Document Detail


Okazaki fragment processing-independent role for human Dna2 enzyme during DNA replication.
MedLine Citation:
PMID:  22570476     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Dna2 is an essential helicase/nuclease that is postulated to cleave long DNA flaps that escape FEN1 activity during Okazaki fragment (OF) maturation in yeast. We previously demonstrated that the human Dna2 orthologue (hDna2) localizes to the nucleus and contributes to genomic stability. Here we investigated the role hDna2 plays in DNA replication. We show that Dna2 associates with the replisome protein And-1 in a cell cycle-dependent manner. Depletion of hDna2 resulted in S/G(2) phase-specific DNA damage as evidenced by increased γ-H2AX, replication protein A foci, and Chk1 kinase phosphorylation, a readout for activation of the ATR-mediated S phase checkpoint. In addition, we observed reduced origin firing in hDna2-depleted cells consistent with Chk1 activation. We next examined the impact of hDna2 on OF maturation and replication fork progression in human cells. As expected, FEN1 depletion led to a significant reduction in OF maturation. Strikingly, the reduction in OF maturation had no impact on replication fork progression, indicating that fork movement is not tightly coupled to lagging strand maturation. Analysis of hDna2-depleted cells failed to reveal a defect in OF maturation or replication fork progression. Prior work in yeast demonstrated that ectopic expression of FEN1 rescues Dna2 defects. In contrast, we found that FEN1 expression in hDna2-depleted cells failed to rescue genomic instability. These findings suggest that the genomic instability observed in hDna2-depleted cells does not arise from defective OF maturation and that hDna2 plays a role in DNA replication that is distinct from FEN1 and OF maturation.
Authors:
Julien P Duxin; Hayley R Moore; Julia Sidorova; Kenneth Karanja; Yuchi Honaker; Benjamin Dao; Helen Piwnica-Worms; Judith L Campbell; Raymond J Monnat; Sheila A Stewart
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.     Date:  2012-05-07
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  287     ISSN:  1083-351X     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2012 Jun 
Date Detail:
Created Date:  2012-06-25     Completed Date:  2012-09-20     Revised Date:  2013-06-25    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  21980-91     Citation Subset:  IM    
Affiliation:
Department of Cell Biology and Physiology, University of Washington, Seattle, Washington 98195, USA.
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MeSH Terms
Descriptor/Qualifier:
Cell Cycle
Cell Line, Tumor
Cell Nucleus / metabolism
Chromatin / metabolism
DNA* / chemistry
DNA Damage
DNA Helicases / chemistry,  physiology*
DNA Repair
DNA Replication*
Gene Expression Regulation
HeLa Cells
Humans
Kinetics
Micronucleus Tests
Microscopy, Fluorescence / methods
Grant Support
ID/Acronym/Agency:
GM04701/GM/NIGMS NIH HHS; GM04707/GM/NIGMS NIH HHS; GM078666/GM/NIGMS NIH HHS; GM95924/GM/NIGMS NIH HHS; P01 CA077852/CA/NCI NIH HHS; P01 CA77852/CA/NCI NIH HHS
Chemical
Reg. No./Substance:
0/Chromatin; 0/Okazaki fragments; 9007-49-2/DNA; EC 3.6.1.-/DNA Helicases; EC 3.6.4.12/DNA2 protein, human
Comments/Corrections

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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