RELOCATION OF THE UNUSUAL VAR1 GENE FROM THE MITOCHONDRION TO THE NUCLEUS

The Var1 protein (Var1p) is an essential, stoichiometric component of the yeast mitochondrial small ribosomal subunit, and it is the only ma jor protein product of the mitochondrial genetic system that is not pa rt of an energy transducing complex of the inner membrane. Interesting ly, no mutations have been reported that affect the function of Var1p, presumably because loss of a functional mitochondrial translation sys tem leads to an instability of mtDNA. To study the structure, function and synthesis of Var1p, we have engineered yeast strains for the expr ession of this protein from a nuclear gene, VAR1(U), in which 39 nonst andard mitochondrial codons were converted to the universal code. Immu noblot analysis using an epitope-tagged form of Var1(U)p showed that t he nuclear-encoded protein was expressed and imported into the mitocho ndria. VAR1(U) was tested for its ability to complement a mutation in mtDNA, PZ206, which disrupts 3'-end processing of the VAR1 mRNA, causi ng greatly reduced synthesis of Var1p and a respiratory-deficient phen otype. Respiratory growth was restored in PZ206 mutants by transformat ion with a centromere plasmid carrying VAR1(U) under ADH1 promoter con trol, thus proving that VAR(1) function can be relocated from the mito chondrion to the nucleus. Moreover, epitope-tagged Var1(U)p co-sedimen ted specifically with small ribosomal subunits in high salt sucrose gr adients. The relocation of VAR1 from the mitochondrion to the nucleus provides an excellent system for the molecular genetic analysis of str ucture-function relationships in the unusual Var1 protein.