The C terminus of beta-tubulin regulates vinblastine-induced tubulin polymerization.
Oligoanions such as sodium triphosphate or GTP prevent and/or reverse vinblastine-induced polymerization of tubulin. We now show that the anions of glutamate-rich extreme C termini of tubulin are similarly involved in the regulation of the vinblastine effect. Cleavage of the C termini by limited proteolysis with subtilisin enhances vinblastine-induced tubulin polymerization and abolishes the anion effect. Only the beta-tubulin C terminus needs to be removed to achieve these changes and the later cleavage of the alpha-tubulin C terminus has little additional effect. In fact, vinblastine concentrations >20 microM block cleavage of the alpha-tubulin C terminus in the polymer, whereas cleavage of the beta-tubulin C terminus proceeds unimpeded over the time used. The vinblastine effect on tubulin polymerization is also highly pH-dependent between pH 6.5 and 7.5; this is less marked, but not absent, after subtilisin treatment. A working model is proposed wherein an anionic domain proximal to the extreme C terminus must interact with a cationic domain to permit vinblastine to promote polymerization. Both exogenous and extreme C-terminal anions compete for the cationic domain with the proximal anionic domain to prevent vinblastine-induced polymerization. We conclude that the electrostatic regulation of tubulin polymerization induced by vinblastine resides primarily in the beta-tubulin C terminus but that additional regulation proximal in the tubulin molecule also plays a role.
Rai
S S
SS
Laboratory of Biochemistry and Genetics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0830, USA.
Wolff
J
J
D000818:Animals; D001921:Brain / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008453:Maytansine / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry; D051381:Rats; D013381:Subtilisins; D014404:Tubulin / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D014747:Vinblastine / Q000494:pharmacology*
D046911:Macromolecular Substances; D010446:Peptide Fragments; D014404:Tubulin; D008453:Maytansine; D014747:Vinblastine; D013381:Subtilisins