Metabolism and translation of mitochondrial RNA in the yeast S. pombd
Mitochondria are organelles present in most eukaryotic cells and specialized in the production of energy via the respiratory chain located in their inner membrane. Mitochondria have their own genome and their own system of gene expression, which is involved in the biogenesis of the respiratory complexes. The mitochondrial translation machinery, like the respiratory complexes, has a dual genetic origin, both nuclear and mitochondrial. Numerous human diseases result from defects in the expression of mitochondrial genes and especially mutations of factors involved in mitochondrial translation. The yeast Schizosaccharomyces pombe is a useful model for the identification and functional analysis of these factors because it is a simple organism that is physiologically closer to higher eukaryotes than Saccharomyces cerevisiae. During my PhD I first participated in the development of new tools to further our understanding of mitochondrial translation, by tagging the small and large subunits of S. pombe mitoribosome. In addition I set up fractionation experiments on sucrose gradients to analyze the sedimentation of associated or dissociated mitochondrial ribosomes and test whether given factors are bound to mitoribosome. I also became interested in factors that could act in the termination mitochondrial translation. Surprisingly, the only factor known to recognize stop codons in S. pombe, Mrf1, is not essential, thus I tried to determine which other proteins might also be involved in translation termination. S. pombe contains four predicted peptidyl tRNA hydrolases (Pth), two of which, Pth3 and Pth4, have a GGQ motif like Mrf1, which is thought to contribute directly to the hydrolysis of the peptidyl-tRNA bond. Thus they seemed to be good candidates to explain how S. pombe can survive without Mrf1. I have shown that Pth3 and Pth4 play a role in mitochondrial biogenesis and that Pth4 is both a high copy suppressor and a synthetic lethal of the ∆mrf1 mutant. Finally I worked on the Pentatrico Peptide Repeat family of proteins (PPR), predicted to be involved in the metabolism of mitochondrial RNA. There are at least nine PPR proteins in S. pombe named Ppr1 to Ppr8 and the mitochondrial RNA polymerase, Rpo41. The study of these PPR proteins has shown that all of them are involved in the metabolism of RNA at different stages, mainly stability and translation, and that they often have specific targets. For example Ppr3 is involved in the stability of the small rRNA rns while Ppr4 is a specific activator of the translation of cox1 and Ppr2 is a general activator of mitochondrial translation whose target remains to be identified. Overall, these studies show that S. pombe is an excellent model for mitochondrial functions, both for fundamental studies and as a tool for understanding more complex organisms such as man.