CELLULAR CONTROL OF NEURONAL CYTOSKELETON ASSEMBLY

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The long term objective of the proposed research is to understand how the cytoskeleton of neuroblastoma cells is assembled and organized. The role of 215,000 dalton microtubule associated protein (215 K MAP) that is present in neurite-bearing neuroblastoma cells but absent in non-differentiated cells will be examined in detail. In vivo analyses will establish the synthetic pattern of this protein during stages of cell attachment and neurite formation. Immunofluorescence microscopy with antisera against the 215 K MAP and tubulin will indicate the distribution of these proteins at various stages of microtubule organization. These results will be compared to those obtained using cellular fractionation to analyze the distribtuion and amount of the 215 K MAP relative to soluble and assembled tubulin. In vitro assays will establish the mechanisms by which the 215 K MAP interacts with tubulin, and biochemical procedures will demonstrate whether this protein is post-translationally modified. Peptide mapping and monoclonal antibodies will be used to investigate the extenet of homology between the 215 K MAP and other microtubule associated proteins in neuroblastoma cells, as well as to microtubule associated proteins from other cell types. Limited studies will be undertaken to examine the relationship between microtubules and neurofilaments during neurite development, and to establish whether any of the microtubule associated proteins coordinate these two cytoskeletal systems. These investigations should provide basic information on the biological role of microtubule associated proteins in the organization of microtubules, and on the biochemical properties of this class of specialized proteins.