Faculty Opinions recommendation of BRCA1/BARD1 orthologs required for DNA repair in Caenorhabditis elegans.

AbstractFanconi Anemia is a rare genetic disease associated with DNA repair defects, congenital abnormalities and infertility. Most of FA pathway is evolutionary conserved, allowing dissection and mechanistic studies in simpler model systems such as Caenorhabditis elegans. In the present study, we employed C. elegans to better understand the role of FA group D2 (FANCD2) protein in vivo, a key player in promoting genome stability. We report that localization of FCD-2/FANCD2 is dynamic during meiotic prophase I and requires its heterodimeric partner FNCI-1/FANCI. Strikingly, we found that FCD-2 recruitment depends on SPO-11-induced double-strand breaks (DSBs) but not RAD-51-mediated strand invasion. Furthermore, exposure to DNA damage-inducing agents boosts FCD-2 recruitment on the chromatin. Finally, analysis of genetic interaction between FCD-2 and BRC-1 (the C. elegans orthologue of mammalian BRCA1) supports a role for these proteins in different DSB repair pathways. Collectively, we showed a direct involvement of FCD-2 at DSBs and speculate on its function in driving meiotic DNA repair.

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A DNA repair protein and histone methyltransferase interact to promote genome stability in the Caenorhabditis elegans germ line

PLoS Genetics ◽

10.1371/journal.pgen.1007992 ◽

2019 ◽

Vol 15 (2) ◽

pp. e1007992 ◽

Cited By ~ 11

Author(s):

Bing Yang ◽

Xia Xu ◽

Logan Russell ◽

Matthew T. Sullenberger ◽

Judith L. Yanowitz ◽

...

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans ◽

Genome Stability ◽

Germ Line ◽

Histone Methyltransferase ◽

Repair Protein ◽

Dna Repair Protein

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Functional characterization of the Caenorhabditis elegans DNA repair enzyme APN-1

DNA Repair ◽

10.1016/j.dnarep.2012.06.009 ◽

2012 ◽

Vol 11 (10) ◽

pp. 811-822 ◽

Cited By ~ 12

Author(s):

Xiaoming Yang ◽

Jinjiang Fan ◽

Alexander A. Ishchenko ◽

Devang Patel ◽

Murat K. Saparbaev ◽

...

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans ◽

Functional Characterization ◽

Repair Enzyme

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Caenorhabditis elegans dna-2 is involved in DNA repair and is essential for germ-line development

FEBS Letters ◽

10.1016/s0014-5793(03)01243-2 ◽

2003 ◽

Vol 555 (2) ◽

pp. 250-256 ◽

Cited By ~ 9

Author(s):

Myon Hee Lee ◽

Sung Min Han ◽

Ji Won Han ◽

Yun Mi Kim ◽

Joohong Ahnn ◽

...

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans ◽

Germ Line ◽

Dna 2

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BRCA1/BARD1 Orthologs Required for DNA Repair in Caenorhabditis elegans

Current Biology ◽

10.1016/j.cub.2003.11.029 ◽

2004 ◽

Vol 14 (1) ◽

pp. 33-39 ◽

Cited By ~ 96

Author(s):

Simon J. Boulton ◽

Julie S. Martin ◽

Jolanta Polanowska ◽

David E. Hill ◽

Anton Gartner ◽

...

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans

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An efficient genome editing strategy to generate putative null mutants in Caenorhabditis elegans using CRISPR/Cas9

10.1101/391243 ◽

2018 ◽

Cited By ~ 1

Author(s):

Han Wang ◽

Heenam Park ◽

Jonathan Liu ◽

Paul W. Sternberg

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans ◽

Genome Editing ◽

Gene Function ◽

Unique Sequence ◽

Target Site ◽

Null Alleles ◽

Wild Type ◽

C Elegans ◽

Null Mutants

AbstractNull mutants are essential for analyzing gene function. Here, we describe a simple and efficient method to generate Caenorhabditis elegans null mutants using CRISPR/Cas9 and short single stranded DNA oligo repair templates to insert a universal 43-nucleotide-long stop knock-in (STOP-IN) cassette into the early exons of target genes. This cassette has stop codons in all three reading frames and leads to frameshifts, which will generate putative null mutations regardless of the reading frame of the insertion position in exons. The STOP-IN cassette also contains an exogenous Cas9 target site that allows further genome editing and provides a unique sequence that simplifies the identification of successful insertion events via PCR. As a proof of concept, we inserted the STOP-IN cassette right at a Cas9 target site in aex-2 to generate new putative null alleles by injecting preassembled Cas9 ribonucleoprotein and a short synthetic single stranded DNA repair template containing the STOP-IN cassette and two 35-nucleotide-long homology arms identical to the sequences flanking the Cas9 cut site. We showed that these new aex-2 alleles phenocopied an existing loss-of-function allele of aex-2. We further showed that the new aex-2 null alleles could be reverted back to the wild-type sequence by targeting exogenous Cas9 cut site included in the STOP-IN cassette and providing a single stranded wild-type DNA repair oligo. We applied our STOP-IN method to generate new putative null mutants for additional 20 genes, including three pharyngeal muscle-specific genes (clik-1, clik-2, and clik-3), and reported a high insertion rate (46%) based on the animals we screened. We showed that null mutations of clik-2 cause recessive lethality with a severe pumping defect and clik-3 null mutants have a mild pumping defect, while clik-1 is dispensable for pumping. We expect that the knock-in method using the STOP-IN cassette will facilitate the generation of new null mutants to understand gene function in C. elegans and other genetic model organisms.SummaryWe report a simple and efficient CRISPR/Cas9 genome editing strategy to generate putative null C. elegans mutants by inserting a small universal stop knock-in (STOP-IN) cassette with stop codons in three frames and frameshifts. The strategy is cloning-free, with the mixture consisting of preassembled Cas9 ribonucleoprotein and single stranded repair DNA oligos directly injected into gonads of adult C. elegans. The universal STOP-IN cassette also contains a unique sequence that simplifies detection of successful knock-in events via PCR and an exogenous Cas9 target sequence that allows further genome editing.

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The Base Excision Repair Pathway in the Nematode Caenorhabditis elegans

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.598860 ◽

2020 ◽

Vol 8 ◽

Author(s):

Noha Elsakrmy ◽

Qiu-Mei Zhang-Akiyama ◽

Dindial Ramotar

Keyword(s):

Dna Repair ◽

Caenorhabditis Elegans ◽

Dna Polymerase ◽

Base Excision Repair ◽

Excision Repair ◽

Dna Glycosylases ◽

Repair Synthesis ◽

C Elegans ◽

Base Excision ◽

Dna Repair Synthesis

Exogenous and endogenous damage to the DNA is inevitable. Several DNA repair pathways including base excision, nucleotide excision, mismatch, homologous and non-homologous recombinations are conserved across all organisms to faithfully maintain the integrity of the genome. The base excision repair (BER) pathway functions to repair single-base DNA lesions and during the process creates the premutagenic apurinic/apyrimidinic (AP) sites. In this review, we discuss the components of the BER pathway in the nematode Caenorhabditis elegans and delineate the different phenotypes caused by the deletion or the knockdown of the respective DNA repair gene, as well as the implications. To date, two DNA glycosylases have been identified in C. elegans, the monofunctional uracil DNA glycosylase-1 (UNG-1) and the bifunctional endonuclease III-1 (NTH-1) with associated AP lyase activity. In addition, the animal possesses two AP endonucleases belonging to the exonuclease-3 and endonuclease IV families and in C. elegans these enzymes are called EXO-3 and APN-1, respectively. In mammalian cells, the DNA polymerase, Pol beta, that is required to reinsert the correct bases for DNA repair synthesis is not found in the genome of C. elegans and the evidence indicates that this role could be substituted by DNA polymerase theta (POLQ), which is known to perform a function in the microhomology-mediated end-joining pathway in human cells. The phenotypes observed by the C. elegans mutant strains of the BER pathway raised many challenging questions including the possibility that the DNA glycosylases may have broader functional roles, as discuss in this review.

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