Abstract

We have developed a soluble, cell-free system that carries out the replication of exogenous DNA templates containing the SV40 origin of DNA replication. We plan to use this system to explore the molecular mechanisms involved in viral and cellular DNA replication.
Our specific aims for the initial grant period are as follows: (1) To analyze the structure and function of the SV40 origin of replication by studying the effects of deletions and point mutations on DNA replication in vitro. (2) To purify and functionally characterize cellular proteins involved in DNA replication with particular emphasis on the proteins required for the initiation reaction. (3) To extend the in vitro system to the analysis of initiation at cellular origins of replication.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA040414-03
Application #
3180303
Study Section
Virology Study Section (VR)
Project Start
1985-07-01
Project End
1990-06-30
Budget Start
1987-07-01
Budget End
1988-06-30
Support Year
3
Fiscal Year
1987
Total Cost
Indirect Cost

  Institution

Name
Johns Hopkins University
Department
Type
Schools of Medicine
DUNS #
045911138
City
Baltimore
State
MD
Country
United States
Zip Code
21218

  Publications

Wu, Min; Lu, Wenyan; Santos, Ruth E et al. (2014) Geminin inhibits a late step in the formation of human pre-replicative complexes. J Biol Chem 289:30810-21
Dai, Jianli; Chuang, Ray-Yuan; Kelly, Thomas J (2005) DNA replication origins in the Schizosaccharomyces pombe genome. Proc Natl Acad Sci U S A 102:337-42
Vashee, S; Simancek, P; Challberg, M D et al. (2001) Assembly of the human origin recognition complex. J Biol Chem 276:26666-73
Collins, K L; Russo, A A; Tseng, B Y et al. (1993) The role of the 70 kDa subunit of human DNA polymerase alpha in DNA replication. EMBO J 12:4555-66
Moarefi, I F; Small, D; Gilbert, I et al. (1993) Mutation of the cyclin-dependent kinase phosphorylation site in simian virus 40 (SV40) large T antigen specifically blocks SV40 origin DNA unwinding. J Virol 67:4992-5002
Virshup, D M; Russo, A A; Kelly, T J (1992) Mechanism of activation of simian virus 40 DNA replication by protein phosphatase 2A. Mol Cell Biol 12:4883-95
Randall, S K; Kelly, T J (1992) The fate of parental nucleosomes during SV40 DNA replication. J Biol Chem 267:14259-65
Erdile, L F; Collins, K L; Russo, A et al. (1991) Initiation of SV40 DNA replication: mechanism and control. Cold Spring Harb Symp Quant Biol 56:303-13
Scheidtmann, K H; Virshup, D M; Kelly, T J (1991) Protein phosphatase 2A dephosphorylates simian virus 40 large T antigen specifically at residues involved in regulation of DNA-binding activity. J Virol 65:2098-101
Weinberg, D H; Collins, K L; Simancek, P et al. (1990) Reconstitution of simian virus 40 DNA replication with purified proteins. Proc Natl Acad Sci U S A 87:8692-6

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