scholarly journals DYRK1A regulates the recruitment of 53BP1 to the sites of DNA damage in part through interaction with RNF169

2018 ◽  
Author(s):  
Vijay R. Menon ◽  
Varsha Ananthapadmanabhan ◽  
Selene Swanson ◽  
Siddharth Saini ◽  
Fatmata Sesay ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Parp Inhibitor ◽  
Therapy Response ◽  
Strand Break ◽  
Dependent Manner ◽  
Dsb Repair ◽  
Altered Expression ◽  
Developmental Abnormalities

SummaryHumanDYRK1Agene encoding Dual-specificity tyrosine (Y)- Regulated Kinase 1A (DYRK1A) is a dosage-dependent gene whereby either trisomy or haploinsufficiency result in developmental abnormalities. However, the function and regulation of this important protein kinase are not fully understood. Here we report proteomic analysis of DYRK1A in human cells that revealed a novel role of DYRK1A in the DNA double-strand break (DSB) repair signaling. This novel function of DYRK1A is mediated in part by its interaction with ubiquitin-binding protein RNF169 that regulates the choice between homologous recombination (HR) and non-homologous end joining (NHEJ) DSB repair. Accumulation of RNF169 at the DSB sites promotes homologous recombination (HR) by limiting the recruitment of the scaffold protein 53BP1 that promotes NHEJ by protecting the DNA ends from resection. Inducible overexpression of active, but not the kinase inactive, DYRK1A in U-2 OS cells inhibited accumulation of 53BP1 at the DSB sites in RNF169-dependent manner. Mutation of DYRK1A phosphorylation sites in RNF169 or pharmacological inhibition of DYRK1A using harmine decreased the ability of RNF169 to displace 53BP1 from radiation-induced DSB sites. In order to further investigate the role of DYRK1A in regulation of DNA repair, we used CRISPR-Cas9 mediated knockout of DYRK1A in human and mouse cells. Interestingly, knockout of DYRK1A also caused a defect in 53BP1 DSB recruitment that was independent of RNF169, suggesting that dosage of DYRK1A can influence the DNA repair processes through several mechanisms. U-2 OS cells devoid of DYRK1A displayed an increased DNA repair and HR efficiency, and showed a decreased sensitivity to the PARP inhibitor olaparib when compared to control cells. Given evidence of its altered expression in human cancers, DYRK1A levels could represent a significant determinant of the DNA damaging therapy response.

scholarly journals The Role of Nonerythroid SpectrinαII in Cancer

2019 ◽  
Vol 2019 ◽  
pp. 1-14 ◽  
Author(s):  
Anne Ackermann ◽  
Angela Brieger
Keyword(s):  
Dna Repair ◽  
Current Knowledge ◽  
Tumor Development ◽  
Therapy Response ◽  
Cell Contact ◽  
Altered Expression ◽  
Potential Marker ◽  
Specific Regulation ◽  
And Migration

Nonerythroid spectrinαII (SPTAN1) is an important cytoskeletal protein that ensures vital cellular properties including polarity and cell stabilization. In addition, it is involved in cell adhesion, cell-cell contact, and apoptosis. The detection of altered expression of SPTAN1 in tumors indicates that SPTAN1 might be involved in the development and progression of cancer. SPTAN1 has been described in cancer and therapy response and proposed as a potential marker protein for neoplasia, tumor aggressiveness, and therapeutic efficiency. On one hand, the existing data suggest that overexpression of SPTAN1 in tumor cells reflects neoplastic and tumor promoting activity. On the other hand, nuclear SPTAN1 can have tumor suppressing effects by enabling DNA repair through interaction with DNA repair proteins. Moreover, SPTAN1 cleavage products occur during apoptosis and could serve as markers for the efficacy of cancer therapy. Due to SPTAN1’s multifaceted functions and its role in adhesion and migration, SPTAN1 can influence tumor growth and progression in both positive and negative directions depending on its specific regulation. This review summarizes the current knowledge on SPTAN1 in cancer and depicts several mechanisms by which SPTAN1 could impact tumor development and aggressiveness.

scholarly journals HDAC8 Maintain Cytoskeleton Integrity Via Homologous Recombination and Represent a Novel Therapeutic Target in Multiple Myeloma

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4385-4385
Author(s):  
Zuzana Chyra ◽  
Maria Gkotzamanidou ◽  
Masood A. Shammas ◽  
Vassilis L. Souliotis ◽  
Yan Xu ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Dna Repair ◽  
Homologous Recombination ◽  
Cell Lines ◽  
Hdac Inhibitors ◽  
Mass Spectrometry Analysis ◽  
Dependent Manner ◽  
Large Panel ◽  
Dose Dependent

Multiple Myeloma (MM) is a plasma cell malignancy vulnerable to epigenetic intervention, with histone deacetylases (HDACs) emerging as the most promising epigenetic targets in combination with current anti-myeloma agents. Pan-HDAC inhibitors are effective as therapeutic agents both in preclinical and clinical setting; however, there is an increasing emphasis on understanding the biological and molecular roles of individual HDACs to limit toxicities observed with pan-HDAC inhibitors. Based on correlation with patient outcome in three independent myeloma datasets, we have evaluated the functional role of HDAC8, a member of Class I HDAC isoenzymes, in MM. Unlike other isoforms, there is limited information about molecular and epigenomic functions of HDAC8. We have previously confirmed expression of HDAC8 in a large panel of MM cell lines, where it is localized predominantly to cytoplasm. Moreover, genetic and pharmacological modulation of HDAC8 with RNAi and specific inhibitor PCI-34051 resulted in a significant inhibition of myeloma cell proliferation and decrease in colony formation (p<.001). HDAC8 inhibition led to an increase in the ongoing spontaneous and radiation-induced DNA damage in MM cells by affecting DNA repair via the homologous recombination (HR) pathway, suggesting a novel function of HDAC8 in promoting HR and DNA repair in MM cells. Using laser micro-irradiation in MM1S and U2OS cells, we observed HDAC8 recruitment to DSBs sites and its co-localization with Rad51 and Scm3, a member of cohesin complex. A transcriptomic analysis of HDAC8 knock-down cells also shows perturbation of number of cytoskeleton-related genes confirming significant role of HAD8 in cytoskeleton rearrangement in MM. Mass-spectrometry analysis to identify the HDAC8 substrates in MM cells is currently ongoing. Classical pan-HDACi, such as SAHA (vorinostat), bind to HDAC8 with substantially diminished activity (IC50 = 2 μM), reflecting a unique binding site of this isoform. To discover and validate new small molecules with HDAC8 subtype selectivity, we have explored the efficacy of OJI-1, a novel selective and potent HDAC8 inhibitor (IC50 = 0.8 nM) with modest inhibition of HDAC6 (1200 nM). Treatment with OJI-1 selectively impact cell viability of a large panel of MM cell lines (n=20) in a time and dose dependent manner, while sparing healthy donors PBMC both in resting and activated state (n=3). The significantly higher IC50 observed in PBMCs suggests a favorable therapeutic index. Western blotting analysis confirmed target selectivity with significant time and dose dependent decrease in H3 and H4 acetylation in MM cells treated with OJI-1. Moreover, pharmacological inhibition of HDAC8 specifically inhibited HR but not non-homologous end joining. These data suggest that targeting of HDAC8 using OJI-1 could be effective treatment approach in MM. Based on molecular data combination studies and in vivo evaluation are ongoing. In conclusion, our results provide insight into the role of HDAC8 in DNA stability and cell growth and viability which can be exploited in future for therapeutic application alone and in combination in MM. Disclosures Munshi: Takeda: Consultancy; Janssen: Consultancy; Amgen: Consultancy; Abbvie: Consultancy; Janssen: Consultancy; Celgene: Consultancy; Takeda: Consultancy; Adaptive: Consultancy; Amgen: Consultancy; Adaptive: Consultancy; Abbvie: Consultancy; Oncopep: Consultancy; Oncopep: Consultancy; Celgene: Consultancy.

scholarly journals Nuclear NAD+ homeostasis is essential for naive and chemoresistant BRCA1/2-deficient tumor survival

2021 ◽  
Author(s):  
Daniele Musiani ◽  
Hatice Yucel ◽  
Laura Sourd ◽  
Elisabetta Marangoni ◽  
Raphael Ceccaldi
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Parp Inhibitors ◽  
Dependent Manner ◽  
Mechanistic Studies ◽  
Resistance Development ◽  
Dna Repair Proteins ◽  
Tumor Survival ◽  
Synthetically Lethal

Resistance to PARP inhibitors (PARPi) is emerging as the major obstacle to their effectiveness for the treatment of BRCA1/2-mutated, also referred as homologous recombination (HR)-deficient, tumors (HRD). Over the years, mechanistic studies gained insights on effectors acting downstream of PARP1, lagging behind the understanding of earlier events upstream - and thus independent - of PARP1. Here, we investigated the role of nuclear NAD+, an essential cofactor for the activity of key DNA repair proteins, including PARP1 and sirtuins. We show that NMNAT1- the enzyme synthesizing nuclear NAD+ - is synthetically lethal with BRCA1/2 in a PARP1-independent but SIRT6-dependent manner. Consequently, inhibition of NMNAT1/SIRT6 axis not only kills naive but also PARPi-resistant HRD cancer cells. Our results unravel a unique vulnerability of HRD tumors, therapeutically exploitable even upon PARPi resistance development.

scholarly journals Rational combination therapy for hepatocellular carcinoma with PARP1 and DNA-PK inhibitors

2020 ◽  
Vol 117 (42) ◽  
pp. 26356-26365 ◽  
Author(s):  
Chen Wang ◽  
Huanyin Tang ◽  
Anke Geng ◽  
Binghua Dai ◽  
Haiping Zhang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Dna Repair ◽  
Strand Break ◽  
Nonhomologous End Joining ◽  
End Joining ◽  
Dsb Repair ◽  
Altered Expression ◽  
Normal Tissues ◽  
Dna Double Strand Break ◽  
Rational Combination

Understanding differences in DNA double-strand break (DSB) repair between tumor and normal tissues would provide a rationale for developing DNA repair-targeted cancer therapy. Here, using knock-in mouse models for measuring the efficiency of two DSB repair pathways, homologous recombination (HR) and nonhomologous end-joining (NHEJ), we demonstrated that both pathways are up-regulated in hepatocellular carcinoma (HCC) compared with adjacent normal tissues due to altered expression of DNA repair factors, including PARP1 and DNA-PKcs. Surprisingly, inhibiting PARP1 with olaparib abrogated HR repair in HCC. Mechanistically, inhibiting PARP1 suppressed the clearance of nucleosomes at DNA damage sites by blocking the recruitment of ALC1 to DSB sites, thereby inhibiting RPA2 and RAD51 recruitment. Importantly, combining olaparib with NU7441, a DNA-PKcs inhibitor that blocks NHEJ in HCC, synergistically suppressed HCC growth in both mice and HCC patient-derived-xenograft models. Our results suggest the combined inhibition of both HR and NHEJ as a potential therapy for HCC.

scholarly journals The dual role of HOP2 in mammalian meiotic homologous recombination

2013 ◽  
Vol 42 (4) ◽  
pp. 2346-2357 ◽  
Author(s):  
Roberto J. Pezza ◽  
Oleg N. Voloshin ◽  
Alexander A. Volodin ◽  
Kingsley A. Boateng ◽  
Marina A. Bellani ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Homologous Chromosome ◽  
Strand Break ◽  
Dsb Repair ◽  
Chromosome Synapsis ◽  
Homologous Chromosome Pairing ◽  
Strand Invasion ◽  
High Level

Abstract Deletion of Hop2 in mice eliminates homologous chromosome synapsis and disrupts double-strand break (DSB) repair through homologous recombination. HOP2 in vitro shows two distinctive activities: when it is incorporated into a HOP2–MND1 complex it stimulates DMC1 and RAD51 recombination activities and the purified HOP2 alone is proficient in promoting strand invasion. We observed that a fraction of Mnd1−/− spermatocytes, which express HOP2 but apparently have inactive DMC1 and RAD51 due to lack of the HOP2–MND1 complex, exhibits a high level of chromosome synapsis and that most DSBs in these spermatocytes are repaired. This suggests that DSB repair catalyzed solely by HOP2 supports homologous chromosome pairing and synapsis. In addition, we show that in vitro HOP2 promotes the co-aggregation of ssDNA with duplex DNA, binds to ssDNA leading to unstacking of the bases, and promotes the formation of a three-strand synaptic intermediate. However, HOP2 shows distinctive mechanistic signatures as a recombinase. Namely, HOP2-mediated strand exchange does not require ATP and, in contrast to DMC1, joint molecules formed by HOP2 are more sensitive to mismatches and are efficiently dissociated by RAD54. We propose that HOP2 may act as a recombinase with specific functions in meiosis.

scholarly journals The Mre11-Rad50-Xrs2 Protein Complex Facilitates Homologous Recombination-Based Double-Strand Break Repair inSaccharomyces cerevisiae

1999 ◽  
Vol 19 (11) ◽  
pp. 7681-7687 ◽  
Author(s):  
Debra A. Bressan ◽  
Bonnie K. Baxter ◽  
John H. J. Petrini
Keyword(s):  
Homologous Recombination ◽  
Sister Chromatid ◽  
Protein Complex ◽  
Cellular Response ◽  
Strand Break ◽  
Double Strand Break ◽  
Dsb Repair ◽  
Synchronous Cultures ◽  
Dna Dsbs

ABSTRACT Saccharomyces cerevisiae mre11Δ mutants are profoundly deficient in double-strand break (DSB) repair, indicating that the Mre11-Rad50-Xrs2 protein complex plays a central role in the cellular response to DNA DSBs. In this study, we examined the role of the complex in homologous recombination, the primary mode of DSB repair in yeast. We measured survival in synchronous cultures following irradiation and scored sister chromatid and interhomologue recombination genetically. mre11Δ strains were profoundly sensitive to ionizing radiation (IR) throughout the cell cycle. Mutant strains exhibited decreased frequencies of IR-induced sister chromatid and interhomologue recombination, indicating a general deficiency in homologous recombination-based DSB repair. Since a nuclease-deficientmre11 mutant was not impaired in these assays, it appears that the role of the S. cerevisiae Mre11-Rad50-Xrs2 protein complex in facilitating homologous recombination is independent of its nuclease activities.

scholarly journals Mrc1-Dependent Chromatin Compaction Represses DNA Double-Stranded Break Repair by Homologous Recombination Upon Replication Stress

2021 ◽  
Vol 9 ◽  
Author(s):  
Poyuan Xing ◽  
Yang Dong ◽  
Jingyu Zhao ◽  
Zhou Zhou ◽  
Zhao Li ◽  
...  
Keyword(s):  
Dna Replication ◽  
Homologous Recombination ◽  
Critical Role ◽  
Protein A ◽  
Replication Stress ◽  
Dependent Manner ◽  
Dsb Repair ◽  
Chromatin Compaction ◽  
End Resection

The coordination of DNA replication and repair is critical for the maintenance of genome stability. It has been shown that the Mrc1-mediated S phase checkpoint inhibits DNA double-stranded break (DSB) repair through homologous recombination (HR). How the replication checkpoint inhibits HR remains only partially understood. Here we show that replication stress induces the suppression of both Sgs1/Dna2- and Exo1-mediated resection pathways in an Mrc1-dependent manner. As a result, the loading of the single-stranded DNA binding factor replication protein A (RPA) and Rad51 and DSB repair by HR were severely impaired under replication stress. Notably, the deletion of MRC1 partially restored the recruitment of resection enzymes, DSB end resection, and the loading of RPA and Rad51. The role of Mrc1 in inhibiting DSB end resection is independent of Csm3, Tof1, or Ctf4. Mechanistically, we reveal that replication stress induces global chromatin compaction in a manner partially dependent on Mrc1, and this chromatin compaction limits the access of chromatin remodeling factors and HR proteins, leading to the suppression of HR. Our study reveals a critical role of the Mrc1-dependent chromatin structure change in coordinating DNA replication and recombination under replication stress.

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.

scholarly journals End-resection at DNA double-strand breaks in the three domains of life

2013 ◽  
Vol 41 (1) ◽  
pp. 314-320 ◽  
Author(s):  
John K. Blackwood ◽  
Neil J. Rzechorzek ◽  
Sian M. Bray ◽  
Joseph D. Maman ◽  
Luca Pellegrini ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homologous Chromosomes ◽  
Domains Of Life ◽  
Strand Invasion ◽  
End Resection

During DNA repair by HR (homologous recombination), the ends of a DNA DSB (double-strand break) must be resected to generate single-stranded tails, which are required for strand invasion and exchange with homologous chromosomes. This 5′–3′ end-resection of the DNA duplex is an essential process, conserved across all three domains of life: the bacteria, eukaryota and archaea. In the present review, we examine the numerous and redundant helicase and nuclease systems that function as the enzymatic analogues for this crucial process in the three major phylogenetic divisions.

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