The phosphorylated polypeptide (pp)38 of oncogenic Marek's disease (MD) herpesvirus (MDV) is expressed during lytic infections in vivo and in vitro, but its functions have not been fully elucidated. The quail cell line QT-35, latently infected with MDV, was used to generate QTP32 in which pp38 is expressed under control of a tetracycline controlled promoter to examine possible functions of pp38. Induction of pp38 did not influence late MDV genes expression, but it enhanced mitochondrial dehydrogenase activity significantly. Two new pp38 splice variants were found in induced QTP32 cells, in additional in vitro systems and MDV-infected chickens. Differential expression of full-length pp38 and splice variants suggests that the splice variants are important during latency and perhaps transformation. Polypeptides of 40 and 20kDa were detected by Western blot using monoclonal antibody H19. These polypeptides were also produced in DF-1 cells transfected with a pp38 construct in which the splice acceptor sites had been mutated. Our results add important new information to the role of pp38 in the pathogenesis of MD. The data suggest that pp38 and the two newly described splice variants may influence metabolic activity, which may have important consequences for the understanding of latency and tumor development.