The Werner syndrome helicase/exonuclease processes mobile D-loops through branch migration and degradation.
RecQ DNA helicases are critical for preserving genome integrity. Of the five RecQ family members identified in humans, only the Werner syndrome protein (WRN) possesses exonuclease activity. Loss of WRN causes the progeroid disorder Werner syndrome which is marked by cancer predisposition. Cellular evidence indicates that WRN disrupts potentially deleterious intermediates in homologous recombination (HR) that arise in genomic and telomeric regions during DNA replication and repair. Precisely how the WRN biochemical activities process these structures is unknown, especially since the DNA unwinding activity is poorly processive. We generated biologically relevant mobile D-loops which mimic the initial DNA strand invasion step in HR to investigate whether WRN biochemical activities can disrupt this joint molecule. We show that WRN helicase alone can promote branch migration through an 84 base pair duplex region to completely displace the invading strand from the D-loop. However, substrate processing is altered in the presence of the WRN exonuclease activity which degrades the invading strand both prior to and after release from the D-loop. Furthermore, telomeric D-loops are more refractory to disruption by WRN, which has implications for tighter regulation of D-loop processing at telomeres. Finally, we show that WRN can recognize and initiate branch migration from both the 5' and 3' ends of the invading strand in the D-loops. These findings led us to propose a novel model for WRN D-loop disruption. Our biochemical results offer an explanation for the cellular studies that indicate both WRN activities function in processing HR intermediates.
Opresko
Patricia L
PL
Department of Environmental and Occupational Health, University of Pittsburgh Graduate School of Public Health, Pittsburgh, Pennsylvania, United States of America. plo4@pitt.edu
Sowd
Gregory
G
Wang
Hong
H
D004260:DNA Repair; D004261:DNA Replication; D005090:Exodeoxyribonucleases / Q000235:genetics*; D005092:Exonucleases / Q000235:genetics*; D006801:Humans; D010957:Plasmids / Q000235:genetics; D053484:RecQ Helicases / Q000235:genetics*; D011995:Recombination, Genetic; D016615:Telomere / Q000235:genetics; D014898:Werner Syndrome / Q000235:genetics*; D000071657:Werner Syndrome Helicase
D005090:Exodeoxyribonucleases; D005092:Exonucleases; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase