Pet111p, an Inner Membrane-bound Translational Activator That Limits Expression of the Saccharomyces cerevisiaeMitochondrial Gene COX2

Abstract

The protein specified by the Saccharomyces cerevisiae nuclear gene PET111 specifically activates translation of the mitochondrially coded mRNA for cytochromec oxidase subunit II (Cox2p). We found Pet111p specifically in mitochondria of both wild-type cells and cells expressing a chromosomal gene for a functional epitope-tagged form of Pet111p. Pet111p was associated with mitochondrial membranes and was highly resistant to extraction with alkaline carbonate. Pet111p was protected from proteolytic digestion by the mitochondrial inner membrane. Thus, it is exposed only on the matrix side, where it could participate directly in organellar translation and localize Cox2p synthesis by virtue of its functional interaction with the COX2 mRNA 5′-untranslated leader. We also found that Pet111p is present at levels limiting the synthesis of Cox2p by examining the effect of alteredPET111 gene dosage in the nucleus on expression of a reporter gene, cox2::ARG8 m, that was inserted into mitochondrial DNA. The level of the reporter protein, Arg8p, was one-half that of wild type in a diploid strain heterozygous for a pet111 deletion mutation, whereas it was increased 2.8-fold in a strain bearing extra copies of PET111 on a high-copy plasmid. Thus, Pet111p could play dual roles in both membrane localization and regulation of Cox2p synthesis within mitochondria. The protein specified by the Saccharomyces cerevisiae nuclear gene PET111 specifically activates translation of the mitochondrially coded mRNA for cytochromec oxidase subunit II (Cox2p). We found Pet111p specifically in mitochondria of both wild-type cells and cells expressing a chromosomal gene for a functional epitope-tagged form of Pet111p. Pet111p was associated with mitochondrial membranes and was highly resistant to extraction with alkaline carbonate. Pet111p was protected from proteolytic digestion by the mitochondrial inner membrane. Thus, it is exposed only on the matrix side, where it could participate directly in organellar translation and localize Cox2p synthesis by virtue of its functional interaction with the COX2 mRNA 5′-untranslated leader. We also found that Pet111p is present at levels limiting the synthesis of Cox2p by examining the effect of alteredPET111 gene dosage in the nucleus on expression of a reporter gene, cox2::ARG8 m, that was inserted into mitochondrial DNA. The level of the reporter protein, Arg8p, was one-half that of wild type in a diploid strain heterozygous for a pet111 deletion mutation, whereas it was increased 2.8-fold in a strain bearing extra copies of PET111 on a high-copy plasmid. Thus, Pet111p could play dual roles in both membrane localization and regulation of Cox2p synthesis within mitochondria. 5′-untranslated leader hemagglutinin The key products of mitochondrial gene expression are integral membrane proteins that are assembled with proteins encoded by nuclear genes to form energy-transducing complexes in the inner membrane (1Tzagoloff A. Myers A.M. Annu. Rev. Biochem. 1986; 55: 249-285Crossref PubMed Scopus (314) Google Scholar, 2Attardi G. Schatz G. Annu. Rev. Cell Biol. 1988; 4: 289-333Crossref PubMed Scopus (1051) Google Scholar, 3Grivell L.A. Artal-Sanz M. Hakkaart G. de Jong L. Nijtmans L.G. van Oosterum K. Siep M. van der Spek H. FEBS Lett. 1999; 452: 57-60Crossref PubMed Scopus (65) Google Scholar). Thus, the activity of mitochondrial genetic systems must be coordinated with cellular gene expression and adapted for efficient delivery of hydrophobic proteins to the inner membrane. Translation of at least five of the seven major Saccharomyces cerevisiae mitochondrially coded mRNAs is under the control of mRNA-specific activator proteins encoded by nuclear genes (4Fox T.D. Hershey J.W.B. Matthews M.B. Sonenberg N. Translational Control. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY1996: 733-758Google Scholar). Genetic analysis has revealed that the activator proteins functionally interact with the 5′-UTLs1 of their target mRNAs and probably mediate productive interactions between those mRNAs and ribosomes (4Fox T.D. Hershey J.W.B. Matthews M.B. Sonenberg N. Translational Control. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY1996: 733-758Google Scholar, 5Green-Willms N.S. Fox T.D. Costanzo M.C. Mol. Cell. Biol. 1998; 18: 1826-1834Crossref PubMed Scopus (50) Google Scholar). As discussed below, several of the activator proteins have been found to be associated with mitochondrial membranes, suggesting that they could function to localize the synthesis of hydrophobic mitochondrial gene products to their sites of insertion in the inner membrane (3Grivell L.A. Artal-Sanz M. Hakkaart G. de Jong L. Nijtmans L.G. van Oosterum K. Siep M. van der Spek H. FEBS Lett. 1999; 452: 57-60Crossref PubMed Scopus (65) Google Scholar, 4Fox T.D. Hershey J.W.B. Matthews M.B. Sonenberg N. Translational Control. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY1996: 733-758Google Scholar, 6Michaelis U. Körte A. Rödel G. Mol. Gen. Genet. 1991; 230: 177-185Crossref PubMed Scopus (53) Google Scholar, 7Costanzo M.C. Fox T.D. Annu. Rev. Genet. 1990; 24: 91-113Crossref PubMed Google Scholar). This hypothesis is strongly supported by the fact that the untranslated regions of the COX2 and COX3 mRNAs contain information that facilitates correct targeting of the Cox2p and Cox3p cytochrome oxidase subunits (8Sanchirico M.E. Fox T.D. Mason T.L. EMBO J. 1998; 17: 5796-5804Crossref PubMed Scopus (101) Google Scholar). The translational activator proteins themselves are present at low levels (9Fox T.D. Experientia. 1996; 52: 1130-1135Crossref PubMed Scopus (65) Google Scholar) and have therefore been difficult to study. In no case have they been shown to be localized within mitochondria isolated from wild-type (nonoverproducing) cells such that they could participate directly in organellar protein synthesis. Cbs1p and Cbs2p, activators required for translation of the cytochrome b mRNA (10Rödel G. Curr. Genet. 1986; 11: 41-45Crossref PubMed Scopus (76) Google Scholar), were detectable at normal levels in wild-type cells. They were found associated with the total mitochondrial membrane fraction. Cbs1p behaved like an integral membrane protein, whereas Cbs2p was peripherally associated, but they were not further localized within the organelle (6Michaelis U. Körte A. Rödel G. Mol. Gen. Genet. 1991; 230: 177-185Crossref PubMed Scopus (53) Google Scholar). Pet309p, required for Cox1p translation (11Manthey G.M. McEwen J.E. EMBO J. 1995; 14: 4031-4043Crossref PubMed Scopus (179) Google Scholar), could only be detected as an epitope-tagged species in cells overproducing the protein (12Manthey G.M. Przybyla-Zawislak B.D. McEwen J.E. Eur. J. Biochem. 1998; 255: 156-161Crossref PubMed Scopus (62) Google Scholar). Under these conditions it fractionated as an integral inner membrane protein, partially exposed on the outer surface (intermembrane space side). Mss51p, also required for Cox1p synthesis, was peripherally associated with mitoplast (mitochondria with ruptured outer membranes) membranes when expressed from a low-copy plasmid (13Siep M. van Oosterum K. Neufeglise H. van der Spek H. Grivell L.A. Curr. Genet. 2000; 37: 213-220Crossref PubMed Scopus (34) Google Scholar). Synthesis of Cox3p requires three interacting proteins, Pet54p, Pet122p, and Pet494p (14Costanzo M.C. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1988; 85: 2677-2681Crossref PubMed Scopus (87) Google Scholar, 15Brown N.G. Costanzo M.C. Fox T.D. Mol. Cell. Biol. 1994; 14: 1045-1053Crossref PubMed Scopus (68) Google Scholar). Only Pet54p could be detected immunologically in wild-type cells and was found to be peripherally associated with the inner membrane (16McMullin T.W. Fox T.D. J. Biol. Chem. 1993; 268: 11737-11741Abstract Full Text PDF PubMed Google Scholar). Pet122p and Pet494p, when overproduced, fractionated as integral inner membrane proteins (16McMullin T.W. Fox T.D. J. Biol. Chem. 1993; 268: 11737-11741Abstract Full Text PDF PubMed Google Scholar). The mRNA-specific activators could regulate expression of mitochondrial genes in addition to targeting synthesis on the membrane if they were present at levels limiting translation. Alternatively, the activators could be present in excess, with regulation of mitochondrial gene expression controlled at some other step. This has been examined to date only in the case of COX3 expression using the synthetic reporter gene ARG8 m inserted into mtDNA at COX3 (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar). Reduction of the gene dosage ofPET494 caused reduced expression of the mitochondrial reporter, although increased dosage of PET494 caused only a modest increase in reporter expression. In this study we have focussed on Pet111p, the only known translational activator for the COX2 mRNA (18Poutre C.G. Fox T.D. Genetics. 1987; 115: 637-647Crossref PubMed Google Scholar, 19Mulero J.J. Fox T.D. Genetics. 1993; 133: 509-516Crossref PubMed Google Scholar). Pet111p is known to be mitochondrially located (20Strick C.A. Fox T.D. Mol. Cell. Biol. 1987; 7: 2728-2734Crossref PubMed Scopus (31) Google Scholar) and to interact functionally with the COX2 mRNA 5′-UTL (21Mulero J.J. Fox T.D. Mol. Biol. Cell. 1993; 4: 1327-1335Crossref PubMed Scopus (69) Google Scholar). Although the amino acid sequence of Pet111p is not highly conserved among budding yeasts, its function in specifically activating translation of the COX2mRNA has been conserved, indicating an important role in the production of cytochrome c oxidase (22Costanzo M.C. Bonnefoy N. Williams E.H. Clark-Walker G.D. Fox T.D. Genetics. 2000; 154: 999-1012PubMed Google Scholar). Here we study the submitochondrial localization of Pet111p in cells expressing thePET111 gene at normal levels and examine the effect of altered PET111 gene dosage on expression of the mitochondrial COX2 gene. Our results are consistent with a direct role for Pet111p in localization of mitochondrial translation and demonstrate that it is a rate-limiting factor in the expression of the COX2 gene. S. cerevisiaestrains used in this study are listed in TableI. All strains are congenic to the wild-type strain D273–10B (ATCC 25657) except DFS160rho0, HMD122, and NSG175, which were derived from DBY947 (23Neff N.F. Thomas J.H. Grisafi P. Botstein D. Cell. 1983; 33: 211-219Abstract Full Text PDF PubMed Scopus (231) Google Scholar). Cells were grown in rich medium (1% yeast extract, 2% bacto-peptone) containing 2% galactose or 2% raffinose or in minimal medium (0.67% yeast nitrogen base) containing 2% raffinose, as indicated in the figure legends. Standard genetic methods were as described (24Sherman F. Fink G.R. Hicks J.B. Methods in Yeast Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY1986Google Scholar, 25Fox T.D. Folley L.S. Mulero J.J. McMullin T.W. Thorsness P.E. Hedin L.O. Costanzo M.C. Methods Enzymol. 1991; 194: 149-165Crossref PubMed Scopus (204) Google Scholar). Strains deleted forPET111 carry either the pet111–14 deletion (26Mulero J.J. Fox T.D. Mol. Gen. Genet. 1994; 242: 383-390Crossref PubMed Scopus (39) Google Scholar) or the pet111–9 deletion with LEU2 in its place (27Strick C.A. A Study of the Expression and Protein Product of the Yeast Positive Regulatory Gene PET 111.Ph.D. thesis. Cornell University, 1988Google Scholar).Table IYeast strains used in this studyStrainGenotypeSourcePTY11MATα ura3–52 trp1-Δ1 [ρ+]P. E. ThorsnessNB39–16DMATα ura3 trp1 pet111–9[ρ+]N. BonnefoyNSG91MATα ura3–52 trp1-Δ1 PET111-HA-TRP1 [ρ+]This studyHMD22MATalys2 leu2–3,112 ura3–52 his3 arg8::hisG [ρ+ cox2::ARG8 m]This studyNB40–20DMATα lys2 arg8::hisG[ρ+]N. BonnefoyGW241rho0MATaade2 ura3–52 leu2–3,112 arg8::hisG[ρ0]G. WiesenbergerNSG176MATaura3–52 leu2–3,112 arg8::his ade2 [ρ+ cox2::ARG8 m]This studyNSG170rho0MATα ade2 lys2 arg8::hisG[ρ0]This studyNSG165rho0MATα ade2 lys2 ura3 arg8::URA3 pet111–14 [ρ0]This studyNSG164rho0MATα lys2 leu2–3,112 arg8::hisG pet494–41 [ρ0]This studyNSG171rho0MATα ade2 lys2 ura3 arg8::hisG[ρ0]This studyDFS160rho0MATα ade2–101 leu2Δ ura3–52 arg8Δ::URA3 kar1–1[ρ0](17)HMD122MATα ade2–101 leu2Δ ura3–52 arg8Δ::URA3 kar1–1 [ρ− cox2::ARG8 m]This studyNSG175MATα ade2–101 ura3–52 kar1–1[ρ+ cox2::ARG8 m]This study Open table in a new tab The PET111 reading frame was modified to encode three copies of the influenza virus HA epitope (28Tyers M. Tokiwa G. Nash R. Futcher B. EMBO J. 1992; 11: 1773-1784Crossref PubMed Scopus (338) Google Scholar) at its 3′ end using the plasmid pCS124 (obtained from C. Shamu and J. Nunnari), an integrative plasmid carrying three copies of the HA sequence and the TRP1 gene. The downstream 556 base pairs of PET111 were inserted as anEcoRI-NcoI fragment into pCS124 such that the epitope-coding sequence was in-frame with PET111. The resulting plasmid, pNSG30, was cut at a unique within PET111 and used to strain to In the resulting integrative the only of PET111 was the was by were from cells grown to in medium containing 2% galactose as described L.A. Methods Enzymol. 1995; PubMed Scopus Google Scholar), except that were using a as described E. S. A. J. Biol. Chem. 1988; Full Text PDF PubMed Google Scholar). mitochondria were by on as described L.A. Methods Enzymol. 1995; PubMed Scopus Google Scholar). were into membrane and as described Methods Enzymol. 1995; PubMed Scopus Google Scholar), and alkaline of membranes in were as described S. J. Cell Biol. 93: PubMed Scopus Google Scholar). were in acid were to by into containing either or and on for Methods Enzymol. 1995; PubMed Scopus Google Scholar, A. K. S. Schatz G. Cell. 1992; Full Text PDF PubMed Scopus Google Scholar). mitochondria were into m containing either or and were to on and as described E. D. a Cold Spring Harbor Laboratory, Cold Spring Harbor, Scholar). The acid fragment of Pet111p was isolated as a protein encoded by J.J. on the Translation of the Yeast COX2 mRNA with on Translational thesis. Cornell University, Scholar). were isolated in with and the protein was by were and as described E. D. a Cold Spring Harbor Laboratory, Cold Spring Harbor, Scholar). was with from a strain E. D. a Cold Spring Harbor Laboratory, Cold Spring Harbor, Scholar), on a and used at a The the HA epitope was from and was used at was a from L. and were from B. and G. of (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar) and P.E. Fox T.D. Mol. Cell. Biol. 1993; PubMed Scopus Google Scholar) was described the of complexes were on using or and the is to mitochondrial were from cells with the gene grown to in galactose medium and of total mitochondrial protein and submitochondrial were by were into and membrane were further with alkaline to membrane proteins from integral membrane proteins The were with to as as inner Cox2p and the matrix by the wild-type gene in this is an inner membrane protein the were from strain and to by in the or of as indicated for were also with in the of the to were by with to as as the matrix the inner protein exposed on the outer and the space protein cytochrome cells were at and total protein was isolated as described Methods Enzymol. 1991; 194: PubMed Scopus (150) Google Scholar). were to in and to were in in containing in and with the in containing either at for or at The were in and with in containing for at The was for and for The membranes were in and to at for at least on a were with (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar) that been on a and with E. D. a Cold Spring Harbor Laboratory, Cold Spring Harbor, Scholar) derived from yeast were with 2% at for with at and to for of the were with were using from the of the of was for the was to the The plasmid was in the the ARG8 m sequence (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar) by of mitochondrial of the COX2 reading frame and of mitochondrial downstream of the The sequence downstream of m an for In a inserted between the and of the to as an mitochondrial genetic The sequence of is was into the mitochondria of strain (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar) by as described G. Costanzo M.C. Fox T.D. Mol. Cell. Biol. 1995; PubMed Scopus (39) Google Scholar, and D. Methods Cell in Scholar), and mitochondrial were by their to the M.C. Fox T.D. Mol. Cell. Biol. 1993; PubMed Scopus Google Scholar). such HMD122, was isolated and used to into mitochondria by to strains and Pet111p is expressed at low levels (9Fox T.D. Experientia. 1996; 52: 1130-1135Crossref PubMed Scopus (65) Google Scholar), and we have not been to the protein in wild-type cells (21Mulero J.J. Fox T.D. Mol. Biol. Cell. 1993; 4: 1327-1335Crossref PubMed Scopus (69) Google Scholar). Yeast cells overproducing Pet111p it in associated with A. and D. localization of a protein be from analysis of cells overproducing We therefore to the of Pet111p A new was a protein bearing the Pet111p amino acid and expressed in E. This was used to from a wild-type strain on a with a protein of the for Pet111p that was highly in mitochondria and from This protein was not detected in isolated from a pet111 deletion in wild-type with the level of this protein was increased to total mitochondrial protein in the wild-type strain containing the plasmid which a fragment the PET111 and (26Mulero J.J. Fox T.D. Mol. Gen. Genet. 1994; 242: 383-390Crossref PubMed Scopus (39) Google Scholar) species of which Pet111p were also detected In we were to Pet111p as a protein bearing three copies of the HA expressed from its chromosomal strains were that is a functional were from a strain expressing and from a wild-type strain and were with an and with The with a protein of the in the mitochondria but not in the wild-type mitochondria. The with the protein in both that the species in is Pet111p. the of Pet111p within the submitochondrial were from a strain expressing mitochondria were by and fractionated by to and The membrane was with alkaline to membrane proteins from integral membrane The resulting were by with to the of of the was in the membrane of the with the integral membrane protein alkaline indicating that Pet111p is The known integral membrane protein Cox2p was associated with membranes in this whereas the matrix protein was were used to the submitochondrial of of mitochondria by detectable that the epitope is not protected by a protein in both mitochondria and in to the outer membrane was protected from digestion by K. Thus, the epitope to be on the of the inner membrane. no species were it that the protein is protected from by the inner membrane. The was for the matrix is an integral inner membrane protein exposed on the outer surface of the inner membrane K. G. EMBO J. 1996; PubMed Scopus Google Scholar) and is not protected from by as these that Pet111p is associated with the inner surface of the inner mitochondrial membrane. Expression of the gene could be at the level of translation by the activity of Pet111p. examine this we m reporter gene that the of ARG8 m to translation and This reporter was inserted into mitochondrial reading frame with ARG8 The resulting gene an mRNA with the Pet111p target in the COX2 of by Expression of this mitochondrial reporter gene the caused by a nuclear This was by a pet111 nuclear mutation, that expression is As the of immunologically detectable encoded by the mitochondrial reporter the level m mRNA translation was by the level of Pet111p, we strains containing the mitochondrial reporter with of thePET111 nuclear gene. We the levels of in by of were controlled by to the level of the Expression of the gene to be under conditions PubMed Scopus Google Scholar, M. 1990; PubMed Scopus Google Scholar). the dosage of PET111 was by a diploid strain heterozygous for a pet111 The level of reporter protein in this strain was that of a control strain for wild-type copies of as be if Pet111p levels COX2 mRNA translation As a control for mRNA we also examined the effect m expression of the gene dosage of which a subunit of mRNA-specific translational activator N.G. Costanzo M.C. Fox T.D. Mol. Cell. Biol. 1994; 14: 1045-1053Crossref PubMed Scopus (68) Google Scholar) and is limiting for m expression (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar). levels were not in heterozygous diploid to the wild-type strain but to be increased We examined the effect of PET111 gene dosage on the expression of the cox2::ARG8 gene. The PET111 plasmid (26Mulero J.J. Fox T.D. Mol. Gen. Genet. 1994; 242: 383-390Crossref PubMed Scopus (39) Google Scholar) was into a wild-type diploid The level of present in this was to that of the strain with the J.E. Myers A.M. A. 1986; PubMed Scopus Google Scholar). The of extra copies of PET111 caused the level of to increase by that Pet111p activity is a limiting factor in the expression of the mitochondrial S. cerevisiae Pet111p is in the as an acid protein, which be into mitochondria. is and has a genetic have for a functional interaction between Pet111p and the COX2 mRNA in the mitochondrial where translation we have detected Pet111p in mitochondria of cells expressing it at normal levels and that it is associated with the inner membrane. Pet111p is protected by the inner membrane from indicating that its must be exposed on the matrix where they could interact with the COX2 mRNA and of the organellar translation Thus, Pet111p is to be the membrane that targeting information in the untranslated of the COX2 efficient cytochrome c oxidase (8Sanchirico M.E. Fox T.D. Mason T.L. EMBO J. 1998; 17: 5796-5804Crossref PubMed Scopus (101) Google Scholar). Pet111p was resistant to from mitochondrial membranes by extraction with alkaline suggesting that it is an integral membrane protein S. J. Cell Biol. 93: PubMed Scopus Google Scholar). it was not as as the known integral membrane protein Although analysis of cerevisiae Pet111p sequence using the and the with the of Pet111p from three budding (22Costanzo M.C. Bonnefoy N. Williams E.H. Clark-Walker G.D. Fox T.D. Genetics. 2000; 154: 999-1012PubMed Google Scholar) not such B. P. Protein Sci. 1996; PubMed Scopus Google Scholar, B. P. J. Mol. Biol. 1994; PubMed Scopus Google Scholar). The Pet122p subunit of the activator as an integral membrane protein both when (16McMullin T.W. Fox T.D. J. Biol. Chem. 1993; 268: 11737-11741Abstract Full Text PDF PubMed Google Scholar) and when expressed from a chromosomal gene. A. Butler and D. Pet122p also and exposed to on the outer surface of the inner This with that of Pet309p, the mRNA-specific that the inner membrane and is to from the (12Manthey G.M. Przybyla-Zawislak B.D. McEwen J.E. Eur. J. Biochem. 1998; 255: 156-161Crossref PubMed Scopus (62) Google Scholar). The of Pet111p to expression of the mitochondrial gene COX2 at the level of the level of encoded by a cox2::ARG8 gene with PET111 gene The of was to PET111 dosage when we reduced the of PET111 genes by The level increased 2.8-fold when PET111 was into cells on a that caused an increase in the level of Pet111p. This is consistent with a that the COX2 mRNA level is not limiting for gene expression of that mRNA not to increased synthesis of Cox2p A.M. Mason T.L. Mol. Cell. Biol. 1994; 14: PubMed Scopus (34) Google Scholar). We that of the level of Pet111p its translational activity could regulate the level of Cox2p synthesis within mitochondria. activity of a limiting translational activator increase translation of the target mitochondrial mRNA to the that some other We be this was in we not it the increase in Pet111p levels in mitochondria from cells with dosage was to the 2.8-fold increase in reporter gene expression. In a study m expression we found that of the limiting activator Pet494p, increased the mitochondrial reporter expression only by (17Steele D.F. Butler C.A. Fox T.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 5253-5257Crossref PubMed Scopus (150) Google Scholar). The of between translational activators for low levels of some translation is by the increase m expression in a diploid heterozygous for a but this effect not be The PET111 mRNA has an 5′-UTL of that reading of the sequence (20Strick C.A. Fox T.D. Mol. Cell. Biol. 1987; 7: 2728-2734Crossref PubMed Scopus (31) Google Scholar), suggesting the expression be controlled at the level of translation The and of the Yeast Protein and Cold Spring Harbor Laboratory, Cold Spring Harbor, Scholar). an mRNA could either as a target for regulation that gene expression of mRNA levels or it could the translational of the PET111 The fact that increased of the PET111 gene an increase in Pet111p levels and activity Pet111p synthesis is not of the mRNA we have not the protein and mRNA levels Translational that synthesis of to the of a membrane in K. R. J. Proc. Natl. Acad. Sci. U. S. A. 1998; PubMed Scopus Google Scholar, L. R. 1999; PubMed Scopus Google Scholar) and of the of A. J. PubMed Google Scholar) have been Our results that Pet111p is to play roles in both the of Cox2p synthesis and that synthesis on the surface of the inner membrane. Thus, Pet111p could participate in a Cox2p translation with its into the cytochrome c oxidase We L. B. and G. Schatz for of