The Werner syndrome protein promotes CAG/CTG repeat stability by resolving large (CAG)(n)/(CTG)(n) hairpins.
Expansion of CAG/CTG repeats causes certain neurological and neurodegenerative disorders, and the formation and subsequent persistence of stable DNA hairpins within these repeats are believed to contribute to CAG/CTG repeat instability. Human cells possess a DNA hairpin repair (HPR) pathway, which removes various (CAG)(n) and (CTG)(n) hairpins in a nick-directed and strand-specific manner. Interestingly, this HPR system processes a (CTG)(n) hairpin on the template DNA strand much less efficiently than a (CAG)(n) hairpin on the same strand (Hou, C., Chan, N. L., Gu, L., and Li, G. M. (2009) Incision-dependent and error-free repair of (CAG)(n)/(CTG)(n) hairpins in human cell extracts. Nat. Struct. Mol. Biol. 16, 869-875), suggesting the involvement of an additional component for (CTG)(n) HPR. To identify this activity, a functional in vitro HPR assay was used to screen partially purified HeLa nuclear fractions for their ability to stimulate (CTG)(n) HPR. We demonstrate here that the stimulating activity is the Werner syndrome protein (WRN). Although WRN contains both a 3'→5' helicase activity and a 3'→5' exonuclease activity, the stimulating activity was found to be the helicase activity, as a WRN helicase mutant failed to enhance (CTG)(n) HPR. Consistently, WRN efficiently unwound large (CTG)(n) hairpins and promoted DNA polymerase δ-catalyzed DNA synthesis using a (CTG)(n) hairpin as a template. We, therefore, conclude that WRN stimulates (CTG)(n) HPR on the template DNA strand by resolving the hairpin so that it can be efficiently used as a template for repair or replicative synthesis.
Graduate Center for Toxicology and Markey Cancer Center, University of Kentucky College of Medicine, Lexington, KY 40536, USA.
Nelson L S
NL
Chan
Caixia
C
Hou
Tianyi
T
Zhang
Fenghua
F
Yuan
Amrita
A
Machwe
Jian
J
Huang
David K
DK
Orren
Liya
L
Gu
Guo-Min
GM
Li
D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004258:DNA Polymerase III / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D055029:Inverted Repeat Sequences; D053484:RecQ Helicases / Q000235:genetics / Q000378:metabolism*; D019680:Trinucleotide Repeat Expansion; D000071657:Werner Syndrome Helicase
D004247:DNA; D004258:DNA Polymerase III; D005090:Exodeoxyribonucleases; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase