Cloning and developmental expression of a nuclear ubiquitin-conjugating enzyme (DmUbc9) that interacts with small heat shock proteins in Drosophila melanogaster.
In a two hybrid screen designed to identify proteins that interact with small heat shock proteins (sHsps), a Drosophila melanogaster homologue of yeast and human ubc9 (Dmubc9) was found to interact with Drosophila Hsp23. Further, two-hybrid system analysis reveals DmUbc9 interaction with Drosophila and mammalian Hsp27. In situ hybridization localizes Dmubc9 as a doublet at locus 21D on chromosome 2L, and genomic cloning of the gene reveals a single open reading frame without introns. The predicted Dmubc9 protein sequence shares a very high level of homology with mouse (85.4%) and human (> or = 82.9%) Ubc9. Genetic complementation analysis show that Dmubc9 functionally rescues a temperature-sensitive S. cerevisiae ubc9ts mutant. Co-immunoprecipitation with antibody raised against DmUbc9 confirms the interaction with Drosophila Hsp23 and Hsp26 and preferentially with Hsp27. The DmUbc9 protein, which localizes primarily to the nucleus in Drosophila S2 cells, is found at high levels in embryos but is also present at lower levels throughout development. The significance of the sHsp-Ubc9 interaction is discussed.
Joanisse
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DR
Laboratoire de Génétique Cellulaire et Développementale, Pavillon C-E. Marchand, Université Laval, Sainte-Foy, Québec, Canada.
Inaguma
Y
Y
Tanguay
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RM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000201:enzymology*; D003001:Cloning, Molecular; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D008025:Ligases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D014407:Tumor Cells, Cultured; D044763:Ubiquitin-Conjugating Enzymes; D014452:Ubiquitins / Q000378:metabolism*; D000098843:Ubiquitin-Conjugating Enzyme UBC9
D005656:Fungal Proteins; D006360:Heat-Shock Proteins; D014452:Ubiquitins; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases; D000098843:Ubiquitin-Conjugating Enzyme UBC9