GTP Is Required for Iron-Sulfur Cluster Biogenesis in Mitochondria

Boominathan Amutha,Donna M Gordon,Yajuan Gu,Elise R Lyver,Andrew Dancis,Debkumar Pain

doi:10.1074/jbc.m706808200

Boominathan Amutha, Donna M Gordon + Show 4 more

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https://doi.org/10.1074/jbc.m706808200

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Abstract

Iron-sulfur (Fe-S) cluster biogenesis in mitochondria is an essential process and is conserved from yeast to humans. Several proteins with Fe-S cluster cofactors reside in mitochondria, including aconitase [4Fe-4S] and ferredoxin [2Fe-2S]. We found that mitochondria isolated from wild-type yeast contain a pool of apoaconitase and machinery capable of forming new clusters and inserting them into this endogenous apoprotein pool. These observations allowed us to develop assays to assess the role of nucleotides (GTP and ATP) in cluster biogenesis in mitochondria. We show that Fe-S cluster biogenesis in isolated mitochondria is enhanced by the addition of GTP and ATP. Hydrolysis of both GTP and ATP is necessary, and the addition of ATP cannot circumvent processes that require GTP hydrolysis. Both in vivo and in vitro experiments suggest that GTP must enter into the matrix to exert its effects on cluster biogenesis. Upon import into isolated mitochondria, purified apoferredoxin can also be used as a substrate by the Fe-S cluster machinery in a GTP-dependent manner. GTP is likely required for a common step involved in the cluster biogenesis of aconitase and ferredoxin. To our knowledge this is the first report demonstrating a role of GTP in mitochondrial Fe-S cluster biogenesis.

Highlights

Outer membrane function in fission and fusion of mitochondria, and these are required for morphologic changes of the organelle associated with the metabolic demand of the cell [4]
We show that isolated intact yeast mitochondria contain a nucleotide (GTP and ATP)-dependent machinery capable of forming new Fe-S clusters and inserting them into endogenous apoaconitase or imported apoferredoxin
We have shown that GTP acts in the mitochondrial matrix and must be hydrolyzed to facilitate Fe-S cluster biogenesis

Summary

Introduction

Outer membrane function in fission and fusion of mitochondria, and these are required for morphologic changes of the organelle associated with the metabolic demand of the cell [4]. Palmieri and co-workers [2] identified one such protein in yeast mitochondria as the GTP/GDP carrier and named the protein Ggc1p They demonstrated that Ggc1p allows exchange of cytosolic GTP for matrix GDP across the inner membrane. The Nm23-H4 enzyme exhibits activity capable of converting GDP to GTP using ATP as the phosphate donor in the mitochondrial matrix, and it is this activity that is responsible for rescuing the iron phenotypic defects of the mutant [8]. These results suggest that GTP in the mitochondrial matrix plays an important role in organellar iron homeostasis

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