Our studies have continued to focus on the molecular interactions between glial cells and their infection with neurotropic JC virus which culminates in demyelination in the human brain, usually in the immunocompromised patient. Our current results reflect the foundation of our basic approach in accomplishing our experimental goals. Recently we have determined that gene sequences shared by a rat neuronal ID sequence and the JC virus do not function similarly as transcription enhancers in glial cells, JCV being highly active and ID inactive. The ID sequence, however, can function as an enhancer if placed in established cell lines. Both JCV sequences and ID sequence have cis acting elements which control transcription depending upon the cellular environment. Further study of the JCV DNA revealed a necessity of a protein binding site located downstream of the replication origin in order for viral DNA synthesis to occur in cells which produce a trans acting SV40 T protein. In attempts to identify proteins of glial cells which may interact with the JCV protein, we did find a potential cellular onc gene, p53, which is made independent of developmental stage in human fetal brain but not functionally associated with viral infection. Further studies of PML brain tissue confirmed the accuracy of using biotinylated DNA probes to provide a laboratory diagnosis of the association of JCV and lesions of PML and showed a significant advantage over using immunocytochemical detection of viral antigen. Several new isolations of JCV have been made: one from a PML patient diagnosed both clinically and in the laboratory using molecular techniques and the other from an owl monkey tumor. The PML virus isolate showed genomic changes compared with the wild type JCV. The owl monkey JCV isolate made a T protein which is structurally different from other isolates. Both of these new JCV strains provide evidence of genetic alteration of this virus.