scholarly journals The Over-expression of the β2 Catalytic Subunit of the Proteasome Decreases Homologous Recombination and Impairs DNA Double-Strand Break Repair in Human Cells

2011 ◽  
Vol 2011 ◽  
pp. 1-7 ◽  
Author(s):  
Anita Collavoli ◽  
Laura Comelli ◽  
Tiziana Cervelli ◽  
Alvaro Galli
Keyword(s):  
Homologous Recombination ◽  
Active Role ◽  
Strand Break ◽  
Double Strand Break ◽  
Human Cells ◽  
Proteasome Activity ◽  
Dsb Repair ◽  
Catalytic Subunits ◽  
Over Expression ◽  
Dna Double Strand Break

By a human cDNA library screening, we have previously identified two sequences coding two different catalytic subunits of the proteasome which increase homologous recombination (HR) when overexpressed in the yeastSaccharomyces cerevisiae. Here, we investigated the effect of proteasome on spontaneous HR and DNA repair in human cells. To determine if the proteasome has a role in the occurrence of spontaneous HR in human cells, we overexpressed the β2 subunit of the proteasome in HeLa cells and determined the effect on intrachromosomal HR. Results showed that the overexpression of β2 subunit decreased HR in human cells without altering the cell proteasome activity and the Rad51p level. Moreover, exposure to MG132 that inhibits the proteasome activity reduced HR in human cells. We also found that the expression of the β2 subunit increases the sensitivity to the camptothecin that induces DNA double-strand break (DSB). This suggests that the β2 subunit has an active role in HR and DSB repair but does not alter the intracellular level of the Rad51p.

scholarly journals The deSUMOylase SENP2 coordinates homologous recombination and non-homologous end joining by independent mechanisms

2018 ◽  
Author(s):  
Alexander J. Garvin ◽  
Alexandra K. Walker ◽  
Ruth M. Densham ◽  
Anoop Singh Chauhan ◽  
Helen R. Stone ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Break ◽  
End Joining ◽  
Dsb Repair ◽  
Dna Double Strand Break ◽  
Non Homologous End Joining

AbstractSUMOylation in the DNA double-strand break (DSB) response regulates recruitment, activity and clearance of repair factors. However, our understanding of a role for deSUMOylation in this process is limited. Here we identify different mechanistic roles for deSUMOylation in homologous recombination (HR) and non-homologous enjoining (NHEJ) through the investigation of the deSUMOylase SENP2. We find regulated deSUMOylation of MDC1 prevents excessive SUMOylation and its RNF4-VCP mediated clearance from DSBs, thereby promoting NHEJ. In contrast we show HR is differentially sensitive to SUMO availability and SENP2 activity is needed to provide SUMO. SENP2 is amplified as part of the chromosome 3q amplification in many cancers. Increased SENP2 expression prolongs MDC1 foci retention and increases NHEJ and radioresistance. Collectively our data reveal that deSUMOylation differentially primes cells for responding to DSBs and demonstrates the ability of SENP2 to tune DSB repair responses.

scholarly journals Live imaging of induced and controlled DNA double-strand break formation reveals extremely low repair by homologous recombination in human cells

Oncogene ◽  
2011 ◽  
Vol 31 (30) ◽  
pp. 3495-3504 ◽  
Author(s):  
O D Shahar ◽  
E V S Raghu Ram ◽  
E Shimshoni ◽  
S Hareli ◽  
E Meshorer ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Strand Break ◽  
Live Imaging ◽  
Double Strand Break ◽  
Human Cells ◽  
Dna Double Strand Break ◽  
Break Formation

scholarly journals Homologous Recombination Subpathways: A Tangle to Resolve

2021 ◽  
Vol 12 ◽  
Author(s):  
Amira Elbakry ◽  
Markus Löbrich
Keyword(s):  
Homologous Recombination ◽  
Cell Types ◽  
Strand Break ◽  
Human Cells ◽  
Dsb Repair ◽  
New Developments ◽  
Alternative Models ◽  
Dna Double Strand Break ◽  
Different Cell Types ◽  
The Impact

Homologous recombination (HR) is an essential pathway for DNA double-strand break (DSB) repair, which can proceed through various subpathways that have distinct elements and genetic outcomes. In this mini-review, we highlight the main features known about HR subpathways operating at DSBs in human cells and the factors regulating subpathway choice. We examine new developments that provide alternative models of subpathway usage in different cell types revise the nature of HR intermediates involved and reassess the frequency of repair outcomes. We discuss the impact of expanding our understanding of HR subpathways and how it can be clinically exploited.

scholarly journals Preserving genome integrity in human cells via DNA double-strand break repair

2020 ◽  
Vol 31 (9) ◽  
pp. 859-865 ◽  
Author(s):  
Ryan B. Jensen ◽  
Eli Rothenberg
Keyword(s):  
Dna Repair ◽  
Strand Break ◽  
Double Strand Break ◽  
Human Cells ◽  
Cell Cycle Checkpoints ◽  
Human Diseases ◽  
Genome Integrity ◽  
Dsb Repair ◽  
Dna Double Strand Break ◽  
Repair Pathways

The efficient maintenance of genome integrity in the face of cellular stress is vital to protect against human diseases such as cancer. DNA replication, chromatin dynamics, cellular signaling, nuclear architecture, cell cycle checkpoints, and other cellular activities contribute to the delicate spatiotemporal control that cells utilize to regulate and maintain genome stability. This perspective will highlight DNA double-strand break (DSB) repair pathways in human cells, how DNA repair failures can lead to human disease, and how PARP inhibitors have emerged as a novel clinical therapy to treat homologous recombination-deficient tumors. We briefly discuss how failures in DNA repair produce a permissive genetic environment in which preneoplastic cells evolve to reach their full tumorigenic potential. Finally, we conclude that an in-depth understanding of DNA DSB repair pathways in human cells will lead to novel therapeutic strategies to treat cancer and potentially other human diseases.

scholarly journals β1-Integrin Impacts Rad51 Stability and DNA Double-Strand Break Repair by Homologous Recombination

2018 ◽  
Vol 38 (9) ◽  
Author(s):  
Kazi Mokim Ahmed ◽  
Raj K. Pandita ◽  
Dharmendra Kumar Singh ◽  
Clayton R. Hunt ◽  
Tej K. Pandita
Keyword(s):  
Breast Cancer ◽  
Homologous Recombination ◽  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Strand Break ◽  
Double Strand Break ◽  
Β1 Integrin ◽  
Integrin Expression ◽  
Dsb Repair ◽  
Dna Double Strand Break

ABSTRACT The molecular mechanisms underlying resistance to radiotherapy in breast cancer cells remain elusive. Previously, we reported that elevated β1-integrin is associated with enhanced breast cancer cell survival postirradiation, but how β1-integrin conferred radioresistance was unclear. Ionizing radiation (IR) induced cell killing correlates with the efficiency of DNA double-strand break (DSB) repair, and we found that nonmalignant breast epithelial (S1) cells with low β1-integrin expression have a higher frequency of S-phase-specific IR-induced chromosomal aberrations than the derivative malignant breast (T4-2) cells with high β1-integrin expression. In addition, there was an increased frequency of IR-induced homologous recombination (HR) repairosome focus formation in T4-2 cells compared with that of S1 cells. Cellular levels of Rad51 in T4-2 cells, a critical factor in HR-mediated DSB repair, were significantly higher. Blocking or depleting β1-integrin activity in T4-2 cells reduced Rad51 levels, while ectopic expression of β1-integrin in S1 cells correspondingly increased Rad51 levels, suggesting that Rad51 is regulated by β1-integrin. The low level of Rad51 protein in S1 cells was found to be due to rapid degradation by the ubiquitin proteasome pathway (UPP). Furthermore, the E3 ubiquitin ligase RING1 was highly upregulated in S1 cells compared to T4-2 cells. Ectopic β1-integrin expression in S1 cells reduced RING1 levels and increased Rad51 accumulation. In contrast, β1-integrin depletion in T4-2 cells significantly increased RING1 protein levels and potentiated Rad51 ubiquitination. These data suggest for the first time that elevated levels of the extracellular matrix receptor β1-integrin can increase tumor cell radioresistance by decreasing Rad51 degradation through a RING1-mediated proteasomal pathway.

scholarly journals A novel nuclear speckle factor, USP42, promotes homologous recombination repair by resolving DNA double-strand break induced R-loop

2019 ◽  
Author(s):  
Misaki Matsui ◽  
Ryo Sakasai ◽  
Masako Abe ◽  
Yusuke Kimura ◽  
Shoki Kajita ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Break ◽  
Nuclear Bodies ◽  
Nuclear Speckles ◽  
Dsb Repair ◽  
Nuclear Speckle ◽  
Spatial Regulation ◽  
Intrinsically Disordered ◽  
Dna Double Strand Break

AbstractThe nucleus of mammalian cells is highly compartmentalized by nuclear bodies, including nuclear speckles. While nuclear bodies are known to function in regulating gene expression, their involvement in DNA repair has not been actively investigated. Here, our focused screen for nuclear speckle factors involved in homologous recombination (HR), which is a faithful DNA double-strand break (DSB) repair mechanism, revealed that nuclear speckle factors regulating transcription are potentially involved in the regulation of HR. Among the top hits, we provide evidence showing that USP42, which is a deubiquitylating enzyme and a hitherto unidentified nuclear speckles factor, promotes HR by facilitating BRCA1 recruitment to DSB sites and DNA-end resection. We further showed that USP42 localizes to nuclear speckles via an intrinsically disordered region, which is required for efficient HR. Furthermore, we established that USP42 interacts with DHX9, which possesses DNA-RNA helicase activity, and is required for efficient resolution of DSB-induced R-loop. Mechanistically, USP42 antagonizes mono-ubiquitylation of DHX9 that is evoked after DSB induction. In conclusion, our data propose a model in which a novel nuclear speckle factor, USP42, facilitates DSB-induced R-loop resolution, BRCA1 loading to DSB sites and preferential DSB repair by HR, indicating the importance of spatial regulation of DSB repair choice mediated by nuclear bodies.Significant statementDefects in the repair of DNA double-strand break (DSB), which is one of the most harmful DNA insults, cause human diseases including cancers. It has been suggested that DSBs generated in the coding region tend to be repaired by homologous recombination (HR) that is error-free DSB repair pathway. To reveal the spatial regulation of HR, in this study, we investigated the potential contribution of nuclear bodies, especially nuclear speckles, to HR, identifying a deubiquitylating enzyme USP42 as a HR promoting factor. We found that USP42 deubiquitylates DHX9, facilitates resolution of DNA-RNA hybrid structure and enhances HR through BRCA1 loading to DSB sites.ClassificationBiological Sciences, Cell Biology

scholarly journals Formation of Large Palindromic DNA by Homologous Recombination of Short Inverted Repeat Sequences in Saccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 161 (3) ◽  
pp. 1065-1075
Author(s):  
David K Butler ◽  
David Gillespie ◽  
Brandi Steele
Keyword(s):  
Homologous Recombination ◽  
Inverted Repeat ◽  
Excision Repair ◽  
Strand Break ◽  
Double Strand Break ◽  
Repeat Sequence ◽  
Inverted Repeat Sequence ◽  
Repeat Sequences ◽  
Dna Double Strand Break ◽  
Intermolecular Reaction

Abstract Large DNA palindromes form sporadically in many eukaryotic and prokaryotic genomes and are often associated with amplified genes. The presence of a short inverted repeat sequence near a DNA double-strand break has been implicated in the formation of large palindromes in a variety of organisms. Previously we have established that in Saccharomyces cerevisae a linear DNA palindrome is efficiently formed from a single-copy circular plasmid when a DNA double-strand break is introduced next to a short inverted repeat sequence. In this study we address whether the linear palindromes form by an intermolecular reaction (that is, a reaction between two identical fragments in a head-to-head arrangement) or by an unusual intramolecular reaction, as it apparently does in other examples of palindrome formation. Our evidence supports a model in which palindromes are primarily formed by an intermolecular reaction involving homologous recombination of short inverted repeat sequences. We have also extended our investigation into the requirement for DNA double-strand break repair genes in palindrome formation. We have found that a deletion of the RAD52 gene significantly reduces palindrome formation by intermolecular recombination and that deletions of two other genes in the RAD52-epistasis group (RAD51 and MRE11) have little or no effect on palindrome formation. In addition, palindrome formation is dramatically reduced by a deletion of the nucleotide excision repair gene RAD1.

scholarly journals RAD6 Promotes Homologous Recombination Repair by Activating the Autophagy-Mediated Degradation of Heterochromatin Protein HP1

2014 ◽  
Vol 35 (2) ◽  
pp. 406-416 ◽  
Author(s):  
Su Chen ◽  
Chen Wang ◽  
Luxi Sun ◽  
Da-Liang Wang ◽  
Lu Chen ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Genome Stability ◽  
Chromosomal Rearrangements ◽  
Strand Break ◽  
Open Chromatin ◽  
Dsb Repair ◽  
Heterochromatin Protein ◽  
Dna Double Strand Break ◽  
Recombination Repair ◽  
Ubiquitin Conjugating Enzyme

Efficient DNA double-strand break (DSB) repair is critical for the maintenance of genome stability. Unrepaired or misrepaired DSBs cause chromosomal rearrangements that can result in severe consequences, such as tumorigenesis. RAD6 is an E2 ubiquitin-conjugating enzyme that plays a pivotal role in repairing UV-induced DNA damage. Here, we present evidence that RAD6 is also required for DNA DSB repair via homologous recombination (HR) by specifically regulating the degradation of heterochromatin protein 1α (HP1α). Our study indicates that RAD6 physically interacts with HP1α and ubiquitinates HP1α at residue K154, thereby promoting HP1α degradation through the autophagy pathway and eventually leading to an open chromatin structure that facilitates efficient HR DSB repair. Furthermore, bioinformatics studies have indicated that the expression of RAD6 and HP1α exhibits an inverse relationship and correlates with the survival rate of patients.

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