VSV RNA TRANSCRIPTION AND REPLICATION

The long term objectives of this research proposal are to understand at
the molecular level the basic mechanisms controlling transcription and
replication of vesicular stomatitis virus (VSV) and its defective
interfering (DI) particle RNAs. VSV, the prototypical rhabdovirus with a
single-stranded negative-sense RNA genome is an excellent model system for
numerous other disease-causing negative-strand RNA viruses. It has also
been used as a model virus of choice for studying basic mechanisms of
negative-strand RNA virus genome transcription and replication and virus
assembly. The proposed research is designed to gain knowledge on various
transcription and replication regulating sequences and their functions.
A recently developed reverse genetic system that utilizes only cDNAs for
VSV proteins and RNA will be used to introduce specific alterations into
the genome of VSV. The effect of these mutations on transcription and
replication of VSV RNA will be measured by various biochemical means.
identification and characterization of the regulatory sequences by the
methods described above will be essential to gain insight into the
mechanisms of RNA transcription and replication in this group of important
human pathogens. The specific aims of this proposal are as follows: 1. Competition and interference studies with different size DI particles. 2. Identification and characterization of signals for encapsidation and
replication of DI particle genomic RNA. 3. Identification and characterization of various transcription regulatory
sequence elements within the genome of VSV. 4. Evaluation of the role of sequences at or near the leader-N gene
junction of VSV in transcription attenuation or readthrough.