title: Detection of O-propargyl-puromycin with SUMO and ubiquitin by click chemistry at PML-nuclear bodies during abortive proteasome activities.
abstract: The amino-nucleoside antibiotic, puromycin, acts by covalently linking to elongating polypeptide chains on ribosomes to generate prematurely terminated immature polypeptides. The trafficking of puromycin-conjugated (puromycylated) immature polypeptides within cell has, however, remained elusive. In this study, using O-propargyl-puromycin (OP-Puro), the distribution of puromycylated polypeptides was assessed in HeLa cells by click chemistry. Under standard culture conditions, OP-Puro signals were detected in the cytoplasm and nucleus with the highest concentrations in the nucleolus. Intriguingly, when proteasome activities were aborted using MG132, OP-Puro signals began to accumulate at promyelocytic leukemia nuclear bodies (PML-NBs) in addition to the nucleolus. We also found promiscuous association of OP-Puro signals with SUMO-2/3 and ubiquitin at PML-NBs, but not at the nucleolus, during abortive proteasome activities. This study reveals a previously unknown distribution of OP-Puro that argues for a nuclear function in regulating immature protein homeostasis.
authors.affiliation: Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Uozumi
authors.affiliation: Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto, Japan.
authors.forename: Hotaru
authors.initials: H
authors.lastname: Matsumoto
authors.affiliation: Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto, Japan; Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, Kumamoto, Japan. Electronic address: hisa@kumamoto-u.ac.jp.
authors.forename: Hisato
authors.initials: H
authors.lastname: Saitoh
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism*; D057930:Click Chemistry / Q000379:methods; D045586:Intranuclear Inclusion Bodies / Q000378:metabolism*; D000071156:Promyelocytic Leukemia Protein / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011691:Puromycin / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D025842:SUMO-1 Protein / Q000737:chemistry / Q000378:metabolism*; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism*
chemical_list: C570448:O-propargyl-puromycin; D000071156:Promyelocytic Leukemia Protein; D025842:SUMO-1 Protein; D025801:Ubiquitin; D011691:Puromycin; D046988:Proteasome Endopeptidase Complex
keywords: Click chemistry; Promyelocytic leukemia nuclear bodies; Protein homeostasis; Puromycin; Small ubiquitin-related modifier (SUMO); Ubiquitin