The aim of this project is to better elucidate the role(s) of specific protein kinases in the regulation of cell growth and in malignant transformation. Protein kinase C (PKC) has been shown to play a crucial role in the regulation of a number of physiological processes. Since PKC consists of a family of at least 10 closely related isotypes, it is important to identify cellular targets and biological systems which are selectively modulated by the different PKC isoforms to regulate specific cellular functions. Studies utilizing PKC isotype-specific inhibitors and antisense oligonucleotides established that PKC epsilon is the isoform involved in mediating the phorbol ester tumor promoter (PMA)-induced activation of the PiT-2 phosphate (Pi) transporter/viral receptor. Further, it was determined that infection of cells with amphotropic murine leukemia virus (A-MuLV) resulted in the specific down-modulation of PiT-2- mediated Pi uptake and resistance to superinfection with A-MuLV. When cells expressing epitope-tagged PiT-2 were infected with A-MuLV the tagged transporters/viral receptors were no longer detected at the plasma membrane, but rather were localized to a cytosolic compartment. It appears that this loss of PiT-2 viral receptors from the cell membrane is responsible for superinfection interference induced with A-MuLV infection. .In other studies to determine biological activities specifically regulated by the different isoforms of protein kinase C, it was found that PMA-induced growth inhibition of low population density NIH 3T3 cells was selectively mediated through activation of PKC epsilon.. Studies also were carried out with PC-12 cells to examine the role of specific PKC isoforms in mediating a mitogenic versus a differentiation signal in response to nerve growth factor (NGF) and epidermal growth factor (EGF). Overexpression of PKC epsilon was found to enhance NGF-induced differentiation, whereas overexpression of a dominant-negative PKC epsilon construct blocked this growth arrest. In studies carried out with human intestinal mucosal cells, results indicated that PKC epsilon also was involved in mediating the expression of two major intestinal mucins in response to PMA, in mediating PMA-induced mucin secretion in these cells. These results indicate that altering the levels of specific PKC isoforms can markedly alter the biological effect noted in response to agents which regulate PKC activation, such as hormones, growth factors, and PMA. These findings also lend further support to a prominent role for PKC epsilon in cell growth regulation, as well as in the regulation of Golgi function and protein secretion. . - cell proliferation, Golgi, Oxidative Regulation, Protein Kinases, Signal Transduction, Subcellular Targeting, viral receptors,
