MITOCHONDRIAL IMPORT OF A CYTOPLASMIC LYSINE-TRANSFER-RNA IN YEAST ISMEDIATED BY COOPERATION OF CYTOPLASMIC AND MITOCHONDRIAL LYSYL-TRANSFER-RNA SYNTHETASES

Cytoplasmic tRNA(CUU)(Lys) is the only nuclear-encoded tRNA of Sacchar omyces cerevisiae found to be associated with mitochondria. Selective import of this tRNA into isolated organelles requires cytoplasmic fact ors. Here we identify two of these factors as the cytoplasmic and mito chondrial lysyl-tRNA synthetases. The cytoplasmic enzyme is obligatory for in vitro import of the deacylated, but not of the aminoacylated t RNA. We thus infer that it is needed for aminoacylation of the tRNA, w hich is a prerequisite for its import. The mitochondrial synthetase, w hich cannot aminoacylate tRNA(CUU)(Lys), is required for import of bot h aminoacylated and deacylated forms. Its depletion leads to a total a rrest of tRNA import, in vitro and in vivo. The mitochondrial lysyl-tR NA synthetase is able to form specific and stable RNP complexes with t he aminoacylated tRNA. Furthermore, an N-terminal truncated form of th e synthetase which cannot be targeted into mitochondria is unable to d irect the import of the tRNA. We therefore hypothesize that the cytoso lic precursor form of the mitochondrial synthetase has a carrier funct ion for translocation of the tRNA across the mitochondrial membranes. However, cooperation of the two synthetases is not sufficient to direc t tRNA import, suggesting the need of additional factor(s).