Abstract
Genetic and microscopic approaches using Saccharomyces cerevisiae have identified many proteins that play a role in mitochondrial dynamics, but it is possible that other proteins and pathways that play a role in mitochondrial division and fusion remain to be discovered. Mutants lacking mitochondrial fusion are characterized by rapid loss of mitochondrial DNA. We took advantage of a petite-negative mutant that is unable to survive mitochondrial DNA loss to select for mutations that allow cells with fusion-deficient mitochondria to maintain the mitochondrial genome on fermentable medium. Next-generation sequencing revealed that all identified suppressor mutations not associated with known mitochondrial division components were localized to PDR1 or PDR3, which encode transcription factors promoting drug resistance. Further studies revealed that at least one, if not all, of these suppressor mutations dominantly increases resistance to known substrates of the pleiotropic drug resistance pathway. Interestingly, hyperactivation of this pathway did not significantly affect mitochondrial shape, suggesting that mitochondrial division was not greatly affected. Our results reveal an intriguing genetic connection between pleiotropic drug resistance and mitochondrial dynamics.
Highlights
Genetic and microscopic approaches using Saccharomyces cerevisiae have identified many proteins that play a role in mitochondrial dynamics, but it is possible that other proteins and pathways that play a role in mitochondrial division and fusion remain to be discovered
With the goal of isolating new mitochondrial division machinery or proteins and pathways impinging upon the mitochondrial fission process, we asked that fusion-incompetent cells maintain mitochondrial DNA (mtDNA), yet we performed our selection on glucose-containing medium
Because genetic evidence suggested that pleiotropic drug resistance (PDR) activation might inhibit mitochondrial division, we examined the mitochondrial morphology of PDR1-249 cells by fluorescence microscopy after transformation with green fluorescent protein (GFP) targeted to mitochondria by the Cox4 presequence (Sesaki and Jensen 1999), but initial observation indicated that there was minimal, if any difference in mitochondrial morphology between WT and PDR1249 cells
Summary
Genetic and microscopic approaches using Saccharomyces cerevisiae have identified many proteins that play a role in mitochondrial dynamics, but it is possible that other proteins and pathways that play a role in mitochondrial division and fusion remain to be discovered. Association with another organelle system, the endoplasmic reticulum (ER), can determine the location of mitochondrial scission (Friedman et al 2011) These exciting findings raise the possibility that additional genes and pathways that influence the rate of mitochondrial division might be identified and mechanistically studied using S. cerevisiae. Mutations blocking mitochondrial fusion result in fragmentation of mitochondrial tubules and, for reasons that are not yet understood, mitochondrial DNA (mtDNA) loss (Hermann et al 1998; Rapaport et al 1998) This mitochondrial fragmentation is dependent upon the mitochondrial division machinery; cells lacking both the ability to fuse mitochondria and the capacity to divide mitochondria are able to maintain both tubular mitochondrial morphology and mtDNA (Bleazard et al 1999; Sesaki and Jensen 1999). By sequencing the entire genomes of suppressorcontaining isolates, we found that dominant mutations activating the pleiotropic drug resistance (PDR) pathway can allow cells lacking mitochondrial fusion components to keep the mitochondrial genome, providing additional evidence of a functional relationship between the PDR pathway and mitochondrial biogenesis
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