1 and its paralog MICU2. The minimal components sufficient for in vivo uniporter activity are unknown. Here we consider Dictyostelium discoideum (Dd), a member of the Amoebazoa outgroup of Metazoa and Fungi, and show that it has a highly simplified uniporter machinery. We show that D. discoideum mitochondria exhibit membrane potential-dependent calcium uptake compatible with uniporter activity, and also that expression of DdMCU complements the mitochondrial calcium uptake defect in human cells lacking MCU or EMRE. Moreover, expression of DdMCU in yeast alone is sufficient to reconstitute mitochondrial calcium uniporter activity. Having established yeast as an in vivo reconstitution system, we then reconstituted the human uniporter. We show that coexpression of MCU and EMRE is sufficient for uniporter activity, whereas expression of MCU alone is insufficient. Our work establishes yeast as a powerful in vivo reconstitution system for the uniporter. Using this system, we confirm that MCU is the pore-forming subunit, define the minimal genetic elements sufficient for metazoan and nonmetazoan uniporter activity, and provide valuable insight into the evolution of the uniporter machinery.

Original languageEnglish
Pages (from-to)8985-8990
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number24
DOIs
Publication statusPublished - 2014

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Yeasts
Dictyostelium
Calcium
mitochondrial calcium uniporter
Calcium Channels
Eukaryota
Membrane Potentials
Saccharomyces cerevisiae
Mitochondria
Fungi
Membranes

ASJC Scopus subject areas

  • General

Cite this

Reconstitution of the mitochondrial calcium uniporter in yeast. / Kovács, E.; Sancak, Yasemin; Kamer, Kimberli J.; Plovanich, Molly; Jambhekar, Ashwini; Huber, Robert J.; Myre, Michael A.; Blower, Michael D.; Mootha, Vamsi K.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 111, No. 24, 2014, p. 8985-8990.

Research output: Contribution to journalArticle

Kovács, E, Sancak, Y, Kamer, KJ, Plovanich, M, Jambhekar, A, Huber, RJ, Myre, MA, Blower, MD & Mootha, VK 2014, 'Reconstitution of the mitochondrial calcium uniporter in yeast', Proceedings of the National Academy of Sciences of the United States of America, vol. 111, no. 24, pp. 8985-8990. https://doi.org/10.1073/pnas.1400514111
Kovács, E. ; Sancak, Yasemin ; Kamer, Kimberli J. ; Plovanich, Molly ; Jambhekar, Ashwini ; Huber, Robert J. ; Myre, Michael A. ; Blower, Michael D. ; Mootha, Vamsi K. / Reconstitution of the mitochondrial calcium uniporter in yeast. In: Proceedings of the National Academy of Sciences of the United States of America. 2014 ; Vol. 111, No. 24. pp. 8985-8990.
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