Bloom helicase mediates formation of large single–stranded DNA loops during DNA end processing

Chaoyou Xue; Sameer J. Salunkhe; Nozomi Tomimatsu; Ajinkya S. Kawale; Youngho Kwon; Sandeep Burma; Patrick Sung; Eric C. Greene

doi:10.1038/s41467-022-29937-7

Bloom helicase mediates formation of large single–stranded DNA loops during DNA end processing

Chaoyou Xue, Sameer J. Salunkhe, Nozomi Tomimatsu, Ajinkya S. Kawale, Youngho Kwon, Sandeep Burma, Patrick Sung, Eric C. Greene

Resultado de la investigación: Article › revisión exhaustiva

4 Citas (Scopus)

Resumen

Bloom syndrome (BS) is associated with a profoundly increased cancer risk and is caused by mutations in the Bloom helicase (BLM). BLM is involved in the nucleolytic processing of the ends of DNA double–strand breaks (DSBs), to yield long 3′ ssDNA tails that serve as the substrate for break repair by homologous recombination (HR). Here, we use single–molecule imaging to demonstrate that BLM mediates formation of large ssDNA loops during DNA end processing. A BLM mutant lacking the N–terminal domain (NTD) retains vigorous in vitro end processing activity but fails to generate ssDNA loops. This same mutant supports DSB end processing in cells, however, these cells do not form RAD51 DNA repair foci and the processed DSBs are channeled into synthesis–dependent strand annealing (SSA) instead of HR–mediated repair, consistent with a defect in RAD51 filament formation. Together, our results provide insights into BLM functions during homologous recombination.

Idioma original	English (US)
Número de artículo	2248
Publicación	Nature communications
Volumen	13
N.º	1
DOI	https://doi.org/10.1038/s41467-022-29937-7
Estado	Published - dic 2022
Publicado de forma externa	Sí

ASJC Scopus subject areas

General
Physics and Astronomy(all)
Chemistry(all)
Biochemistry, Genetics and Molecular Biology(all)

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title = "Bloom helicase mediates formation of large single–stranded DNA loops during DNA end processing",

abstract = "Bloom syndrome (BS) is associated with a profoundly increased cancer risk and is caused by mutations in the Bloom helicase (BLM). BLM is involved in the nucleolytic processing of the ends of DNA double–strand breaks (DSBs), to yield long 3′ ssDNA tails that serve as the substrate for break repair by homologous recombination (HR). Here, we use single–molecule imaging to demonstrate that BLM mediates formation of large ssDNA loops during DNA end processing. A BLM mutant lacking the N–terminal domain (NTD) retains vigorous in vitro end processing activity but fails to generate ssDNA loops. This same mutant supports DSB end processing in cells, however, these cells do not form RAD51 DNA repair foci and the processed DSBs are channeled into synthesis–dependent strand annealing (SSA) instead of HR–mediated repair, consistent with a defect in RAD51 filament formation. Together, our results provide insights into BLM functions during homologous recombination.",

author = "Chaoyou Xue and Salunkhe, {Sameer J.} and Nozomi Tomimatsu and Kawale, {Ajinkya S.} and Youngho Kwon and Sandeep Burma and Patrick Sung and Greene, {Eric C.}",

note = "Funding Information: We thank Prof. Jeremy Stark (Department of Cancer Genetics and Epigenetics, City of Hope Comprehensive Cancer Center) for reporter assay cell lines. This research was funded by National Institutes of Health grants R01CA236606, R01CA221858 and R35GM118026 (E.C.G.); National Institutes of Health grants R35CA241801, P01CA092584, ES007061, a Gray Foundation Team Science Grant (P.S.); a Wellcome Trust Collaborative Award in Science (Grant. No. 206292/D/17/Z; E.C.G.); and National Institutes of Health Grants R01CA258381 and R01CA246807 (S.B.) and National Aeronautics and Space Administration Award 80NSSC20K0732 (S.B.). P.S. is the recipient of a CPRIT REI Award (RR180029) and holder of the Robert A. Welch Distinguished Chair in Biochemistry (AQ-0012). A.S.K. was the recipient of a CPRIT Postdoctoral Fellowship (RP170345). Publisher Copyright: {\textcopyright} 2022, The Author(s).",

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doi = "10.1038/s41467-022-29937-7",

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AU - Xue, Chaoyou

AU - Salunkhe, Sameer J.

AU - Tomimatsu, Nozomi

AU - Kawale, Ajinkya S.

AU - Kwon, Youngho

AU - Burma, Sandeep

AU - Sung, Patrick

AU - Greene, Eric C.

N1 - Funding Information: We thank Prof. Jeremy Stark (Department of Cancer Genetics and Epigenetics, City of Hope Comprehensive Cancer Center) for reporter assay cell lines. This research was funded by National Institutes of Health grants R01CA236606, R01CA221858 and R35GM118026 (E.C.G.); National Institutes of Health grants R35CA241801, P01CA092584, ES007061, a Gray Foundation Team Science Grant (P.S.); a Wellcome Trust Collaborative Award in Science (Grant. No. 206292/D/17/Z; E.C.G.); and National Institutes of Health Grants R01CA258381 and R01CA246807 (S.B.) and National Aeronautics and Space Administration Award 80NSSC20K0732 (S.B.). P.S. is the recipient of a CPRIT REI Award (RR180029) and holder of the Robert A. Welch Distinguished Chair in Biochemistry (AQ-0012). A.S.K. was the recipient of a CPRIT Postdoctoral Fellowship (RP170345). Publisher Copyright: © 2022, The Author(s).

PY - 2022/12

Y1 - 2022/12

N2 - Bloom syndrome (BS) is associated with a profoundly increased cancer risk and is caused by mutations in the Bloom helicase (BLM). BLM is involved in the nucleolytic processing of the ends of DNA double–strand breaks (DSBs), to yield long 3′ ssDNA tails that serve as the substrate for break repair by homologous recombination (HR). Here, we use single–molecule imaging to demonstrate that BLM mediates formation of large ssDNA loops during DNA end processing. A BLM mutant lacking the N–terminal domain (NTD) retains vigorous in vitro end processing activity but fails to generate ssDNA loops. This same mutant supports DSB end processing in cells, however, these cells do not form RAD51 DNA repair foci and the processed DSBs are channeled into synthesis–dependent strand annealing (SSA) instead of HR–mediated repair, consistent with a defect in RAD51 filament formation. Together, our results provide insights into BLM functions during homologous recombination.

AB - Bloom syndrome (BS) is associated with a profoundly increased cancer risk and is caused by mutations in the Bloom helicase (BLM). BLM is involved in the nucleolytic processing of the ends of DNA double–strand breaks (DSBs), to yield long 3′ ssDNA tails that serve as the substrate for break repair by homologous recombination (HR). Here, we use single–molecule imaging to demonstrate that BLM mediates formation of large ssDNA loops during DNA end processing. A BLM mutant lacking the N–terminal domain (NTD) retains vigorous in vitro end processing activity but fails to generate ssDNA loops. This same mutant supports DSB end processing in cells, however, these cells do not form RAD51 DNA repair foci and the processed DSBs are channeled into synthesis–dependent strand annealing (SSA) instead of HR–mediated repair, consistent with a defect in RAD51 filament formation. Together, our results provide insights into BLM functions during homologous recombination.

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Bloom helicase mediates formation of large single–stranded DNA loops during DNA end processing

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