The broad, long-term objective is to understand the Molecular mechanisms of chronic virus infections in the human central nervous system (CNS). Measles virus (MV) causes millions of death annually in infants and children on a global scale. MV can cause acute and subacute CNS infections. Recent research suggests that a neural cell RNA-modifying activity may introduce genetic changes called biased hypermutation in MV and may alter its mode of replication in a CNS infection. The present project investigates this unique virus-host interaction.
The specific aims of this project are (1) to test the model that biased hypermutation occurs in the MV transcription complex, (2) to purify the RNA-modifying enzyme implicated in hypermutation and clone the corresponding cellular gene, and (3) to characterize the RNA-modifying enzyme and prove its role in hypermutation by use of the cloned gene. The first goal will be accomplished by studying MV transcription in neuroblastoma cells which contain high RNA-modifying activity or in vitro in neuroblastoma cell lysates. The second goal will be achieved by purifying the RNA-modifying enzyme from bovine brain, which contains the RNA-modifying activity. The purified protein will be sequenced to generate oligonucleotide probes for cloning the cDNA encoding the enzyme. The final goal will be accomplished by generating antibodies and gene probes to study the expression of the RNA-modifying protein in different cell types. The RNA-modifying enzyme will be overexpressed in mammalian cells with low intrinsic RNA-modifying activity. MV will be passaged in those cells to see whether hypermutation occurs with increased frequency.
