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Link to original content: https://pubmed.ncbi.nlm.nih.gov/20829486
RETRACTED: Human SIRT6 promotes DNA end resection through CtIP deacetylation - PubMed Skip to main page content
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. 2010 Sep 10;329(5997):1348-53.
doi: 10.1126/science.1192049.

RETRACTED: Human SIRT6 promotes DNA end resection through CtIP deacetylation

Abderrahmane Kaidi  1 , Brian T Weinert, Chunaram Choudhary, Stephen P Jackson
Affiliations

RETRACTED: Human SIRT6 promotes DNA end resection through CtIP deacetylation

Abderrahmane Kaidi et al. Science. .

Retraction in

  • Retraction.
    Kaidi A, Weinert BT, Choudhary C, Jackson SP. Kaidi A, et al. Science. 2019 Apr 19;364(6437):247. doi: 10.1126/science.aax4558. Epub 2019 Apr 11. Science. 2019. PMID: 30975768 Free PMC article. No abstract available.

Expression of concern in

  • Editorial expression of concern.
    Berg J. Berg J. Science. 2018 Sep 28;361(6409):1322. doi: 10.1126/science.aav4528. Epub 2018 Sep 27. Science. 2018. PMID: 30262489 No abstract available.

Abstract

SIRT6 belongs to the sirtuin family of protein lysine deacetylases, which regulate aging and genome stability. We found that human SIRT6 has a role in promoting DNA end resection, a crucial step in DNA double-strand break (DSB) repair by homologous recombination. SIRT6 depletion impaired the accumulation of replication protein A and single-stranded DNA at DNA damage sites, reduced rates of homologous recombination, and sensitized cells to DSB-inducing agents. We identified the DSB resection protein CtIP [C-terminal binding protein (CtBP) interacting protein] as a SIRT6 interaction partner and showed that SIRT6-dependent CtIP deacetylation promotes resection. A nonacetylatable CtIP mutant alleviated the effect of SIRT6 depletion on resection, thus identifying CtIP as a key substrate by which SIRT6 facilitates DSB processing and homologous recombination. These findings further clarify how SIRT6 promotes genome stability.

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Figures

Fig. 1
Fig. 1
Nicotinamide treatment impairs resection and HR. (A) U2OS cells were untreated (Con) or pre-treated with NA and/or NaB, then exposed to CPT. Cell extracts were analyzed by western blotting (WB). (B) U2OS cells were treated and examined by immunofluorescence (IF). ssDNA was detected with anti-BrdU antibody. (C) HR (gene conversion) was measured (Ref ; also see methods) and data from three experiments are presented as mean +/− S.D. (D) U2OS survival upon CPT treatment in the presence or absence of NA. Results represent three experiments; error bars, S.D. (E) U2OS cells were treated as (A) with 1μM etoposide.
Fig. 2
Fig. 2
SIRT6 promotes resection and HR. (A) SIRT1 and SIRT6 depleted U2OS cells were treated and analysed. Luciferase siRNA (Luc) was used as control. (B) RPA IF staining in U2OS cells after SIRT1/SIRT6 depletion (upper panel). Lower panel: the proportion of γH2AX-positive cells with RPA foci; 1000 cells were counted. (C) IF staining for ssDNA after SIRT6 depletion. (D) Mouse ESCs from WT or Sirt6-knockout (S6−/−) mice were treated and analysed. SIRT6 depletion impairs HR (E), increases sensitivity to CPT (F) and PARP inhibitor (PARPi; G). Data in (E-G) represent averages from three experiments +/−S.D.
Fig. 3
Fig. 3
SIRT6 recruitment to DNA-damage sites and interaction with CtIP. (A) SIRT6 was depleted in U2OS cells stably expressing siRNA-resistant GFP-SIRT6 (WT or enzymatically-inactive H133Y; e and g indicate endogenous, and GFP-tagged SIRT6, respectively). (B) U2OS cells treated as in (A) were analysed by IF. (C) Fractionation of U2OS cells after CPT treatment: S1=cytoplasmic; S2=nuclear soluble; S3=200mM NaCl-extracted chromatin; P=pellet. (D) Recruitment kinetics of GFP-SIRT6 to laser-induced lesions. (E) Purified GFP-SIRT6 (asterisk denotes the band identified as CtIP). (F) Interaction between SIRT6 and CtIP by co-immunoprecipitation from U2OS cells stably transfected with GFP-SIRT6.
Fig. 4
Fig. 4
SIRT6 deacetylates CtIP to promote resection and HR. (A) Constitutive CtIP acetylation is removed by SIRT6 in vitro. (B) GFP-CtIP is acetylated in HEK293 cells and deacetylated upon treatment with CPT (1μM), etoposide (1μM) or IR (10Gy). Immunoprecipitated material and cell extract (CE) were analysed. (C) NA or WOR block GFP-CtIP de-acetylation in HEK293 cells after DNA damage. (D) Endogenous CtIP is acetylated in U2OS cells. A negative-control IP is shown in fig. S12D. (E) SIRT6 mediates CtIP deacetylation upon DNA damage. CtIP acetylation status in U2OS cells stably expressing siRNA-resistant GFP-SIRT6. (F) Identification of CtIP acetylation sites; mutations were introduced into GFP-CtIP and analysed by IP-WB. Mutants: 2KA (K513A+K515A), 2KR (K513R+K515R), 3KR (K432A+K526A+K604A) and 3KR (K432R+K526R+K604R). (G) Mass-spectrometry data for intensity of acetylated CtIP peptides. (H) Non-acetylatable CtIP (3KR) alleviates resection defects caused by SIRT6 depletion. U2OS cells expressing siRNA-resistant GFP-CtIP (WT/3KR), where SIRT6 and CtIP were depleted, were treated with CPT and analysed. (I) U2OS cells treated as in (H) were analysed by IF. (J) CtIP-3KR mutant partially rescues HR defect of SIRT6 depleted cells. Cells contain with vector (Vec), Flag-CtIP-WT or 3KR. Data are from two experiments.
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