Wild-type class I beta-tubulin sensitizes Taxol-resistant breast adenocarcinoma cells harboring a beta-tubulin mutation.
A Taxol-resistant cell line, K20T, which does not express P-glycoprotein, was selected with Taxol from human MDA-MB-231 breast adenocarcinoma cells and maintained in the presence of 20nM Taxol. K20T cells were approximately 18-fold resistant to Taxol, displayed cross-resistance to Taxotere and the epothilones, but little cross-resistance to discodermolide. Sequence analysis of the class I beta-tubulin indicated that it harbored an A593G mutation resulting in a change from glutamate to glycine at amino acid 198, which is near the intradimer interface within the alpha/beta-tubulin heterodimer. An HA-tagged wild-type class I beta-tubulin expression vector was transfected into the K20T cells. Immunofluorescence studies demonstrated that this exogenous tubulin was incorporated into cellular microtubules and Western blot analysis indicated that the K20T transfectants predominantly expressed the exogenous wild-type class I beta-tubulin. The transfected cells were only approximately 5-fold resistant to Taxol. Our results, plus the knowledge that Glu198 is the target for other anti-tubulin agents, suggest that glutamate198 in beta-tubulin is a critical determinant for microtubule stability and Taxol resistance.
Wiesen
Kenneth M
KM
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Xia
Shujun
S
Yang
Chia-Ping Huang
CP
Horwitz
Susan Band
SB
D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000473:Alkanes / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D002219:Carbamates / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D019281:Dimerization; D000077143:Docetaxel; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D034261:Epothilones / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D018698:Glutamic Acid / Q000235:genetics; D005998:Glycine / Q000235:genetics; D006801:Humans; D007783:Lactones / Q000494:pharmacology; D008958:Models, Molecular; D020125:Mutation, Missense; D017239:Paclitaxel / Q000494:pharmacology*; D020836:Protein Structure, Quaternary; D011753:Pyrones / Q000494:pharmacology; D043823:Taxoids / Q000494:pharmacology; D014162:Transfection; D014404:Tubulin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D050257:Tubulin Modulators / Q000494:pharmacology
D000473:Alkanes; D000972:Antineoplastic Agents, Phytogenic; D002219:Carbamates; D034261:Epothilones; D007783:Lactones; D011753:Pyrones; D043823:Taxoids; D014404:Tubulin; D050257:Tubulin Modulators; D000077143:Docetaxel; D018698:Glutamic Acid; C071229:discodermolide; D017239:Paclitaxel; D005998:Glycine