I. Robert Lehman
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Our laboratory is concerned with the enzymology of eukaryotic DNA replication using as a model genome, the linear chromosome of herpes simplex Type I virus (HSV-1). The HSV-1 genome, like cellular chromosomes, contains multiple, redundant, origins of replication. The 150 kilobase HSV-1 genome has been completely sequenced. Of the seventy-five genes in the genome, seven are essential for HSV-1 DNA replication. The proteins encoded by these seven genes have been purified and analyzed in detail. They include a DNA polymerase, a protein that enhances the processivity of deoxynucleotide polymerization by the polymerase, a single-strand DNA binding protein, a heterotrimeric enzyme with both DNA helicase and primase activities, and an origin binding protein that binds specifically to the HSV-1 origins of replication.
Upon infection of susceptible cells, the linear HSV-1 chromosome circularizes. On the basis of two dimensional gel analysis of replicating HSV-1 chromosomes in vivo, we have found that HSV-1 chromosome enters a theta-mode of DNA replication, very likely initiated by the interaction of the origin binding protein with one of the HSV-1 origins. The products of theta replication then serve as substrates for rolling circle DNA replication. We have isolated a complex from infected cells that consists of the DNA polymerase, the processivity factor, the helicase-primase, and single strand DNA binding protein. This complex can promote rolling circle DNA replication in vitro. Our current efforts are directed toward reconstituting in vitro with purified enzymes, including host cell components, both phases of HSV-1 DNA replication.
A particularly intriguing feature of HSV-1 is its ability to infect non-dividing cells e.g., neurons, and enter a latent, quiescent state of infection. Upon introduction of the appropriate stimulus, the virus initiates its replication. We wish to understand the factors that regulate this latent-virulent switch.
Boehmer, P. E., Dodson, M. S. and Lehman, I. R. (1993) The herpes simplex virus type-1 origin binding protein. DNA helicase activity. J Biol Chem 268, (2): 1220-1225. (Medline)
Boehmer, P. E. and Lehman, I. R. (1993) Physical interaction between the herpes simplex virus 1 origin-binding protein and single-stranded DNA-binding protein ICP8. Proc Natl Acad Sci U S A 90, (18): 8444-8448. (Medline)
Dodson, M. S. and Lehman, I. R. (1993) The herpes simplex virus type I origin binding protein. DNA-dependent nucleoside triphosphatase activity. J Biol Chem 268, (2): 1213-1219. (Medline)
Lee, S.S.-K., Dong, Q., Wang, T.S.-F., and Lehman, I.R. (1994) Interaction of herpes simplex virus 1 origin-binding protein with DNA polymerase a. Proc. Natl. Acad. Sci. USA 92: 7882-7786. (Medline)
Skaliter, R., Makhov, A.M., Griffith, J.D., and Lehman, I.R. (1996) Rolling Circle DNA Replication by Extracts of Herpes Simplex Virus Type 1-Infected Human Cells. J. Virol. 70, 1132-1136. (Medline)