The essential products of the yeast mitochondrial translation system are seven hydrophobic membrane proteins and Var1p, an unusual yet essential protein in the small ribosomal subunit. Translation of the membrane proteins is dependent on nucleus-encoded, mRNA-specific translational activators that recognize sites with the 5'-untranslated leaders of the target mRNAs. These translational activators are membrane associated and appear to modulate the expression of the membrane proteins and target their translation to the inner membrane. It is not known whether the synthesis of Var1p also requires specific translational activation. The identification of nuclear genes for VAR1-specific translational activators is complicated by the instability of rho+ mtDNA in the absence of a functional mitochondrial translation system. Thus, mutations that impair VAR1 expression will cause inactivation of the mitochondrial genetic system through the secondary conversion of the mutants to rho- or rho cytoplasmic petites. The research objectives in this proposal are based on the use of a novel experimental system in which functional Var1p is supplied from a recorded gene in the nucleus (VAR1/u) and the VAR1 coding sequences in mtDNA is replaced with a sequence recorded for the mitochondrial expression of a nuclear encoded arginine biosynthetic enzyme (Arg8p), thus creating a var1::ARG8/m reporter gene. This set of relocated genes opens the way to address the following questions about the VAR1 gene, its protein product and the formation and function of mitochondrial translational apparatus in general:
Aim 1 : Are mRNA-specific translational activations required for VAR1 expression? Aim 2: What is the functional role of Var1P in the ribosome? Aim 3: Can a mitochondria-free translation system be developed using VAR1 mRNA? Aim 4: What roles does mitochondrial translation play in the complementation, recombination, and segregation of two different rho- mitochondrial genomes in newly formed zygotes.
| Fiori, Alessandro; Mason, Thomas L; Fox, Thomas D (2003) Evidence that synthesis of the Saccharomyces cerevisiae mitochondrially encoded ribosomal protein Var1p may be membrane localized. Eukaryot Cell 2:651-3 |
