In this application we propose to examine myelin protein gene expression and related events in the differentiating and maturing oligodendrocyte both in vivo and in vitro. We have been successful at immortalizing oligodendrocytes from normal and shiverer mice with a retrovirus containing a temperature sensitive SV40 large T antigen oncogene, many of which may be useful models of oligodendrocyte differentiation. We propose to characterize these lines with respect to their differentiation state and their expression of oligodendrocyte-specific and myelin protein genes. We propose to test the potential of these lines to differentiate in vivo through transplantation studies and in vivo through exposure to a variety of growth factors and hormones. A major portion of this application is to investigate the mechanisms by which MBP mRNAs and other mRNAs are translocated from oligodendrocyte cell bodies to their processes in a variety of in vitro models including primary cultures and immortalized cells. We wish to examine the role that cytoskeletal elements play in this process and the nature of the ribonucleoprotein particles with which MBP mRNAs become associated at different stages in oligodendrocyte differentiation. Preliminary data indicate that astrocytes, and possibly astrocyte-conditioned media can influence the translocation of MBP mRNAs in oligodendrocytes. We propose to pursue these findings by determining the specificity of the response and the astrocytic components responsible for the effect. We have found that certain cytokeratins are expressed in cells with phenotypes similar to immature oligodendrocytes or their precursors. We propose to investigate the expression of these and any other cytokeratins in the differentiating oligodendrocyte in vivo and in vitro. A final set of experiments are proposed to examine myelin protein gene expression, MBP mRNA translocation and oligodendrocyte differentiation in primary cultures of jimpy mouse brains and jimpy oligodendrocytes immortalized by transfection with a retrovirus containing the temperature-sensitive large T antigen oncogene.
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