title: Increased expression of a COOH-truncated nucleophosmin resulting from alternative splicing is associated with cellular resistance to ionizing radiation in HeLa cells.
abstract: We previously demonstrated that transfecting HeLa cells with the 24 kDa basic fibroblast growth factor-2 (FGF-2) isoform dramatically increased cell survival after irradiation. To investigate genes implicated in this radioresistance acquisition, we compared mRNA expression between radioresistant 24 kDa FGF-2-expressing cells (HeLa 3A) and radiosensitive control HeLa PINA cells using the differential display technique. Of the 32 differentially expressed mRNAs, 1 presented a significant homology with a known gene. This 378 bp fragment presented 100% identity with exon 11 and 12 of human nucleophosmin (NPM) but differed by including a part of intron 9 in its 5' end. The differential expression of this fragment was confirmed using an RNase protection assay. We then cloned the entire corresponding mRNA and showed that it contained all the exons of NPM plus intron 9, suggesting that it was a splicing product of the NPM gene. This variant encoded for a 35-amino acid truncated NPM (NPM2). NPM2 expression was increased in HeLa 3A. To investigate NPM2's role in radioresistance acquisition, we transfected HeLa cells with NPM2 cDNA and analyzed survival after irradiation of the clones obtained. After transfection with NPM2, radiosensitive HeLa cells exhibited a dramatic increase in cell survival after irradiation. Taken together, our results demonstrate that expression of a COOH-truncated NPM form resulting from the alternative splicing of NPM mRNA is able to increase cell survival after irradiation and suggests that it might be involved in cellular response to ionizing radiation.
authors.affiliation: Laboratoire d'Oncologie Cellulaire et Moléculaire, UPRES-EA 2048, Université Paul Sabatier, Toulouse, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Dalenc
authors.forename: Jerome
authors.initials: J
authors.lastname: Drouet
authors.forename: Isabelle
authors.initials: I
authors.lastname: Ader
authors.forename: Caroline
authors.initials: C
authors.lastname: Delmas
authors.forename: Philippe
authors.initials: P
authors.lastname: Rochaix
authors.forename: Gilles
authors.initials: G
authors.lastname: Favre
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Cohen-Jonathan
authors.forename: Christine
authors.initials: C
authors.lastname: Toulas
mesh_terms: D017398:Alternative Splicing; D001483:Base Sequence; D015153:Blotting, Western; D002470:Cell Survival; D018076:DNA, Complementary / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D005091:Exons; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry; D020869:Gene Expression Profiling; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D007438:Introns; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D000090243:Nucleophosmin; D010957:Plasmids / Q000378:metabolism; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011836:Radiation Tolerance; D011839:Radiation, Ionizing; D014162:Transfection
chemical_list: D018076:DNA, Complementary; C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D016222:Fibroblast Growth Factor 2; D000090243:Nucleophosmin; D012313:RNA