DNA Repair in<i>Drosophila</i>: Mutagens, Models, and Missing Genes

Sekelsky, Jeff

doi:10.1534/genetics.116.186759

Cited by 109 publications

(134 citation statements)

References 194 publications

(204 reference statements)

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“…Drosophila melanogaster presents an interesting case where the GGR pathway for NER is present but TCR was thought to be missing (11, 12). The lack of fly homologs for genes required for TCR in other organisms — CSA/ERCC8 and CSB/ERCC6 — suggested that the pathway was lost during evolution (11).…”

Section: Introductionmentioning

confidence: 99%

Transcription-coupled repair inDrosophila melanogasteris independent of the mismatch repair pathway

Törmä¹,

Burny²,

Nolte³

et al. 2020

Preprint

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1Transcription-coupled repair (TCR) removes base damage on the transcribed strand of a gene to 2 ensure a quick resumption of transcription. Based on the absence of key enzymes for TCR and 3 empirical evidence, TCR was thought to be missing in Drosophila melanogaster. The recent 4 demonstration of TCR in S2 cells raises the question about the involved genes. Since the 5 mismatch repair (MMR) pathway serves a central role in TCR, at least in Escherichia coli, we 6 studied the mutational signatures in flies with a deletion of the MMR gene spellchecker1 (spel1), 7 a MutS homolog. Whole-genome sequencing of mutation accumulation (MA) lines obtained 7,345 8 new single nucleotide variants (SNVs) and 5,672 short indel mutations, the largest data set from 9 an MA study in D. melanogaster. Based on the observed mutational strand-asymmetries, we 10 conclude that TCR is still active without spel1. The operation of TCR is further confirmed by a 11 negative association between mutation rate and gene expression. Surprisingly, the TCR 12 signatures are detected for introns, but not for exons. We propose that an additional exon-specific 13 repair pathway is masking the signature of TCR. This study presents the first step towards 14 understanding the molecular basis of TCR in Drosophila melanogaster. 15

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Section: Introductionmentioning

confidence: 99%

Transcription-coupled repair inDrosophila melanogasteris independent of the mismatch repair pathway

Törmä¹,

Burny²,

Nolte³

et al. 2020

Preprint

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“…The elevated level of DNA damages under dehydration has been reported for different species, including Diptera [69][70][71]. The damage-induced DNA repair may also intensify recombination due to the upregulation of some common enzymatic pathways [72,73].…”

Section: (C) Desiccation-induced Changes In Recombination Rate and Thmentioning

confidence: 98%

Seasonal changes in recombination rate, crossover interference, and their response to desiccation stress in a natural population ofDrosophila melanogasterfrom India

Rybnikov

Sapielkin

et al. 2020

Preprint

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Environmental seasonality is a potent evolutionary force, capable to maintain polymorphism, promote phenotypic plasticity, and cause bet-hedging. In Drosophila, it has been reported to affect life-history traits, tolerance to abiotic stressors, and immunity. Oscillations in frequencies of alleles underlying fitness-related traits were also documented alongside SNP alleles across genome. Here, we test for seasonal changes in recombination in a natural D. melanogaster population from India using morphological markers of the three major chromosomes. We show that winter flies (collected after the dry season) have significantly higher desiccation tolerance than their autumn counterparts. This difference proved to hold also for hybrids with three independent marker stocks, suggesting its genetic rather than plastic nature. Significant segment-specific changes are documented for recombination rate (in five of 13 intervals) and crossover interference (in five of 16 studied pairs of intervals); both singleand double-crossover rates tended to increase in the winter cohort. The winter flies also display weaker plasticity of recombination characteristics to desiccation. We ascribe the observed differences to indirect selection on recombination caused by directional selection on desiccation tolerance. Our findings suggest that changes in recombination can arise even after a short period of seasonal adaptation (~8-10 generations).

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“…Thanks to CRISPR/Cas technology, generating such mutants has become within the reach of any research laboratory. Indels are readily created by inducing two site-specific double strand breaks (DSBs), thus deleting the intervening genomic sequences ( [4]). With this approach, any gene can be inactivated in most model organisms and the phenotypic consequences assessed.…”

Section: Introductionmentioning

confidence: 99%

Lessons in genome engineering: opportunities, tools and pitfalls

Poernbacher¹,

Crossman

Kurth

et al. 2019

Preprint

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CRISPR/Cas technology allows the creation of double strand breaks and hence loss of function mutations at any location in the genome. This technology is now routine for many organisms and cell lines. Here we describe how CRISPR/Cas can be combined with other DNA manipulation techniques (e.g. homology-based repair, site-specific integration and Cre or FLP-mediated recombination) to create sophisticated tools to measure and manipulate gene activity. In one class of applications, a single site-specific insertion generates a transcriptional reporter, a loss-of function allele, and a tagged allele. In a second class of modifications, essential sequences are deleted and replaced with an integrase site, which serves as a platform for the creation of custom reporters, transcriptional drivers, conditional alleles and regulatory mutations. We describe how these tools and protocols can be implemented easily and efficiently. Importantly, we also highlight unanticipated failures, which serve as cautionary tales, and suggest mitigating measures. Our tools are designed for use in Drosophila but the lessons we draw are likely to be widely relevant. AUTHOR SUMMARYThe genome contains all the information that an organism needs to develop and function throughout its life. One of the goal of genetics is to decipher the role of all the genes (typically several thousands for an animal) present in the genome. One approach is to delete each gene and assay the consequences. Deletion of individual genes is now readily achieved with a technique called CRISPR/Cas9. However, simple genetic deletion provides limited information. Here we describe strains and DNA vectors that streamline the generation of more sophisticated genetic tools. We describe general means of creating alleles (genetic variants) that enable gene activity to be measured and experimentally modulated in space and time. Although the tools we describe are universally applicable, each gene requires special consideration. Based on our experience of successes and failures, we suggest measures to maximise the chances that engineered alleles serve their intended purpose. Although our methods are designed for use in Drosophila, they could be adapted to any organism that is amenable to CRISPR/Cas9 genome modification.

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DNA Repair inDrosophila: Mutagens, Models, and Missing Genes

Cited by 109 publications

References 194 publications

Transcription-coupled repair inDrosophila melanogasteris independent of the mismatch repair pathway

Transcription-coupled repair inDrosophila melanogasteris independent of the mismatch repair pathway

Seasonal changes in recombination rate, crossover interference, and their response to desiccation stress in a natural population ofDrosophila melanogasterfrom India

Lessons in genome engineering: opportunities, tools and pitfalls

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