title: Alternative translation initiation augments the human mitochondrial proteome.
abstract: Alternative translation initiation (ATI) is a mechanism of producing multiple proteins from a single transcript, which in some cases regulates trafficking of proteins to different cellular compartments, including mitochondria. Application of a genome-wide computational screen predicts a cryptic mitochondrial targeting signal for 126 proteins in mouse and man that is revealed when an AUG codon located downstream from the canonical initiator methionine codon is used as a translation start site, which we term downstream ATI (dATI). Experimental evidence in support of dATI is provided by immunoblotting of endogenous truncated proteins enriched in mitochondrial cell fractions or of co-localization with mitochondria using immunocytochemistry. More detailed cellular localization studies establish mitochondrial targeting of a member of the cytosolic poly(A) binding protein family, PABPC5, and of the RNA/DNA helicase PIF1α. The mitochondrial isoform of PABPC5 co-immunoprecipitates with the mitochondrial poly(A) polymerase, and is markedly reduced in abundance when mitochondrial DNA and RNA are depleted, suggesting it plays a role in RNA metabolism in the organelle. Like PABPC5 and PIF1α, most of the candidates identified by the screen are not currently annotated as mitochondrial proteins, and so dATI expands the human mitochondrial proteome.
authors.affiliation: MRC-Mitochondrial Biology Unit, Wellcome Trust-MRC Building, Cambridge CB2 0XY, UK.
authors.forename: Lawrence
authors.initials: L
authors.lastname: Kazak
authors.forename: Aurelio
authors.initials: A
authors.lastname: Reyes
authors.forename: Anna L
authors.initials: AL
authors.lastname: Duncan
authors.forename: Joanna
authors.initials: J
authors.lastname: Rorbach
authors.forename: Stuart R
authors.initials: SR
authors.lastname: Wood
authors.forename: Gloria
authors.initials: G
authors.lastname: Brea-Calvo
authors.forename: Payam A
authors.initials: PA
authors.lastname: Gammage
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Robinson
authors.forename: Michal
authors.initials: M
authors.lastname: Minczuk
authors.forename: Ian J
authors.initials: IJ
authors.lastname: Holt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D018387:Codon, Initiator; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000302:isolation & purification; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000201:enzymology / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000032:analysis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010442:Peptide Chain Initiation, Translational; D039102:Poly(A)-Binding Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011062:Polynucleotide Adenylyltransferase / Q000302:isolation & purification; D020033:Protein Isoforms / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D020543:Proteome / Q000235:genetics* / Q000378:metabolism
chemical_list: D018387:Codon, Initiator; D004272:DNA, Mitochondrial; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; C580881:PABPC5 protein, human; D039102:Poly(A)-Binding Proteins; D020033:Protein Isoforms; D020543:Proteome; C504091:NABP2 protein, human; D011062:Polynucleotide Adenylyltransferase; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; D004265:DNA Helicases; C508765:PIF1 protein, human; C517519:Pif1 protein, mouse