Activation of PKC has been postulated to play key roles in physiological cellular responses to a variety of signals. This includes various membrane-associated controls, such as exocytosis, receptor down-regulation and cross-talk among various signalling pathways. Evidence obtained with several cell systems suggests that sustained activation of PKC is needed for long-term cellular responses such as cell proliferation and differentiation. For example, the PKC activator, phorbol myristate acetate (PMA), will induce U937 cells to differentiate towards more mature macrophages. The work in this thesis describes studies of a putative Phospholipase D-Protein kinase C-MAPKinase pathway that leads to differentiation. U937 human promonocytic leukemia cells were chosen for these studies, since a clear relationship between PKC and differentiation was established. This was known to be associated with microtubule reorganisation and granule release. The studies here investigated specific molecular aspects of these events and the associated signals. It was established that sustained MAPK was itself sensitive to treatments designed to block PLD. This correlated with the release of granules and cell surface expression of β2 integrins. In parallel with this, PLD was found in fractionated membrane extracts alongside PKCβ1 and β2 integrins consistent with the notion that PKCβ1 is responsible for stimulating PLD1 in this compartment. The relationship between granule movement and MAPK activation led to an investigation into possible TPA-induced autocrine mechanisms. Partial inhibition of TPA-induced MAPK activation by marimastat and also blockade of endocytosis, led to the conclusion that an autocrine response contributed through the action of matrix-metalloproteinases to trigger a signal to MAPK downstream of an endocytic step. The implications of these conclusions are discussed throughout the thesis
