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Purification of Viral DNA for the Identification of Associated Viral and Cellular Proteins
Published on: August 31, 2017
Herpes simplex virus type 1 single strand DNA binding protein and helicase/primase complex disable cellular ATR
Kareem N Mohni1, Samantha Smith, Alexander R Dee
1Department of Molecular, Microbial and Structural Biology and the Molecular Biology and Biochemistry Graduate Program, University of Connecticut Health Center, Farmington, Connecticut, United States of America.
Plos Pathogens
|October 8, 2013
Summary
Herpes Simplex Virus type 1 (HSV-1) disables the cell's ATR DNA damage response. Viral proteins ICP8 and helicase/primase block the 9-1-1 clamp, preventing ATR activation and cell signaling.
Area of Science:
- Virology
- Molecular Biology
- Cellular Biology
Background:
- Herpes Simplex Virus type 1 (HSV-1) infection interferes with cellular DNA damage response pathways.
- HSV-1-infected cells exhibit impaired Chk1 phosphorylation, a key event in ATR signaling, even when replication forks stall.
Purpose of the Study:
- To elucidate the mechanism by which HSV-1 disables the ATR DNA damage response kinase.
- To identify the specific viral proteins involved in inhibiting ATR signaling.
Main Methods:
- Utilized transfected cells to study the interaction of HSV-1 proteins with cellular DNA damage response machinery.
- Investigated the localization and complex formation of viral proteins (ICP8, UL8/UL5/UL52) at sites of DNA damage.
- Assessed the recruitment of ATR/ATRIP, RPA, and the 9-1-1 checkpoint clamp in the presence of viral proteins.
Main Results:
- The HSV-1 single-stranded DNA binding protein (ICP8) and the helicase/primase complex (UL8/UL5/UL52) form a nuclear complex that disables ATR signaling.
- This viral complex localizes to DNA damage sites and interacts with ATR/ATRIP and RPA but excludes the 9-1-1 checkpoint clamp.
- The viral proteins are proposed to bind the DNA substrate, preventing 9-1-1 clamp loading and subsequent recruitment of TopBP1, the ATR activator.
Conclusions:
- HSV-1 viral proteins ICP8 and helicase/primase directly inhibit ATR signaling by obstructing the DNA damage recognition site.
- This mechanism prevents the loading of the 9-1-1 checkpoint clamp and TopBP1, effectively disabling the ATR-mediated DNA damage response.
- This represents a novel strategy where viral replication proteins interfere with host DNA damage signaling pathways, offering potential therapeutic targets.
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