bims-cytox1 2017-08-27 papers

J Biol Chem. 2017 Aug 16. pii: jbc.M117.801811. [Epub ahead of print]

Cox16 protein is physically associated with Cox1p assembly intermediates and with cytochrome oxidase.

Mitochondrial cytochrome oxidase (COX) catalyzes the last step in the respiratory pathway. In Saccharomyces cervisiae this inner membrane complex is composed of 11 protein subunits. Expression of COX is assisted by some 2 dozen ancillary proteins that intercede at different stages of the assembly pathway. One such protein, Cox16p, encoded by COX16, was shown to be essential for activity and assembly of COX. The function of Cox16p, however, has not been determined. We present evidence that Cox16p is present in Cox1p assembly intermediates and in COX. This is based on the finding that Cox16p, tagged with a dual poly-histidine and protein C tag, co-immunopurified with Cox1p assembly intermediates. The pull-down assays also indicated the presence of Cox16p in mature COX and in supercomplexes consisting of COX and the bc1 complex. From the Western signal strengths, Cox16p appears to be substoichiometric with Cox1p and Cox4p, which could indicate that Cox16p is only present in a fraction of COX. In conclusion, our results indicate that Cox16p is a constituent of several Cox1p assembly intermediates and of COX.

Keywords: cytochrome c oxidase (Complex IV); electron transfer complex; membrane biogenesis; mitochondria; yeast

DOI: https://doi.org/10.1074/jbc.M117.801811

J Biol Chem. 2017 Aug 18. pii: jbc.M117.788588. [Epub ahead of print]

Proteolytic cleavage by the IMP complex or Oct1 peptidase controls the localization of the yeast peroxiredoxin Prx1 to distinct mitochondrial compartments.

Fernando Gomes, Flavio Romero Palma, Mario H Barros, Eduardo T Tsuchida, Helena G Turano, Thiago G P Alegria, Marilene Demasi, Luis E S Netto.

Yeast Prx1 is a mitochondrial 1-Cys peroxiredoxin that catalyzes the reduction of endogenously generated H2O2 Prx1 is synthesized on cytosolic ribosomes as a preprotein with a cleavable N-terminal presequence that is the mitochondrial targeting signal, but the mechanisms underlying Prx1 distribution to distinct mitochondrial subcompartments are unknown. Here, we provide direct evidence of the following dual mitochondrial localization of Prx1: a soluble form in the intermembrane space and a form in the matrix weakly associated with the inner mitochondrial membrane. We show that Prx1 sorting into the intermembrane space likely involves the release of the protein precursor within the lipid bilayer of the inner membrane, followed by cleavage by the inner membrane peptidase (IMP). We also found that during its import into the matrix compartment, Prx1 is sequentially cleaved by mitochondrial processing peptidase (MPP) and then by octapeptidyl aminopeptidase 1 (Oct1). Oct1 cleaved eight amino acid residues from the N-terminal region of Prx1 inside the matrix, without interfering with its peroxidase activity in vitro Remarkably, the processing of Prx proteins by Oct1 appears to be an evolutionarily conserved process since yeast Oct1 could cleave the human mitochondrial peroxiredoxin Prx3 when expressed in Saccharomyces cerevisiae Altogether, the processing of peroxiredoxins by Imp2 or Oct1 likely represents systems that control the localization of Prxs into distinct compartments and thereby contribute to various mitochondrial redox processes..

Keywords: Inner membrane peptidase complex IMP complex; Mitochondrial processing peptidase (MPP); Octapeptidylpeptidase 1 Oct1; hydrogen peroxide; mitochondria; oxidative stress; peroxiredoxin; protein import; protein sorting; yeast

DOI: https://doi.org/10.1074/jbc.M117.788588

J Biol Chem. 2017 Aug 18. pii: jbc.M117.781773. [Epub ahead of print]

Two Plant-Derived Aporphinoid Alkaloids Exert Their Antifungal Activity by Disrupting Mitochondrial Iron-Sulfur Cluster Biosynthesis.

Siddharth K Tripathi, Tao Xu, Qin Feng, Bharathi Avula, Xiaomin Shi, Xuewen Pan, Melanie M Mask, Scott R Baerson, Melissa R Jacob, Ranga Rao Ravu, Shabana I Khan, Xing-Cong Li, Ikhlas A Khan, Alice M Clark, Ameeta K Agarwal.

Eupolauridine and liriodenine are plant-derived aporphinoid alkaloids that exhibit potent inhibitory activity against the opportunistic fungal pathogens Candida albicans and Cryptococcus neoformans However, the molecular mechanism of this antifungal activity is unknown. In this study, we show that eupolauridine 9591 (E9591), a synthetic analog of eupolauridine, and liriodenine methiodide (LMT), a methiodide salt of liriodenine, mediate their antifungal activities by disrupting mitochondrial iron-sulfur (Fe-S) cluster synthesis. Several lines of evidence supported this conclusion. First, both E9591 and LMT elicited a transcriptional response indicative of iron imbalance, causing the induction of genes that are required for iron uptake and for the maintenance of cellular iron homeostasis. Second, a genome-wide fitness profile analysis showed that yeast mutants with deletions in iron homeostasis-related genes were hypersensitive to E9591 and LMT. Third, treatment of wild-type yeast cells with E9591 or LMT generated cellular defects that mimicked deficiencies in mitochondrial Fe-S cluster synthesis including an increase in mitochondrial iron levels, a decrease in the activities of Fe-S cluster enzymes, a decrease in respiratory function, and an increase in oxidative stress. Collectively, our results demonstrate that E9591 and LMT perturb mitochondrial Fe-S cluster biosynthesis; thus, these two compounds target a cellular pathway that is distinct from the pathways commonly targeted by clinically used antifungal drugs. Therefore, the identification of this pathway as a target for antifungal compounds has potential applications in the development of new antifungal therapies.

Keywords: drug action; infectious disease; iron-sulfur protein; small molecule; yeast

DOI: https://doi.org/10.1074/jbc.M117.781773