title: Quantitative and subcellular localization analysis of the nuclear isoform dUTP pyrophosphatase in alkylating agent-induced cell responses.
abstract: Our previous proteome analysis showed that the nuclear isoform of dUTP pyrophosphatase (DUT-N) was identified in the culture medium of human amnion FL cells after exposure to the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). These results suggest that DUT-N may be a potential early biomarker to assess the risk of alkylating agents exposure. DUT-N is one of the two isoforms of deoxyuridine triphosphate nucleotidohydrolase (dUTPase). Our current knowledge of DUT-N expression in human cells is very limited. In the current study, we first investigated the appearance of DUT-N in the culture medium of different human cell lines in response to a low concentration of MNNG exposure. We verified that the MNNG-induced appearance of DUT-N in the extracellular environment is cell-specific. Western blot analysis confirmed that the intracellular DUT-N changes responded to MNNG in a concentration-dependent and cell-specific manner. Furthermore, subcellular fraction experiments showed that 0.25μM MNNG treatment dramatically increased the DUT-N expression levels in the cytoplasmic extracts prepared from both FL and HepG2 cells, increased DUT-N levels in nuclear extracts prepared from HepG2 cells, and decreased DUT-N levels in nuclear extracts from FL cells. Morphological studies using immunofluorescence showed that a low concentration of MNNG could alter the distribution of DUT-N in FL and HepG2 cells in different ways. Taken together, these studies indicate a role of DUT-N in alkylating agent-induced cell responses.
authors.affiliation: Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, 388 Yuhangtang Road, Hangzhou 310058, China. huxiaolan1998@yahoo.com.cn
authors.forename: Xiaolan
authors.initials: X
authors.lastname: Hu
authors.forename: Yingnian
authors.initials: Y
authors.lastname: Yu
authors.forename: Qian
authors.initials: Q
authors.lastname: Li
authors.forename: Danxiao
authors.initials: D
authors.lastname: Wu
authors.forename: Zhengning
authors.initials: Z
authors.lastname: Tan
authors.forename: Cheng
authors.initials: C
authors.lastname: Wang
authors.forename: Jvping
authors.initials: J
authors.lastname: Wang
authors.forename: Meiping
authors.initials: M
authors.lastname: Wu
mesh_terms: D000477:Alkylating Agents / Q000633:toxicity*; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D003593:Cytoplasm / Q000201:enzymology; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D006801:Humans; D008769:Methylnitronitrosoguanidine / Q000633:toxicity*; D011755:Pyrophosphatases / Q000032:analysis / Q000502:physiology*
chemical_list: D000477:Alkylating Agents; D008769:Methylnitronitrosoguanidine; D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase