title: DNA Damage-Induced Foci of E2 Ubiquitin-Conjugating Enzyme are Detectable upon Co-transfection with an Interacting E3 Ubiquitin Ligase.
abstract: DNA damage repair elements accumulate at DNA damage sites to form ionizing radiation-induced foci (IRIF) for damage repair. IRIF, which represent direct evidence of DNA damage response activity, which are conveniently to be observed via immunofluorescence staining. Protein ubiquitination plays an important role in initiating the DNA damage response. Following DNA damage, the substrate binding protein E3 ubiquitin-ligases enzymes are recruited to DNA damage sites, then the E2 ubiquitin-conjugating enzymes are recruited to these sites by the E3 where they catalyze protein ubiquitination. However, IRIF of E2 enzymes are relatively transient and unstable in vivo and difficult to detect. Here, we present a new method for the observation of E2 IRIF. This method is based on the co-transfection of interacting E2 and E3 enzymes into cells and identifies IRIF via immunofluorescence following DNA damage.
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China.
authors.forename: Degui
authors.initials: D
authors.lastname: Wang
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China. tianyxlz@163.com.|Department of Internal Medicine, Gansu Provincial Academic Institute for Medical Research, Lanzhou, 730050, China. tianyxlz@163.com.
authors.forename: Yingxia
authors.identifier: http://orcid.org/0000-0003-0693-3031
authors.initials: Y
authors.lastname: Tian
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China.
authors.forename: Dong
authors.initials: D
authors.lastname: Wei
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China.
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Jing
authors.affiliation: Department of Urology, The Affiliated Hospital of Qingdao University, Qingdao, China. Niuht0532@126.com.
authors.forename: Haitao
authors.initials: H
authors.lastname: Niu
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Xie
authors.affiliation: Department of Anatomy and Histology, School of Basic Medical Sciences, Lanzhou University, Lanzhou, 730000, China. songyanfeng@lzu.edu.cn.
authors.forename: Yanfeng
authors.initials: Y
authors.lastname: Song
mesh_terms: D000818:Animals; D004249:DNA Damage / Q000235:genetics*; D004260:DNA Repair; D019085:Fluorescent Antibody Technique, Direct; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D025801:Ubiquitin; C471545:UBE2A protein, human; C471546:UBE2B protein, human; D044763:Ubiquitin-Conjugating Enzymes; C535210:RNF168 protein, human; D044767:Ubiquitin-Protein Ligases
keywords: DNA damage; Foci; Ubiquitin; Ubiquitin-conjugating enzyme