DELETION OF THE LEADER PEPTIDE OF THE MITOCHONDRIALLY ENCODED PRECURSOR OF SACCHAROMYCES-CEREVISIAE CYTOCHROME-C-OXIDASE SUBUNIT-II

Cytochrome c oxidase subunit II (CoxPp) of Saccharomyces cerevisiae is synthesized within mitochondria as a precursor, pre-Cox2p. The 15-ami no acid leader peptide is processed after export to the intermembrane space. Leader peptides are relatively unusual in mitochondrially coded proteins: indeed mammalian CoxPp lacks a leader peptide. We generated two deletions in the S. cerevisiae COX2 gene, removing either the lea der peptide (cox2-20) or the leader peptide and processing site (cox2- 21) without altering either the promoter or the mRNa-specific translat ional activation site. When inserted into mtDNA, both deletions substa ntially reduced the steady-state levels of CoxPp and caused a tight no nrespiratory phenotype. A respiring pseudorevertant of the cox2-20 mut ant was heteroplasmic for the original mutant mtDNA and a p(-) mtDNA w hose deletion fused the first 251 codons of the mitochondrial gene enc oding cytochrome b to the cox2-20 sequence. The resulting fusion prote in was processed to yield functional Cox2p. Thus, the presence of amin o-terminal cytochrome b sequence bypassed the need for the pre CoxPp l eader peptide. We propose that the pre-Cox2p leader peptide contains a targeting signal necessary for membrane insertion, without which it r emains in the matrix and is rapidly degraded.