title: Sumoylation of Mdm2 by protein inhibitor of activated STAT (PIAS) and RanBP2 enzymes.
abstract: Mdm2, a ubiquitin ligase that acts on p53, is regulated by sumoylation. In the current study, we identify the enzymes responsible for the sumoylation of Mdm2. When mammalian cells are co-transfected with cDNAs encoding Mdm2 and PIAS1 or PIASxbeta (protein inhibitor of activated STAT) as sumoylation enzymes, Mdm2 is highly sumoylated. Mdm2 is also sumoylated in an in vitro system containing PIASxbeta, PIAS1, and RanBP2. When several lysine residues of Mdm2 were sequentially mutated to arginine, the K182R mutant was not sumoylated in intact cells; however, in the in vitro system this mutant was sumoylated by PIAS1, PIASxbeta, and RanBP2 as efficiently as the wild-type Mdm2 protein. Lysine residues 182 and 185 map within the nuclear localization signal of Mdm2. A K185R mutant of Mdm2 is sumoylated in intact cells, whereas a K182R protein is not. Only a Mdm2 protein bearing the K182R mutation is localized exclusively in the cytoplasm. Because RanBP2 is a nuclear pore protein and PIAS proteins are localized within the nucleus, our data suggest that Mdm2 is sumoylated during nuclear translocation by RanBP2 and then further sumoylated once in the nucleus by PIASxbeta and PIAS1.
authors.affiliation: Division of Molecular Life Science, School of Life Science, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Miyauchi
authors.forename: Satomi
authors.initials: S
authors.lastname: Yogosawa
authors.forename: Reiko
authors.initials: R
authors.lastname: Honda
authors.forename: Tamotsu
authors.initials: T
authors.lastname: Nishida
authors.forename: Hideyo
authors.initials: H
authors.lastname: Yasuda
mesh_terms: D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000737:chemistry / Q000378:metabolism; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones; D009154:Mutation; D028861:Nuclear Pore Complex Proteins / Q000378:metabolism*; D009687:Nuclear Proteins; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D050828:Protein Inhibitors of Activated STAT; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051736:Proto-Oncogene Proteins c-mdm2; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D018076:DNA, Complementary; D018832:Molecular Chaperones; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D050828:Protein Inhibitors of Activated STAT; D011506:Proteins; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; C094493:ran-binding protein 2; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D008239:Lysine