Test of four possible mechanisms for the temporal control of spindle and cytoplasmic microtubule assembly in HeLa cells.

We have compared tubulin from exponentially growing and mitotic enriched populations of suspensioncultured HeLa cells. The proportion of the protein in the cell extract that was tubulin (4.6%) was the same in populations of both mitotic and log phase cells. The self assembly and copolymerization activity of extract tubulin were also compared for the two populations of cells. Approximately 60% of the tubulin in extracts of both mitotic enriched and log phase cells self assembled reversibly as determined by quantitative sedimentation assays. In copolymerization experiments, isotopically labeled tubulin from both mitotic enriched and log phase cell extracts was incorporated with equal efficiency into microtubules formed with microtubule protein purified from porcine brain. Greater than 80% of the tubulin in HeLa extracts participated in one cycle of reversible polymerization with carrier brain tubulin. In addition, the amount and type of microtubule-associated proteins (MAPs) were similar in the extracts of mitotic and log phase cells. The capacity of tubulin from the two populations to incorporate tyrosine into the COOH terminus of the (Y chain was also compared before and after partiaI digestion with carboxypeptidase A to release any endogenously bound COOH-terminal tyrosine. Once again, the results obtained for the material from the two sources showed no significant difference. In summary, no differences have yet been detected in the properties of microtubule protein in interphase and mitotic HeLa cells.