Detection of human betaV-tubulin expression in epithelial cancer cell lines by tubulin proteomics.
Tubulin, the constitutive protein of microtubules, is a heterodimeric protein with an alpha and beta subunit, encoded in vertebrates by six and seven different genes, respectively. Each tubulin isotype can be identified by its divergent C-terminal sequence. Nevertheless, two groups of beta-tubulin isotypes can be distinguished by sequence alignment; one includes betaI-, betaII-, betaIVa-, and betaIVb-tubulin, and the other includes betaIII-, betaV-, and betaVI-tubulin. betaIII-tubulin overexpression has been associated with microtubule destabilization and resistance to Taxol. Recent data indicate that mouse betaV-tubulin overexpression in CHO cells results in profound microtubule disorganization and dependence of cells on Taxol for growth. Mouse and human betaV-tubulin sequences display several differences, such as their respective extreme C-terminus, suggesting that they may have different effects on microtubule stability and different affinities for drugs. When high-resolution isoelectric focusing, in-gel CNBr cleavage, and mass spectrometry were combined, we detected for the first time the betaV-tubulin protein in human cell lines and found that it was highly expressed in Hey, an epithelial ovarian cancer cell line. Our data confirm that human and rodent betaV-tubulins are distinct and indicate that, regardless of species, betaIII- and betaV-tubulin may be expressed in a complementary pattern at the protein level. Therefore, both betaIII- and betaV-tubulin expression levels should be systematically determined to assess the role of differential tubulin isotype expression in the response of tumors to drugs targeting microtubules.
Verdier-Pinard
Pascal
P
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Shahabi
Shohreh
S
Wang
Fang
F
Burd
Berta
B
Xiao
Hui
H
Goldberg
Gary L
GL
Orr
George A
GA
Horwitz
Susan Band
SB
D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D002645:Chickens; D006224:Cricetinae; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007525:Isoelectric Focusing; D051379:Mice; D008969:Molecular Sequence Data; D017239:Paclitaxel / Q000494:pharmacology; D020033:Protein Isoforms / Q000235:genetics / Q000302:isolation & purification; D016415:Sequence Alignment; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014404:Tubulin / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification
D020033:Protein Isoforms; D014404:Tubulin; C507416:tubulin betaV, human; D017239:Paclitaxel