Strand Annealing and Terminal Transferase Activities of a B-family DNA Polymerase

Biochemistry ◽  
2011 ◽  
Vol 50 (23) ◽  
pp. 5379-5390 ◽  
Author(s):  
Zhongfeng Zuo ◽  
Hsiang-Kai Lin ◽  
Michael A. Trakselis
Keyword(s):  
Dna Polymerase ◽  
Terminal Transferase ◽  
Strand Annealing

scholarly journals Limited terminal transferase in human DNA polymerase   defines the required balance between accuracy and efficiency in NHEJ

2009 ◽  
Vol 106 (38) ◽  
pp. 16203-16208 ◽  
Author(s):  
P. Andrade ◽  
M. J. Martin ◽  
R. Juarez ◽  
F. Lopez de Saro ◽  
L. Blanco
Keyword(s):  
Dna Polymerase ◽  
Terminal Transferase ◽  
Human Dna

scholarly journals Author response: Srs2 promotes synthesis-dependent strand annealing by disrupting DNA polymerase δ-extending D-loops

2017 ◽  
Author(s):  
Jie Liu ◽  
Christopher Ede ◽  
William D Wright ◽  
Steven K Gore ◽  
Shirin S Jenkins ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Author Response ◽  
Dna Polymerase Δ ◽  
Strand Annealing

scholarly journals DNA Polymerase δ Is Preferentially Recruited during Homologous Recombination To Promote Heteroduplex DNA Extension

2007 ◽  
Vol 28 (4) ◽  
pp. 1373-1382 ◽  
Author(s):  
Laurent Maloisel ◽  
Francis Fabre ◽  
Serge Gangloff
Keyword(s):  
Homologous Recombination ◽  
Gene Conversion ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Strand Break ◽  
Gene Encoding ◽  
Repair Synthesis ◽  
Dna Polymerase Δ ◽  
Strand Annealing ◽  
Gene Conversion Tract

ABSTRACT DNA polymerases play a central role during homologous recombination (HR), but the identity of the enzyme(s) implicated remains elusive. The pol3-ct allele of the gene encoding the catalytic subunit of DNA polymerase δ (Polδ) has highlighted a role for this polymerase in meiotic HR. We now address the ubiquitous role of Polδ during HR in somatic cells. We find that pol3-ct affects gene conversion tract length during mitotic recombination whether the event is initiated by single-strand gaps following UV irradiation or by site-specific double-strand breaks. We show that the pol3-ct effects on gene conversion are completely independent of mismatch repair, indicating that shorter gene conversion tracts in pol3-ct correspond to shorter extensions of primed DNA synthesis. Interestingly, we find that shorter repair tracts do not favor synthesis-dependent strand annealing at the expense of double-strand-break repair. Finally, we show that the DNA polymerases that have been previously suspected to mediate HR repair synthesis (Polε and Polη) do not affect gene conversion during induced HR, including in the pol3-ct background. Our results argue strongly for the preferential recruitment of Polδ during HR.

scholarly journals A Rad51-Independent Pathway Promotes Single-Strand Template Repair in Gene Editing

2020 ◽  
Author(s):  
Danielle N. Gallagher ◽  
Nhung Pham ◽  
Annie M. Tsai ◽  
Abigail N. Janto ◽  
Jihyun Choi ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Gene Editing ◽  
Fold Increase ◽  
Single Strand ◽  
Critical Function ◽  
Strand Breaks ◽  
Independent Events ◽  
Single Strand Annealing ◽  
Strand Annealing

AbstractThe Rad51/RecA family of recombinases perform a critical function in typical repair of double-strand breaks (DSBs): strand invasion of a resected DSB end into a homologous double-stranded DNA (dsDNA) template sequence to facilitate repair. However, repair of a DSB using single stranded DNA (ssDNA) as a template, a common method of CRISPR/Cas9-mediated gene editing, is Rad51-independent. We have analyzed the genetic requirements for these Rad51-independent events in Saccharomyces cerevisiae in two different assays. Gene editing events were carried out either by creating a DSB with the site-specific HO endonuclease and repairing the DSB with 80-nt single-stranded oligonucleotides (ssODNs) or by using a bacterial retron system that produces ssDNA templates in vivo in combination with DSBs created by Cas9. We show that single strand template repair (SSTR), is dependent on Rad52, Rad59, Srs2 and the Mre11-Rad50-Xrs2 (MRX) complex, but not Rad51, Rad54 or Rad55. Srs2 acts to prevent overloading of Rad51 on the ssDNA filament, whereas Rad59 appears to alleviate the inhibition of Rad51 on Rad52’s strand annealing activity; thus, deletion of RAD51 suppresses both the srs2Δ and rad59Δ phenotypes. This same suppression by rad51Δ of rad59Δ is found in another DSB repair pathway, single strand annealing (SSA). In contrast, gene targeting using an 80-bp dsDNA template of the same sequence is Rad51-dependent. We also examined SSTR events in which the ssODN carried several mismatches. In the absence of the mismatch repair protein, Msh2, we found that the fate of mismatches carried on the ssDNA template are very different at the 3’ end, which can anneal directly to the resected DSB end, compared to the 5’ end. We also find that DNA polymerase Polδ’s 3’ to 5’ proofreading activity frequently excises a mismatch close to the 3’ end of the template, similar to its removal of heterologies close to the 3’ invading end of the DSB. We further report that SSTR is accompanied by a 600-fold increase in mutations in a region adjacent to the sequences directly undergoing repair. These DNA polymerase ζ-dependent mutations may compromise the accuracy of gene editing.

scholarly journals Control of DNA polymerase gp5 chain substitution by DNA double strand annealing pressure

2021 ◽  
Vol 70 (15) ◽  
pp. 158701-158701
Author(s):  
Jia Qi ◽  
◽  
Fan Qin-Kai ◽  
Hou Wen-Qing ◽  
Yang Chen-Guang ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Annealing Pressure ◽  
Strand Annealing
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