Mdm38 protein depletion causes loss of mitochondrial K+/H+ exchange activity, osmotic swelling and mitophagy

Abstract

Loss of the MDM38 gene product in yeast mitochondria results in a variety of phenotypic effects including reduced content of respiratory chain complexes, altered mitochondrial morphology and loss of mitochondrial K þ /H þ exchange activity resulting in osmotic swelling. By use of doxycycline-regulated shut-off of MDM38 gene expression, we show here that loss of K þ /H þ exchange activity and mitochondrial swelling are early events, associated with a reduction in membrane potential and fragmentation of the mitochondrial reticulum. Changes in the pattern of mitochondrially encoded proteins are likely to be secondary to the loss of K þ /H þ exchange activity. The use of a novel fluorescent biosensor directed to the mitochondrial matrix revealed that the loss of K þ /H þ exchange activity was immediately followed by morphological changes of mitochondria and vacuoles, the close association of these organelles and finally uptake of mitochondrial material by vacuoles. Nigericin, a K þ /H þ ionophore, fully prevented these effects of Mdm38p depletion. We conclude that osmotic swelling of mitochondria triggers selective mitochondrial autophagy or mitophagy.

Novel Components of an Active Mitochondrial K+/H+ Exchange

“…Further tests involving submitochondrial inner membrane particles (SMPs) confirmed the near total lack of KHE activity (2). Addition of the synthetic KHE nigericin to mdm38⌬ cells restored all mitochondrial functions, including growth on nonfermentable substrates, ⌬, morphology, and KHE activity (4,5). This result strongly supported the conclusion that Mdm38 acts as an essential regulator or subunit of the mitochondrial KHE, because it is unlikely that a protein with only one transmembrane domain like Mdm38 forms the KHE itself.…”

“…Overexpression of Mrs3p, a Fe 2ϩ carrier, increased the mitochondrial ⌬ and moderately the KHE. Deletion of MRS3 had no effect on KHE, and mrs3⌬ mdm38⌬ mutants remained without synthetic phenotype, 5 excluding a role of Mrs3p as the KHE. These findings suggest an indirect role of Mrs3p on the KHE activity.…”

“…Cryofixation, freeze substitution, thin sectioning, and image acquisition were performed as described previously (5).…”

The pathways of mitophagy for quality control and clearance of mitochondria

“…Although mitochondria can be engulfed non-selectively along with other cytosolic contents during bulk autophagy, 31 32,33 Similarly, knockdown of an inhibitor of the mammalian F 0 F 1 ATPsynthase, IF1, causes a dramatic reduction in mitochondrial mass that correlates with increased autophagic activity. 34 Depolarization of mitochondria with the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) 35 or photo-irradiation 36 induces autophagic clearance of mammalian mitochondria while leaving other organelles such as peroxisomes intact.…”

Autophagy in the liver

“…86 In addition, microautophagy, rather than macroautophagy, may be preferentially utilized to remove mitochondria following nitrogen deprivation. Both selective and nonselective mitophagy can occur, and a mitochondrial outer membrane protein, Uth1, is required for selective degradation.…”