The aims of this proposal remain the detailed characterization of (1) the mechanisms by which efficient reproduction of measles virus is achieved and (2) the ways in which these pathways of replication are altered in the evolution of persistent measles virus infections, in order, ultimately, to (3) gain insight into the role played by such persistent infection in the pathogenesis of chronic human disease. Despite recent progress in defining the sequence, structure, and organization of the measles virus genome, the pathways by which transcription and genomic replication are accomplished remain vague and uncertain. Particular attention will be given to these, as yet undefined, features of viral reproduction by exploiting systems and preliminary observations recently generated by this laboratory, including: (1) an in vitro system able to accomplish transcription only when supplemented by a host cell factor -- a system in which the role(s) played by virus-specified and by host cell proteins can now be dissected and reconstructed, and (2) a membrane blot-hybridization system showing binding of measles virus genome and anti-genome RNA to a approximately 90 kD. host-cell protein --a method by which the polarity and sequences of viral RNA responsible for this host-cell protein:nucleic acid interaction can be analyzed and perhaps by which the possible significance of this interaction to transcription and/or replication can be assessed. In addition, using our extensively studied SSPE cell line (IP-3-Ca), efforts to identify the mechanism(s) by which measles virus matrix protein expression is restricted in the persistently infected brain cells of patients with SSPE will continue. The major aim now is to clone the matrix genes of the IP-3-Ca cell and of the genome of a revertent SSPE virus that emerged from this cell line, and to subject these cDNAs to sequence analysis in order to evaluate the hypothesis that genomic mutations accumulated in the course of measles virus persistence ultimately determine both the nature of and the level at which matrix protein expression is restricted in SSPE.
