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Engineered proteins detect spontaneous DNA breakage in human and bacterial cells.

作者信息

Shee Chandan, Cox Ben D, Gu Franklin, Luengas Elizabeth M, Joshi Mohan C, Chiu Li-Ya, Magnan David, Halliday Jennifer A, Frisch Ryan L, Gibson Janet L, Nehring Ralf Bernd, Do Huong G, Hernandez Marcos, Li Lei, Herman Christophe, Hastings P J, Bates David, Harris Reuben S, Miller Kyle M, Rosenberg Susan M

机构信息

Department of Molecular and Human Genetics , Baylor College of Medicine , Houston , United States ; Department of Molecular Virology and Microbiology , Baylor College of Medicine , Houston , United States ; Dan L Duncan Cancer Center, Baylor College of Medicine , Houston , United States ; Department of Biochemistry, Molecular Biology , Baylor College of Medicine , Houston , United States.

出版信息

Elife. 2013 Oct 29;2:e01222. doi: 10.7554/eLife.01222.

DOI:10.7554/eLife.01222
PMID:24171103
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC3809393/
Abstract

Spontaneous DNA breaks instigate genomic changes that fuel cancer and evolution, yet direct quantification of double-strand breaks (DSBs) has been limited. Predominant sources of spontaneous DSBs remain elusive. We report synthetic technology for quantifying DSBs using fluorescent-protein fusions of double-strand DNA end-binding protein, Gam of bacteriophage Mu. In Escherichia coli GamGFP forms foci at chromosomal DSBs and pinpoints their subgenomic locations. Spontaneous DSBs occur mostly one per cell, and correspond with generations, supporting replicative models for spontaneous breakage, and providing the first true breakage rates. In mammalian cells GamGFP-labels laser-induced DSBs antagonized by end-binding protein Ku; co-localizes incompletely with DSB marker 53BP1 suggesting superior DSB-specificity; blocks resection; and demonstrates DNA breakage via APOBEC3A cytosine deaminase. We demonstrate directly that some spontaneous DSBs occur outside of S phase. The data illuminate spontaneous DNA breakage in E. coli and human cells and illustrate the versatility of fluorescent-Gam for interrogation of DSBs in living cells. DOI:http://dx.doi.org/10.7554/eLife.01222.001.

摘要
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/ee6f92b65db8/elife01222f008.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/dcb556a59eef/elife01222f001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/3cc1fe36f639/elife01222fs001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/af725c6db308/elife01222fs002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/a7b873a07ae4/elife01222f002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/81cdd771fe4f/elife01222fs003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/1f3fad30dc5a/elife01222fs004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/0d78ac2508e5/elife01222f003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/18b78786e7f6/elife01222f004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/febb5933c2e8/elife01222fs005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/ea6c1f63e5d5/elife01222f005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/96da158ea4fc/elife01222fs006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/9d7148f5132a/elife01222fs007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/5469b829a59b/elife01222f006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/81b3d7baa92c/elife01222f007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/ee6f92b65db8/elife01222f008.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/dcb556a59eef/elife01222f001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/3cc1fe36f639/elife01222fs001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/af725c6db308/elife01222fs002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/a7b873a07ae4/elife01222f002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/81cdd771fe4f/elife01222fs003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/1f3fad30dc5a/elife01222fs004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/0d78ac2508e5/elife01222f003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/18b78786e7f6/elife01222f004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/febb5933c2e8/elife01222fs005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/ea6c1f63e5d5/elife01222f005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/96da158ea4fc/elife01222fs006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/9d7148f5132a/elife01222fs007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/5469b829a59b/elife01222f006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/81b3d7baa92c/elife01222f007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4d97/3809393/ee6f92b65db8/elife01222f008.jpg

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本文引用的文献

[1]
A new method for high-resolution imaging of Ku foci to decipher mechanisms of DNA double-strand break repair.

J Cell Biol. 2013-7-29

[2]
Evidence for APOBEC3B mutagenesis in multiple human cancers.

Nat Genet. 2013-7-14

[3]
R-loops and nicks initiate DNA breakage and genome instability in non-growing Escherichia coli.

Nat Commun. 2013

[4]
53BP1 is a reader of the DNA-damage-induced H2A Lys 15 ubiquitin mark.

Nature. 2013-6-12

[5]
Standard fluorescent imaging of live cells is highly genotoxic.

Cytometry A. 2013-5-6

[6]
DNA deaminases induce break-associated mutation showers with implication of APOBEC3B and 3A in breast cancer kataegis.

Elife. 2013-4-16

[7]
Protein-DNA complexes are the primary sources of replication fork pausing in Escherichia coli.

Proc Natl Acad Sci U S A. 2013-4-15

[8]
APOBEC3B is an enzymatic source of mutation in breast cancer.

Nature. 2013-2-6

[9]
Identity and function of a large gene network underlying mutagenic repair of DNA breaks.

Science. 2012-12-7

[10]
APOBEC3B can impair genomic stability by inducing base substitutions in genomic DNA in human cells.

Sci Rep. 2012-11-13

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