The isolation and characterization of microtubule mutants using drug-resistance selections has provided insights into the role of microtubules in mitosis, the mechanism of drug-resistance in mammalian cells, and regulatory mechanisms in microtubule assembly. Recently, methods have been devised which enhance our ability to isolate mutants; and ideas on how to use mutants to study microtubule function, assembly, and regulation have been refined. Using this information, we propose to isolate many new mutants to determine the range of mutant phenotypes and to test developing ideas on the mechanism of drug action and cellular mechanisms of resistance to their effects. Considerable effort will be devoted toward the development of new selections for obtaining mutants in microtubule assembly and in the expression of microtubule associated proteins (MAPs). We already have one mutant of the first type and believe we may have one of the second. At the same time it will be necessary to develop tools to fully characterize these mutants. Included in these studies will be the isolation and characterization of the spindle apparatus from CHO cells, the identification of MAPs, and the generation of antibodies against various MAP and spindle components. This work should complement and extend our current knowledge of microtubule physiology and provide a basis for understanding drug action and cellular mechanisms of resistance.
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