Microinjection of proteins and antibodies into living somatic cells is a powerful approach to studying the function of regulatory components of the cells. This method has been particularly useful in studies of oncogene proteins and their function in mammalian cell growth control, where it is difficult to alter the complement of expressed proteins by any other means. By introducing a functional oncogene protein, cell growth can be stimulated in a specific way; whereas by injection of an inhibitory antibody or oligonucleotide, cell growth can be specifically blocked. We will continue to utilize an efficient microinjection facility established during the past project period to study the regulation of nuclear oncogene activity by protein phosphorylation and dephosphorylation. In addition to microinjection of specific neutralizing antibodies and antisense oligonucleotides directed against components of the growth factor signal transduction system, we will microinject various signalling proteins and nuclear oncoproteins. These will include cJun, cJun mutants, PP2A catalytic and regulatory subunits, SH2 domains, tyrosine phosphatases, E2A-Pbx1, vAb1, Ha-Ras, casein kinase II, erkI and erkII. The effects of these macromolecules on cell proliferation and gene expression will be examined.
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