Interaction of MDM33 with mitochondrial inner membrane homeostasis pathways in yeast

Membrane homeostasis affects mitochondrial dynamics, morphology, and function. Here we report genetic and proteomic data that reveal multiple interactions of Mdm33, a protein essential for normal mitochondrial structure, with components of phospholipid metabolism and mitochondrial inner membrane homeostasis. We screened for suppressors of MDM33 overexpression-induced growth arrest and isolated binding partners by immunoprecipitation of cross-linked cell extracts. These approaches revealed genetic and proteomic interactions of Mdm33 with prohibitins, Phb1 and Phb2, which are key components of mitochondrial inner membrane homeostasis. Lipid profiling by mass spectrometry of mitochondria isolated from Mdm33-overexpressing cells revealed that high levels of Mdm33 affect the levels of phosphatidylethanolamine and cardiolipin, the two key inner membrane phospholipids. Furthermore, we show that cells lacking Mdm33 show strongly decreased mitochondrial fission activity indicating that Mdm33 is critical for mitochondrial membrane dynamics. Our data suggest that MDM33 functionally interacts with components important for inner membrane homeostasis and thereby supports mitochondrial division

The UbiB family member Cqd1 forms a novel membrane contact site in mitochondria

Mitochondria are essential organelles of eukaryotic cells and are characterized by their unique and complex membrane system. They are confined from the cytosol by an envelope consisting of two membranes. Signals, metabolites, proteins and lipids have to be transferred across these membranes via proteinaceous contact sites to keep mitochondria functional. In the present study, we identified a novel mitochondrial contact site in Saccharomyces cerevisiae that is formed by the inner membrane protein Cqd1 and the outer membrane proteins Por1 and Om14. Similar to what is found for the mitochondrial porin Por1, Cqd1 is highly conserved, suggesting that this complex is conserved in form and function from yeast to human. Cqd1 is a member of the UbiB protein kinase-like family (also called aarF domain-containing kinases). It was recently shown that Cqd1, in cooperation with Cqd2, controls the cellular distribution of coenzyme Q by a yet unknown mechanism. Our data suggest that Cqd1 is additionally involved in phospholipid homeostasis. Moreover, overexpression of CQD1 and CQD2 causes tethering of mitochondria to the endoplasmic reticulum, which might explain the ability of Cqd2 to rescue ERMES deletion phenotypes

Intramitochondrial Transport of Phosphatidic Acid in Yeast by a Lipid Transfer Protein

Mitochondria are dynamic organelles whose function depends on intramitochondrial phospholipid synthesis and the supply of membrane lipids from the endoplasmic reticulum. How phospholipids are transported to and in-between mitochondrial membranes remained unclear. We identified Ups1, a yeast member of a conserved family of intermembrane space proteins, as a lipid transfer protein that can shuttle phosphatidic acid between mitochondrial membranes. Lipid transfer required the dynamic assembly of Ups1 with Mdm35 and allowed conversion of phosphatidic acid to cardiolipin in the inner membrane. High cardiolipin concentrations prevented membrane dissociation of Ups1, leading to its proteolysis and inhibiting transport of phosphatidic acid and cardiolipin synthesis. Thus, intramitochondrial lipid trafficking may involve a regulatory feedback mechanism that limits the accumulation of cardiolipin in mitochondria

Selective retention of dysfunctional mitochondria during asymmetric cell division in yeast.

Decline of mitochondrial function is a hallmark of cellular aging. To counteract this process, some cells inherit mitochondria asymmetrically to rejuvenate daughter cells. The molecular mechanisms that control this process are poorly understood. Here, we made use of matrix-targeted D-amino acid oxidase (Su9-DAO) to selectively trigger oxidative damage in yeast mitochondria. We observed that dysfunctional mitochondria become fusion-incompetent and immotile. Lack of bud-directed movements is caused by defective recruitment of the myosin motor, Myo2. Intriguingly, intact mitochondria that are present in the same cell continue to move into the bud, establishing that quality control occurs directly at the level of the organelle in the mother. The selection of healthy organelles for inheritance no longer works in the absence of the mitochondrial Myo2 adapter protein Mmr1. Together, our data suggest a mechanism in which the combination of blocked fusion and loss of motor protein ensures that damaged mitochondria are retained in the mother cell to ensure rejuvenation of the bud

Cristae-dependent quality control of the mitochondrial genome.

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Cristae-dependent quality control of the mitochondrial genome.

Mitochondrial genomes (mtDNA) encode essential subunits of the mitochondrial respiratory chain. Mutations in mtDNA can cause a shortage in cellular energy supply, which can lead to numerous mitochondrial diseases. How cells secure mtDNA integrity over generations has remained unanswered. Here, we show that the single-celled yeast Saccharomyces cerevisiae can intracellularly distinguish between functional and defective mtDNA and promote generation of daughter cells with increasingly healthy mtDNA content. Purifying selection for functional mtDNA occurs in a continuous mitochondrial network and does not require mitochondrial fission but necessitates stable mitochondrial subdomains that depend on intact cristae morphology. Our findings support a model in which cristae-dependent proximity between mtDNA and the proteins it encodes creates a spatial “sphere of influence,” which links a lack of functional fitness to clearance of defective mtDNA