Translation Initiation inSaccharomyces cerevisiaeMitochondria: Functional Interactions Among Mitochondrial Ribosomal Protein Rsm28p, Initiation Factor 2, Methionyl-tRNA-Formyltransferase and Novel Protein Rmd9p

Abstract

Rsm28p is a dispensable component of the mitochondrial ribosomal small subunit in Saccharomyces cerevisiae that is not related to known proteins found in bacteria. It was identified as a dominant suppressor of certain mitochondrial mutations that reduced translation of the COX2 mRNA. To explore further the function of Rsm28p, we isolated mutations in other genes that caused a synthetic respiratory defective phenotype together with rsm28D. These mutations identified three nuclear genes: IFM1, which encodes the mitochondrial translation initiation factor 2 (IF2); FMT1, which encodes the methionyl-tRNA-formyltransferase; and RMD9, a gene of unknown function. The observed genetic interactions strongly suggest that the ribosomal protein Rsm28p and Ifm1p (IF2) have similar and partially overlapping functions in yeast mitochondrial translation initiation. Rmd9p, bearing a TAP-tag, was localized to mitochondria and exhibited roughly equal distribution in soluble and membrane-bound fractions. A small fraction of the Rmd9-TAP sedimented together with presumed monosomes, but not with either individual ribosomal subunit. Thus, Rmd9 is not a ribosomal protein, but may be a novel factor associated with initiating monosomes. The poorly respiring rsm28D, rmd9-V363I double mutant did not have a strong translationdefective phenotype, suggesting that Rmd9p may function upstream of translation initiation, perhaps at the level of localization of mitochondrially coded mRNAs.

Rmd9p Controls the Processing/Stability of Mitochondrial mRNAs and Its Overexpression Compensates for a Partial Deficiency of Oxa1p in Saccharomyces cerevisiae

“…Thus, the absence of Rmd9p leads to some mtDNA instability on glucose-rich medium. However, the absence of Rmd9p leads to the rapid loss of mtDNA in the strains whose mtDNA contains introns (Williams et al 2007, this issue; data not shown).…”

Yeast AEP3p Is an Accessory Factor in Initiation of Mitochondrial Translation

“…The function(s) of the PPR motifs shared by these four proteins are not known in any detail, but PPR motifs have been implicated in RNA binding and post-transcriptional gene regulation in organelles, including transcript processing and stabilization, tRNA binding, and translation (42,60,61). The discovery that Aep3p (this work) and Rmd9p (49) are also functionally linked to mitochondrial translation initiation factors suggests that the coupling between mRNA stability and translation may be a general phenomenon mediated by RNA-binding proteins and occurs mechanistically at the initiation stage.…”

“…Genetic interaction with the genes encoding methionyl-tRNA formyltransferase and ymIF2 provides strong evidence that Rsm28p has a role in translation initiation. Williams et al (49) speculate that the small subunit protein Rsm28p might provide IF1 and/or IF3-like functions in yeast mitochondrial translation initiation.…”

Yeast PPR proteins, watchdogs of mitochondrial gene expression

“…In S. cerevisiae, Rmd9 encodes a PPR protein and the gene was isolated in two independent screens: as a multi-copy suppressor of point mutations in Oxa1, which encodes an inner membrane insertase for newly translated mitochondrial proteins ( Table 2), and as a synthetic respiratory defective mutant of the deletion of a small ribosomal subunit gene, Rsm28 (Δrsm28 alone only leads to a partial respiratory deficiency) ( Table 2). 19,30 It was also shown that a small fraction of Rmd9 co-sediments with the small subunit of the mitochondrial ribosome in sucrose gradients. Thus, Rmd9 interacts with the ribosome but is not a ribosomal subunit per se.…”