Functional Interactions between Yeast Mitochondrial Ribosomes and mRNA 5′ Untranslated Leaders

While mitochondrial ribosomes exhibit distinct similarities to bacterial (eubacterial) ribosomes (30, 69), the yeast mitochondrial translation system has many intriguing differences from bacterial systems. Mitochondrial ribosomes have more proteins than do bacterial ribosomes (23, 43). Of the mitochondrial ribosomal proteins whose sequences are known, some are simple homologs of their bacterial counterparts, others have domains homologous to bacterial ribosomal proteins attached to domains with no recognizable homology to any known proteins, and still others are completely unrelated to bacterial ribosomal proteins (reviewed in reference 32). Saccharomyces cerevisiae mitochondrial mRNAs generally have long, A+U-rich 5′ untranslated leaders (5′-UTLs) lacking a typical Shine-Dalgarno sequence (11, 27, 31). While the mechanism of start site selection remains obscure in this system, translation initiation on most or all yeast mitochondrial mRNAs requires membrane-bound mRNA-specific activator proteins whose targets lie in the 5′-UTLs (reviewed in reference 27). These mRNA-specific activators appear to play a dual role in mitochondrial gene expression: tethering the synthesis of the very hydrophobic mitochondrial gene products to the inner membrane (27) and modulating the translation levels of individual mRNAs (63). We have focused on translation of the COX2 and COX3 mRNAs, which encode subunits II and III of cytochrome c oxidase, respectively. Previous studies have identified their mRNA-specific translational activators and established functional interactions among activator proteins, their mRNA targets, and other components of the mitochondrial translation system. The COX2-specific translational activator protein is encoded by the nuclear gene PET111 (46, 54), while the COX3-specific activator is a complex containing three proteins encoded by the nuclear genes PET54, PET122, and PET494 (6, 9, 14, 38). Using suppressor analysis, we have shown that one subunit of the COX3-specific activator, Pet122p, interacts functionally with the small subunit of mitochondrial ribosomes (33, 35, 42) and that each translational activator interacts functionally with the 5′-UTL of its target mRNA (12, 45, 67). These findings suggested that yeast mitochondrial ribosomes were unable to recognize mRNAs unless the ribosome-mRNA interaction was mediated by translational activators that recognized sites unique to each mRNA. Here we report that certain mutations in the COX2 and COX3 mRNA 5′-UTLs that are suppressible by alterations of mRNA-specific activators can also be suppressed by mutations in nuclear genes encoding two mitochondrial ribosomal small-subunit proteins. The suppression is allele specific, indicating that the ribosomes play an active role in the recognition of translation start signals. Surprisingly, however, suppression is not gene specific, indicating that the ribosomes are recognizing features of the 5′-UTLs that are common to at least several mRNAs. One of these yeast mitochondrial ribosomal proteins is homologous to bacterial S21 and to the products of unidentified genes from several animals.