Geminiviruses constitute a family of plant viruses with single- stranded DNA genomes that replicate via double-stranded DNA intermediates in the nuclei of infected cells. They encode a single protein for their replication and can recruit the remainder of the replication machinery from their host. These characteristics suggest that geminiviruses are excellent models for studying plant nuclear DNA replication. The overall goal of our research is to understand the molecular events that mediate geminivirus DNA replication using tomato golden mosaic virus (TGMV) as our prototype. We will identify the TGMV replication origin by deletion analysis and by assaying viral DNA fragments for their ability to support replication of nonviral DNA in plant protoplasts. The sequence and structural requirements of the TGMV origin will be characterized by analyzing replications containing site-specific mutations. Our initial studies of trans-acting factors involved in replication will focus on the single TGMV- encoded protein, AL1, required for replication. Two important questions we will ask are does AL1 bind specifically to the TGMV replication origin and what is its catalytic role in replication. We plan to purify AL1, study its catalytic properties, and determine if it requires host proteins for activity. The long term goal of our research is to identify and characterize the plant proteins involved in TGMV replication and to determine their roles in normal cellular processes.***//
