Identification of cytoskeletal [14C]carboplatin-binding proteins reveals reduced expression and disorganization of actin and filamin in cisplatin-resistant cell lines.
Cisplatin resistant (CP-r) cells often show decreased uptake of cisplatin in association with reduced cell surface proteins and decreased endocytosis. In this report, two major [14C]carboplatin-binding proteins were identified as filamin and actin by photoaffinity labeling and mass spectrometry. Decreased expression of these two proteins was found in two different human CP-r cell lines (KB-CP20 and 7404-CP20), in comparison with their parental cell lines (KB-3-1 and BEL-7404), respectively. Disorganization of beta-actin and filamin 250 and 90 was also detected in these CP-r cells by confocal microscopy. Transfection of a wild-type actin-enhanced green fluorescent protein (EGFP) expression vector into 7404-CP20 cells resulted in a nonfilamentous actin-EGFP distribution compared with a normal distribution in the cisplatin-sensitive BEL-7404 cells, suggesting that cytoskeletal organization is disturbed in the CP-r cells. The identification of actin and filamin as [14C]carboplatin-binding proteins and decreased expression and disorganization of several cytoskeletal proteins in CP-r cells provide a molecular and cellular basis for the known defect in endocytosis in these cells.
Shen
Ding-Wu
DW
Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, 37 Convent Drive, Room 1A09, Bethesda, MD 20892-4254, USA.
Liang
Xing-Jie
XJ
Gawinowicz
Mary Ann
MA
Gottesman
Michael M
MM
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology; D002250:Carbon Radioisotopes; D016190:Carboplatin / Q000378:metabolism*; D002945:Cisplatin / Q000494:pharmacology; D003285:Contractile Proteins / Q000302:isolation & purification / Q000378:metabolism*; D003599:Cytoskeleton / Q000737:chemistry / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000502:physiology*; D034285:Dynamin II / Q000378:metabolism; D064448:Filamins; D015870:Gene Expression; D006801:Humans; D007633:Keratins / Q000378:metabolism; D008840:Microfilament Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D014404:Tubulin / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000199:Actins; D000970:Antineoplastic Agents; D002250:Carbon Radioisotopes; D003285:Contractile Proteins; D064448:Filamins; D008840:Microfilament Proteins; D014404:Tubulin; D007633:Keratins; D016190:Carboplatin; D034285:Dynamin II; D002945:Cisplatin