title: Quantitative proteomic analysis and functional studies reveal that nucleophosmin is involved in cell death in glioblastoma cell line transfected with siRNA.
abstract: Previously, we reported that nucleophosmin (NPM) was increased in glioblastoma multiforme (GBM). NPM is a phosphoprotein related to apoptosis, ribosome biogenesis, mitosis, and DNA repair, but details about its function remain unclear. We treated U87MG and A172 cells with small interference RNA (siRNA) and obtained a reduction of 80% in NPM1 expression. Knockdown at the protein level was evident after the 4th day and was maintained until the 7th day of transfection that was investigated by quantitative proteomic analysis using isobaric tags. The comparison of proteomic analysis of NPM1-siRNA against controls allowed the identification of 14 proteins, two proteins showed increase and 12 presented a reduction of expression levels. Gene ontology assigned most of the hypoexpressed proteins to apoptosis regulation, including GRP78. NPM1 silencing did not impair cell proliferation until the 7th day after transfection, but sensitized U87MG cells to temozolomide (TMZ), culminating with an increase in cell death and provoking at a later period a reduction of colony formation. In a large data set of GBM patients, both GRP78 and NPM1 genes were upregulated and presented a tendency to shorter overall survival time. In conclusion, NPM proved to participate in the apoptotic process, sensitizing TMZ-treated U87MG and A172 cells to cell death, and in association with upregulation of GRP78 may be helpful as a predictive factor of poor prognosis in GBM patients.
authors: Protein Chemistry Center and Department of Molecular and Cell Biology, Medical School of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil.
authors: Marcela
authors: M
authors: Gimenez
authors: Suely K N
authors: SK
authors: Marie
authors: Sueli M
authors: SM
authors: Oba-Shinjo
authors: Miyuki
authors: M
authors: Uno
authors: Roseli
authors: R
authors: da Silva
authors: Helen Julie
authors: HJ
authors: Laure
authors: Clarice
authors: C
authors: Izumi
authors: Andreia
authors: A
authors: Otake
authors: Roger
authors: R
authors: Chammas
authors: Jose Cesar
authors: JC
authors: Rosa
mesh_terms: D000328:Adult; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000091342:Endoplasmic Reticulum Chaperone BiP; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D000090243:Nucleophosmin; D011379:Prognosis; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D000077204:Temozolomide; D014162:Transfection
chemical_list: D018906:Antineoplastic Agents, Alkylating; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D020543:Proteome; D034741:RNA, Small Interfering; D000090243:Nucleophosmin; D003606:Dacarbazine; D000077204:Temozolomide