Role of microtubules and related proteins on cellular function

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The purpose of this co-operative research project has been to clarify the mechanism of regulation of various cellular functions by cytoskeletons. There were six subprojects. 1. Function of microtubule-dynein system in mitosis The motive force for the chromosome movements was localized in the half spindle region near chromosomes. The formation of barrel-shaped spindle in the presence of T-1 was thought to be a result of scattering of the microtubule-organizing granules by T-1. MAP1 was localized on the cytoskeletal microtubules at <G_0> stage of cultured cells while it became concentrated in the nucleus at <G_1> stage. 2. Cytoskeletal proteins and cell motility Microtubules were found to undergo spontaneous assembly-disassembly cycle by dark field microscopy. Intermediate filaments were found to be relevant to mitosis and exchange of nuclei after conjugation in Tetrahymena. Complete amino acid sequences of Tetrahymena actin and of -tubulin from lymphoma cells were determined. 3. Dynamic state of microtubules in axons It was found that the 200K subunit of neurofilament is phosphorylated and transported more faster than the bulk neurofilament proteins. Axolinin was found to co-exist with microtubules in squid giant axon. 4. Function of cytoskeleton-modulating proteins A 100K-molecular weight Casensitive actin filament-severing protein was found in sea urchin eggs. Alphaactinin was found to be concentrated in the cortex of the fertilized sea urchin egg together with actin filaments. 5. Nerve excitability and microtubules Calmodulin antagonists were found to interfere with Na-current in squid nerve. 6. Cytoskeleton and cellular function Ultrastructure of the plasma membrane undercoat in skeletal muscle and astrocytes where myofibrils and intermediate filaments, respectively, attach was visualized.