Glucagon responsiveness was selectively lost in a dog kidney cell line, MDCK cells, after transformation by Harvey murine sarcoma virus and this loss can be restored to the transformed cells by culturing the cells in the presence of various inducers, including buytyrate and prostaglandin E-1 or E-2. The induction by prostaglandins is seen only when the treatment is carried out in the absence of serum. This inhibitor of differentiation in the serum has been partially purified. However, the nature of this factor, other than its ability to completely inhibit the induction, remains to be identified. We also found that EGF and a phorbol ester (TPA) inhibit the induction of glucagon sensitivity by PGE-2. The inhibitory effect of TPA is seen at a concentration as low as 0.1 nM. We have observed that acute treatment of normal MDCK cells with TPA leads to loss of hormone response, however, transformed cells appear to be resistant to this uncoupling effect of TPA. Therefore, it is likely that TPA inhibits the induction process itself rather than the expression of induced glucagon sensitivity. TPA is known to activate protein kinase C, which can phosphorylate tyrosine residues in proteins. We are currently studying the involvement of protein kinase C in the susceptibility of normal cells and resistance of transformed cells to the uncoupling effect of TPA. We are also exploring the possible interaction between protein kinase C and the cyclic AMP-dependent protein kinase, since TPA antagonizes the induction by PGE-2, which seems to be mediated by cyclic AMP. We have continued our effort to raise antibodies against glucagon receptors and other cell surface markers which are modulated by cellular differentiation. We have recently, using intact MDCK cells and plasma membranes from rat liver or MDCK cells as antigens, attempted to prepare monoclonal antibodies. Our initial results indicate that we have monoclonal antibodies inhibiting binding of l25I-glucagon to MDCK cells. We are now expanding the hybridoma cultures and the conditioned media will be further characterized to establish the nature of these antibodies.