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Article: Functions of the Bloom Syndrome Helicase N-terminal Intrinsically Disordered Region.

Bereda, Colleen C / Dewey, Evan B / Nasr, Mohamed A / Sekelsky, Jeff

bioRxiv : the preprint server for biology

2024

Abstract: Bloom Syndrome helicase (Blm) is a RecQ family helicase involved in DNA repair, cell-cycle progression, and development. Pathogenic variants in ... ...

Abstract	Bloom Syndrome helicase (Blm) is a RecQ family helicase involved in DNA repair, cell-cycle progression, and development. Pathogenic variants in human
Language	English
Publishing date	2024-04-15
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.04.12.589165
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Article: Centromere-Proximal Suppression of Meiotic Crossovers in

Pazhayam, Nila M / Frazier, Leah K / Sekelsky, Jeff

bioRxiv : the preprint server for biology

2023

Abstract: Accurate segregation of homologous chromosomes during meiosis depends on both the presence and regulated placement of crossovers (COs). The centromere effect (CE), or CO exclusion in pericentromeric regions of the chromosome, is a meiotic CO patterning ... ...

Abstract	Accurate segregation of homologous chromosomes during meiosis depends on both the presence and regulated placement of crossovers (COs). The centromere effect (CE), or CO exclusion in pericentromeric regions of the chromosome, is a meiotic CO patterning phenomenon that helps prevent nondisjunction (NDJ), thereby protecting against chromosomal disorders and other meiotic defects. Despite being identified nearly a century ago, the mechanisms behind this fundamental cellular process remain unknown, with most studies of the
Language	English
Publishing date	2023-10-20
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.10.17.562696
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Article ; Online: Centromere-proximal suppression of meiotic crossovers in Drosophila is robust to changes in centromere number, repetitive DNA content, and centromere-clustering.

Pazhayam, Nila M / Frazier, Leah K / Sekelsky, Jeff

Genetics

2023 Volume 226, Issue 3

Abstract: Accurate segregation of homologous chromosomes during meiosis depends on both the presence and the regulated placement of crossovers (COs). The centromere effect, or CO exclusion in pericentromeric regions of the chromosome, is a meiotic CO patterning ... ...

Abstract	Accurate segregation of homologous chromosomes during meiosis depends on both the presence and the regulated placement of crossovers (COs). The centromere effect, or CO exclusion in pericentromeric regions of the chromosome, is a meiotic CO patterning phenomenon that helps prevent nondisjunction, thereby protecting against chromosomal disorders and other meiotic defects. Despite being identified nearly a century ago, the mechanisms behind this fundamental cellular process remain unknown, with most studies of the Drosophila centromere effect focusing on local influences of the centromere and pericentric heterochromatin. In this study, we sought to investigate whether dosage changes in centromere number and repetitive DNA content affect the strength of the centromere effect, using phenotypic recombination mapping. Additionally, we studied the effects of repetitive DNA function on centromere effect strength using satellite DNA-binding protein mutants displaying defective centromere-clustering in meiotic nuclei. Despite what previous studies suggest, our results show that the Drosophila centromere effect is robust to changes in centromere number, repetitive DNA content, as well as repetitive DNA function. Our study suggests that the centromere effect is unlikely to be spatially controlled, providing novel insight into the mechanisms behind the Drosophila centromere effect.
MeSH term(s)	Animals ; Drosophila/genetics ; Drosophila/metabolism ; Centromere/genetics ; Centromere/metabolism ; Drosophila Proteins/genetics ; Drosophila Proteins/metabolism ; Meiosis/genetics ; DNA ; Chromosome Segregation
Chemical Substances	Drosophila Proteins ; DNA (9007-49-2)
Language	English
Publishing date	2023-12-27
Publishing country	United States
Document type	Journal Article
ZDB-ID	2167-2
ISSN	1943-2631 ; 0016-6731
ISSN (online)	1943-2631
ISSN	0016-6731
DOI	10.1093/genetics/iyad216
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Article ; Online: DNA Repair in Drosophila: Mutagens, Models, and Missing Genes.

Sekelsky, Jeff

Genetics

2017 Volume 205, Issue 2, Page(s) 471–490

Abstract: The numerous processes that damage DNA are counterbalanced by a complex network of repair pathways that, collectively, can mend diverse types of damage. Insights into these pathways have come from studies in many different organisms, including Drosophila ...

Abstract	The numerous processes that damage DNA are counterbalanced by a complex network of repair pathways that, collectively, can mend diverse types of damage. Insights into these pathways have come from studies in many different organisms, including Drosophila melanogaster Indeed, the first ideas about chromosome and gene repair grew out of Drosophila research on the properties of mutations produced by ionizing radiation and mustard gas. Numerous methods have been developed to take advantage of Drosophila genetic tools to elucidate repair processes in whole animals, organs, tissues, and cells. These studies have led to the discovery of key DNA repair pathways, including synthesis-dependent strand annealing, and DNA polymerase theta-mediated end joining. Drosophila appear to utilize other major repair pathways as well, such as base excision repair, nucleotide excision repair, mismatch repair, and interstrand crosslink repair. In a surprising number of cases, however, DNA repair genes whose products play important roles in these pathways in other organisms are missing from the Drosophila genome, raising interesting questions for continued investigations.
MeSH term(s)	Animals ; DNA Repair ; DNA Repair Enzymes/genetics ; Drosophila/genetics ; Drosophila Proteins/genetics ; Models, Genetic ; Mutagenesis
Chemical Substances	Drosophila Proteins ; DNA Repair Enzymes (EC 6.5.1.-)
Language	English
Publishing date	2017-01-30
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	2167-2
ISSN	1943-2631 ; 0016-6731
ISSN (online)	1943-2631
ISSN	0016-6731
DOI	10.1534/genetics.116.186759
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Article ; Online: ATM/Chk2 and ATR/Chk1 Pathways Respond to DNA Damage Induced by Movento

González-Marín, Berenyce / Calderón-Segura, María Elena / Sekelsky, Jeff

Toxics

2023 Volume 11, Issue 9

Abstract: DNA damage response (DDR) pathways in keto-enol genotoxicity have not been characterized, and few studies have reported genotoxic effects in non-target organisms. The present study shows that concentrations of 11.2, 22.4, 37.3 mg/L of ... ...

Abstract	DNA damage response (DDR) pathways in keto-enol genotoxicity have not been characterized, and few studies have reported genotoxic effects in non-target organisms. The present study shows that concentrations of 11.2, 22.4, 37.3 mg/L of Movento
Language	English
Publishing date	2023-09-06
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2733883-6
ISSN	2305-6304 ; 2305-6304
ISSN (online)	2305-6304
ISSN	2305-6304
DOI	10.3390/toxics11090754
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Article: Meiotic Crossover Patterning.

Pazhayam, Nila M / Turcotte, Carolyn A / Sekelsky, Jeff

Frontiers in cell and developmental biology

2021 Volume 9, Page(s) 681123

Abstract: Proper number and placement of meiotic crossovers is vital to chromosome segregation, with failures in normal crossover distribution often resulting in aneuploidy and infertility. Meiotic crossovers are formed via homologous repair of programmed double- ... ...

Abstract	Proper number and placement of meiotic crossovers is vital to chromosome segregation, with failures in normal crossover distribution often resulting in aneuploidy and infertility. Meiotic crossovers are formed via homologous repair of programmed double-strand breaks (DSBs). Although DSBs occur throughout the genome, crossover placement is intricately patterned, as observed first in early genetic studies by Muller and Sturtevant. Three types of patterning events have been identified. Interference, first described by Sturtevant in 1915, is a phenomenon in which crossovers on the same chromosome do not occur near one another. Assurance, initially identified by Owen in 1949, describes the phenomenon in which a minimum of one crossover is formed per chromosome pair. Suppression, first observed by Beadle in 1932, dictates that crossovers do not occur in regions surrounding the centromere and telomeres. The mechanisms behind crossover patterning remain largely unknown, and key players appear to act at all scales, from the DNA level to inter-chromosome interactions. There is also considerable overlap between the known players that drive each patterning phenomenon. In this review we discuss the history of studies of crossover patterning, developments in methods used in the field, and our current understanding of the interplay between patterning phenomena.
Language	English
Publishing date	2021-07-22
Publishing country	Switzerland
Document type	Journal Article ; Review
ZDB-ID	2737824-X
ISSN	2296-634X
ISSN	2296-634X
DOI	10.3389/fcell.2021.681123
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Article ; Online: DNA polymerase theta suppresses mitotic crossing over.

Carvajal-Garcia, Juan / Crown, K Nicole / Ramsden, Dale A / Sekelsky, Jeff

PLoS genetics

2021 Volume 17, Issue 3, Page(s) e1009267

Abstract: Polymerase theta-mediated end joining (TMEJ) is a chromosome break repair pathway that is able to rescue the lethality associated with the loss of proteins involved in early steps in homologous recombination (e.g., BRCA1/2). This is due to the ability of ...

Abstract	Polymerase theta-mediated end joining (TMEJ) is a chromosome break repair pathway that is able to rescue the lethality associated with the loss of proteins involved in early steps in homologous recombination (e.g., BRCA1/2). This is due to the ability of polymerase theta (Pol θ) to use resected, 3' single stranded DNA tails to repair chromosome breaks. These resected DNA tails are also the starting substrate for homologous recombination. However, it remains unknown if TMEJ can compensate for the loss of proteins involved in more downstream steps during homologous recombination. Here we show that the Holliday junction resolvases SLX4 and GEN1 are required for viability in the absence of Pol θ in Drosophila melanogaster, and lack of all three proteins results in high levels of apoptosis. Flies deficient in Pol θ and SLX4 are extremely sensitive to DNA damaging agents, and mammalian cells require either Pol θ or SLX4 to survive. Our results suggest that TMEJ and Holliday junction formation/resolution share a common DNA substrate, likely a homologous recombination intermediate, that when left unrepaired leads to cell death. One major consequence of Holliday junction resolution by SLX4 and GEN1 is cancer-causing loss of heterozygosity due to mitotic crossing over. We measured mitotic crossovers in flies after a Cas9-induced chromosome break, and observed that this mutagenic form of repair is increased in the absence of Pol θ. This demonstrates that TMEJ can function upstream of the Holiday junction resolvases to protect cells from loss of heterozygosity. Our work argues that Pol θ can thus compensate for the loss of the Holliday junction resolvases by using homologous recombination intermediates, suppressing mitotic crossing over and preserving the genomic stability of cells.
MeSH term(s)	Animals ; Apoptosis/genetics ; BRCA2 Protein/genetics ; Crossing Over, Genetic ; DNA End-Joining Repair ; DNA-Directed DNA Polymerase/genetics ; DNA-Directed DNA Polymerase/metabolism ; Drosophila melanogaster/genetics ; Gene Expression Regulation ; Holliday Junction Resolvases/genetics ; Homologous Recombination ; Mitosis/genetics ; Synthetic Lethal Mutations ; DNA Polymerase theta
Chemical Substances	BRCA2 Protein ; DNA-Directed DNA Polymerase (EC 2.7.7.7) ; Holliday Junction Resolvases (EC 3.1.21.-)
Language	English
Publishing date	2021-03-22
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2186725-2
ISSN	1553-7404 ; 1553-7390
ISSN (online)	1553-7404
ISSN	1553-7390
DOI	10.1371/journal.pgen.1009267
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Article ; Online: The effect of repeat length on Marcal1-dependent single-strand annealing in Drosophila.

Dewey, Evan B / Korda Holsclaw, Julie / Saghaey, Kiyarash / Wittmer, Mackenzie E / Sekelsky, Jeff

Genetics

2022 Volume 223, Issue 1

Abstract: Proper repair of DNA double-strand breaks is essential to the maintenance of genomic stability and avoidance of genetic disease. Organisms have many ways of repairing double-strand breaks, including the use of homologous sequences through homology- ... ...

Abstract	Proper repair of DNA double-strand breaks is essential to the maintenance of genomic stability and avoidance of genetic disease. Organisms have many ways of repairing double-strand breaks, including the use of homologous sequences through homology-directed repair. While homology-directed repair is often error free, in single-strand annealing homologous repeats flanking a double-strand break are annealed to one another, leading to the deletion of one repeat and the intervening sequences. Studies in yeast have shown a relationship between the length of the repeat and single-strand annealing efficacy. We sought to determine the effects of homology length on single-strand annealing in Drosophila, as Drosophila uses a different annealing enzyme (Marcal1) than yeast. Using an in vivo single-strand annealing assay, we show that 50 base pairs are insufficient to promote single-strand annealing and that 500-2,000 base pairs are required for maximum efficiency. Loss of Marcal1 generally followed the same homology length trend as wild-type flies, with single-strand annealing frequencies reduced to about a third of wild-type frequencies regardless of homology length. Interestingly, we find a difference in single-strand annealing rates between 500-base pair homologies that align to the annealing target either nearer or further from the double-strand break, a phenomenon that may be explained by Marcal1 dynamics. This study gives insights into Marcal1 function and provides important information to guide the design of genome engineering strategies that use single-strand annealing to integrate linear DNA constructs into a chromosomal double-strand break.
MeSH term(s)	Animals ; DNA Repair ; Drosophila/genetics ; Saccharomyces cerevisiae/genetics ; DNA Breaks, Double-Stranded ; DNA
Chemical Substances	DNA (9007-49-2)
Language	English
Publishing date	2022-10-27
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2167-2
ISSN	1943-2631 ; 0016-6731
ISSN (online)	1943-2631
ISSN	0016-6731
DOI	10.1093/genetics/iyac164
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Article ; Online: Annealing of Complementary DNA Sequences During Double-Strand Break Repair in

Holsclaw, Julie Korda / Sekelsky, Jeff

Genetics

2017 Volume 206, Issue 1, Page(s) 467–480

Abstract: DNA double-strand breaks (DSBs) pose a serious threat to genomic integrity. If unrepaired, they can lead to chromosome fragmentation and cell death. If repaired incorrectly, they can cause mutations and chromosome rearrangements. DSBs are repaired using ... ...

Abstract	DNA double-strand breaks (DSBs) pose a serious threat to genomic integrity. If unrepaired, they can lead to chromosome fragmentation and cell death. If repaired incorrectly, they can cause mutations and chromosome rearrangements. DSBs are repaired using end-joining or homology-directed repair strategies, with the predominant form of homology-directed repair being synthesis-dependent strand annealing (SDSA). SDSA is the first defense against genomic rearrangements and information loss during DSB repair, making it a vital component of cell health and an attractive target for chemotherapeutic development. SDSA has also been proposed to be the primary mechanism for integration of large insertions during genome editing with CRISPR/Cas9. Despite the central role for SDSA in genome stability, little is known about the defining step: annealing. We hypothesized that annealing during SDSA is performed by the annealing helicase SMARCAL1, which can anneal RPA-coated single DNA strands during replication-associated DNA damage repair. We used unique genetic tools in
MeSH term(s)	Adenosine Triphosphate/genetics ; Animals ; CRISPR-Cas Systems ; DNA Breaks, Double-Stranded ; DNA Damage/genetics ; DNA End-Joining Repair/genetics ; DNA, Complementary/genetics ; DNA-Binding Proteins/genetics ; Drosophila melanogaster/genetics ; Gene Editing ; Genomic Instability/genetics ; SMARCB1 Protein/genetics
Chemical Substances	DNA, Complementary ; DNA-Binding Proteins ; SMARCB1 Protein ; Adenosine Triphosphate (8L70Q75FXE)
Language	English
Publishing date	2017-03-03
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2167-2
ISSN	1943-2631 ; 0016-6731
ISSN (online)	1943-2631
ISSN	0016-6731
DOI	10.1534/genetics.117.200238
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Article ; Online: Cross-species investigation into the requirement of XPA for nucleotide excision repair.

Kose, Cansu / Cao, Xuemei / Dewey, Evan B / Malkoç, Mustafa / Adebali, Ogün / Sekelsky, Jeff / Lindsey-Boltz, Laura A / Sancar, Aziz

Nucleic acids research

2023 Volume 52, Issue 2, Page(s) 677–689

Abstract: After reconstitution of nucleotide excision repair (excision repair) with XPA, RPA, XPC, TFIIH, XPF-ERCC1 and XPG, it was concluded that these six factors are the minimal essential components of the excision repair machinery. All six factors are highly ... ...

Abstract	After reconstitution of nucleotide excision repair (excision repair) with XPA, RPA, XPC, TFIIH, XPF-ERCC1 and XPG, it was concluded that these six factors are the minimal essential components of the excision repair machinery. All six factors are highly conserved across diverse organisms spanning yeast to humans, yet no identifiable homolog of the XPA gene exists in many eukaryotes including green plants. Nevertheless, excision repair is reported to be robust in the XPA-lacking organism, Arabidopsis thaliana, which raises a fundamental question of whether excision repair could occur without XPA in other organisms. Here, we performed a phylogenetic analysis of XPA across all species with annotated genomes and then quantitatively measured excision repair in the absence of XPA using the sensitive whole-genome qXR-Seq method in human cell lines and two model organisms, Caenorhabditis elegans and Drosophila melanogaster. We find that although the absence of XPA results in inefficient excision repair and UV-sensitivity in humans, flies, and worms, excision repair of UV-induced DNA damage is detectable over background. These studies have yielded a significant discovery regarding the evolution of XPA protein and its mechanistic role in nucleotide excision repair.
MeSH term(s)	Animals ; Humans ; DNA Damage ; DNA Repair ; DNA-Binding Proteins/genetics ; DNA-Binding Proteins/metabolism ; Drosophila melanogaster/metabolism ; Excision Repair ; Nucleotides/metabolism ; Phylogeny ; Xeroderma Pigmentosum Group A Protein/genetics ; Xeroderma Pigmentosum Group A Protein/metabolism ; Plants/metabolism ; Evolution, Molecular
Chemical Substances	DNA-Binding Proteins ; Nucleotides ; Xeroderma Pigmentosum Group A Protein ; XPA protein, human
Language	English
Publishing date	2023-12-05
Publishing country	England
Document type	Journal Article
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad1104
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