Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

Patrick M. Grierson; Samir Acharya; Joanna Groden

doi:10.1016/j.mrfmmm.2012.12.002

Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

Patrick M. Grierson, Samir Acharya, Joanna Groden

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Abstract

Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.

Original language	English
Pages (from-to)	89-96
Number of pages	8
Journal	Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis
Volume	743-744
DOIs	https://doi.org/10.1016/j.mrfmmm.2012.12.002
State	Published - Mar 2013

Keywords

BLM helicase
Bloom's syndrome
DNA topoisomerase I
Nucleolus
RNA polymerase I
RRNA transcription

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10.1016/j.mrfmmm.2012.12.002

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@article{279362e111294f9481514de559e4a07c,

title = "Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription",

abstract = "Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.",

keywords = "BLM helicase, Bloom's syndrome, DNA topoisomerase I, Nucleolus, RNA polymerase I, RRNA transcription",

author = "Grierson, {Patrick M.} and Samir Acharya and Joanna Groden",

note = "Funding Information: The authors thank Dr. April Sandy Gocha for help with experiments associated with the Bloom's syndrome cell line, GM8505; Jeremy Kiersey for help in purification of BLM K695E . This work was supported by the National Institutes of Health [ CA117898 to J.G.]; The Ohio State University Pelotonia Fellowship Program to [P.G.]; and The Ohio State University Medical Scientist Training Program (MSTP) to [P.G.]. The project described was supported by Award Number 8UL1TR000090-05 , 8KL2TR000112-05 , and 8TL1TR000091-05 from the National Center For Advancing Translational Sciences . The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Center For Advancing Translational Sciences or the National Institutes of Health. ",

year = "2013",

month = mar,

doi = "10.1016/j.mrfmmm.2012.12.002",

language = "English",

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pages = "89--96",

journal = "Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis",

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AU - Grierson, Patrick M.

AU - Acharya, Samir

AU - Groden, Joanna

N1 - Funding Information: The authors thank Dr. April Sandy Gocha for help with experiments associated with the Bloom's syndrome cell line, GM8505; Jeremy Kiersey for help in purification of BLM K695E . This work was supported by the National Institutes of Health [ CA117898 to J.G.]; The Ohio State University Pelotonia Fellowship Program to [P.G.]; and The Ohio State University Medical Scientist Training Program (MSTP) to [P.G.]. The project described was supported by Award Number 8UL1TR000090-05 , 8KL2TR000112-05 , and 8TL1TR000091-05 from the National Center For Advancing Translational Sciences . The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Center For Advancing Translational Sciences or the National Institutes of Health.

PY - 2013/3

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N2 - Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.

AB - Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.

KW - BLM helicase

KW - Bloom's syndrome

KW - DNA topoisomerase I

KW - Nucleolus

KW - RNA polymerase I

KW - RRNA transcription

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U2 - 10.1016/j.mrfmmm.2012.12.002

DO - 10.1016/j.mrfmmm.2012.12.002

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C2 - 23261817

AN - SCOPUS:84877927942

SN - 0027-5107

VL - 743-744

SP - 89

EP - 96

JO - Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis

JF - Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis

ER -

Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

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