title: Synthesis-dependent microhomology-mediated end joining accounts for multiple types of repair junctions.
abstract: Ku or DNA ligase 4-independent alternative end joining (alt-EJ) repair of DNA double-strand breaks (DSBs) frequently correlates with increased junctional microhomology. However, alt-EJ also produces junctions without microhomology (apparent blunt joins), and the exact role of microhomology in both alt-EJ and classical non-homologous end joining (NHEJ) remains unclear. To better understand the degree to which alt-EJ depends on annealing at pre-existing microhomologies, we examined inaccurate repair of an I-SceI DSB lacking nearby microhomologies of greater than four nucleotides in Drosophila. Lig4 deficiency affected neither frequency nor length of junctional microhomology, but significantly increased insertion frequency. Many insertions appeared to be templated. Based on sequence analysis of repair junctions, we propose a model of synthesis-dependent microhomology-mediated end joining (SD-MMEJ), in which de novo synthesis by an accurate non-processive DNA polymerase creates microhomology. Repair junctions with apparent blunt joins, junctional microhomologies and short indels (deletion with insertion) are often considered to reflect different repair mechanisms. However, a majority of each type had structures consistent with the predictions of our SD-MMEJ model. This suggests that a single underlying mechanism could be responsible for all three repair product types. Genetic analysis indicates that SD-MMEJ is Ku70, Lig4 and Rad51-independent but impaired in mus308 (POLQ) mutants.
authors.affiliation: Department of Biology, Tufts University, Medford, MA 02155, USA.
authors.forename: Amy Marie
authors.initials: AM
authors.lastname: Yu
authors.forename: Mitch
authors.initials: M
authors.lastname: McVey
mesh_terms: D000818:Animals; D004247:DNA / Q000096:biosynthesis / Q000737:chemistry; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004331:Drosophila melanogaster / Q000235:genetics; D005838:Genotype; D008957:Models, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D017384:Sequence Deletion; D012689:Sequence Homology, Nucleic Acid; D013698:Templates, Genetic
chemical_list: D004247:DNA
references.citation: Science. 1993 Apr 9;260(5105):207-10
references.pmid: 8469973
references.citation: Trends Genet. 2000 Jun;16(6):276-7
references.pmid: 10827456
references.citation: J Exp Med. 2010 Feb 15;207(2):417-27
references.pmid: 20142431
references.citation: Environ Mol Mutagen. 2003;42(4):288-98
references.pmid: 14673874
references.citation: DNA Repair (Amst). 2004 Jan 5;3(1):13-21
references.pmid: 14697755
references.citation: Genetics. 2007 Sep;177(1):63-77
references.pmid: 17660539
references.citation: Nat Struct Mol Biol. 2009 Aug;16(8):819-24
references.pmid: 19633668
references.citation: DNA Repair (Amst). 2005 Aug 15;4(9):1038-46
references.pmid: 15935739
references.citation: Cell Mol Life Sci. 2009 Mar;66(6):1039-56
references.pmid: 19153654
references.citation: Genes Dev. 2008 Feb 15;22(4):411-5
references.pmid: 18281457
references.citation: Genetics. 2003 Dec;165(4):1831-42
references.pmid: 14704169
references.citation: Nature. 2007 Sep 27;449(7161):483-6
references.pmid: 17898768
references.citation: Immunity. 1997 Jul;7(1):37-47
references.pmid: 9252118
references.citation: Mutat Res. 2004 Oct 4;554(1-2):351-64
references.pmid: 15450431
references.citation: Genetics. 2004 Jun;167(2):699-705
references.pmid: 15238522
references.citation: Mol Cell Biol. 1996 Oct;16(10):5764-71
references.pmid: 8816490
references.citation: Science. 2007 Oct 19;318(5849):456-9
references.pmid: 17947582
references.citation: Proc Natl Acad Sci U S A. 1996 Aug 20;93(17):8929-33
references.pmid: 8799130
references.citation: EMBO J. 2007 Feb 21;26(4):1010-23
references.pmid: 17290226
references.citation: EMBO J. 2003 Nov 3;22(21):5863-74
references.pmid: 14592983
references.citation: Mol Cell Biol. 1996 May;16(5):2164-73
references.pmid: 8628283
references.citation: Blood. 2000 Jun 1;95(11):3520-9
references.pmid: 10828038
references.citation: Biochem J. 2009 Feb 1;417(3):639-50
references.pmid: 19133841
references.citation: Oncogene. 2005 Mar 3;24(10):1663-72
references.pmid: 15688024
references.citation: Proc Natl Acad Sci U S A. 2010 Feb 16;107(7):3034-9
references.pmid: 20133803
references.citation: Genetics. 1993 Sep;135(1):81-95
references.pmid: 8224830
references.citation: Nat Struct Mol Biol. 2009 Aug;16(8):814-8
references.pmid: 19633669
references.citation: Mol Cell Biol. 2003 Dec;23(23):8820-8
references.pmid: 14612421
references.citation: DNA Repair (Amst). 2008 Jan 1;7(1):67-76
references.pmid: 17881298
references.citation: Genetics. 2007 Aug;176(4):2003-14
references.pmid: 17565964
references.citation: Blood. 2010 Mar 18;115(11):2214-9
references.pmid: 19965626
references.citation: Cancer Res. 2001 Feb 15;61(4):1629-36
references.pmid: 11245476
references.citation: Mol Cell Biol. 2005 Feb;25(3):896-906
references.pmid: 15657419
references.citation: Genetics. 2006 Feb;172(2):1055-68
references.pmid: 16299390
references.citation: Philos Trans R Soc Lond B Biol Sci. 2009 Mar 12;364(1517):653-65
references.pmid: 19008195
references.citation: Nucleic Acids Res. 2001 Oct 1;29(19):3975-81
references.pmid: 11574679
references.citation: Mol Cell. 2006 May 19;22(4):511-9
references.pmid: 16713581
references.citation: J Biol Chem. 2004 Dec 31;279(53):55117-26
references.pmid: 15498778
references.citation: Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6409-14
references.pmid: 10823903
references.citation: Cancer Res. 1996 May 1;56(9):2130-6
references.pmid: 8616861
references.citation: Trends Genet. 2008 Nov;24(11):529-38
references.pmid: 18809224
references.citation: Proc Natl Acad Sci U S A. 1982 Jul;79(13):4128-32
references.pmid: 7051004
references.citation: Cancer Res. 2005 May 15;65(10):4020-30
references.pmid: 15899791
references.citation: Proc Natl Acad Sci U S A. 2007 Dec 26;104(52):20902-7
references.pmid: 18093953
references.citation: Nature. 2007 Sep 27;449(7161):478-82
references.pmid: 17713479
references.citation: Genetics. 2004 Dec;168(4):2067-76
references.pmid: 15611176
references.citation: Cancer Res. 2003 Jan 1;63(1):22-4
references.pmid: 12517771
references.citation: Nucleic Acids Res. 1997 Nov 15;25(22):4650-7
references.pmid: 9358178
references.citation: J Biol Chem. 1996 Jun 14;271(24):14405-11
references.pmid: 8662903
references.citation: Cell. 2007 Dec 28;131(7):1235-47
references.pmid: 18160035
references.citation: EMBO J. 2002 Jun 17;21(12):3192-200
references.pmid: 12065431
references.citation: Hum Mol Genet. 2008 Sep 15;17(18):2910-8
references.pmid: 18632684
references.citation: Int J Cancer. 2004 Mar;109(1):9-16
references.pmid: 14735462
references.citation: J Mol Biol. 1998 Feb 27;276(3):559-69
references.pmid: 9551097
references.citation: Genes Cells. 2006 Aug;11(8):935-48
references.pmid: 16866876
references.citation: Subcell Biochem. 2010;50:279-96
references.pmid: 20012587
references.citation: Genetics. 2004 May;167(1):203-6
references.pmid: 15166147
references.citation: Nucleic Acids Res. 2008 May;36(8):2561-9
references.pmid: 18332040
references.citation: Cell Res. 2008 Jan;18(1):125-33
references.pmid: 18087292
references.citation: PLoS Genet. 2007 Apr 13;3(4):e50
references.pmid: 17432935