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Niels Mailand - One of the best experts on this subject based on the ideXlab platform.
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RNF8 and RNF168 but not HERC2 are required for DNA damage-induced ubiquitylation in chicken DT40 cells.
DNA repair, 2012Co-Authors: Vibe H. Oestergaard, Simon Bekker-jensen, Niels Mailand, Constanze Pentzold, Rune Troelsgaard Pedersen, Silviu Iosif, Arno F. Alpi, Michael LisbyAbstract:The ubiquitylation cascade plays an important role in the recruitment of repair factors at DNA double-strand breaks. The involvement of a growing number of ubiquitin E3 ligases adds to the complexity of the DNA damage-induced ubiquitin signaling. Here we use the genetically tractable avian cell line DT40 to investigate the role of HERC2, RNF8 and RNF168 in the DNA damage-induced ubiquitylation pathway. We show that formation of ubiquitin foci as well as cell survival after DNA damage depends on both RNF8 and RNF168. However, we find that RNF8 and RNF168 knockout cell lines respond differently to treatment with camptothecin indicating that they do not function in a strictly linear manner. Surprisingly, we show that HERC2 is required neither for survival nor for ubiquitin foci formation after DNA damage in DT40. Moreover, the E3 ubiquitin ligase activity of HERC2 is not redundant to that of RNF8 or RNF168.
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A new non-catalytic role for ubiquitin ligase RNF8 in unfolding higher-order chromatin structure
The EMBO journal, 2012Co-Authors: Martijn S. Luijsterburg, Klara Acs, Leena Ackermann, Wouter W. Wiegant, Simon Bekker-jensen, Dorthe Helena Larsen, Kum Kum Khanna, Haico Van Attikum, Niels Mailand, Nico P. DantumaAbstract:The ubiquitin ligases RNF8 and RNF168 orchestrate DNA damage signalling through the ubiquitylation of histone H2A and the recruitment of downstream repair factors. Here, we demonstrate that RNF8, but not RNF168 or the canonical H2A ubiquitin ligase RNF2, mediates extensive chromatin decondensation. Our data show that CHD4, the catalytic subunit of the NuRD complex, interacts with RNF8 and is essential for RNF8-mediated chromatin unfolding. The chromatin remodelling activity of CHD4 promotes efficient ubiquitin conjugation and assembly of RNF168 and BRCA1 at DNA double-strand breaks. Interestingly, RNF8-mediated recruitment of CHD4 and subsequent chromatin remodelling were independent of the ubiquitin-ligase activity of RNF8, but involved a non-canonical interaction with the forkhead-associated (FHA) domain. Our study reveals a new mechanism of chromatin remodelling-assisted ubiquitylation, which involves the cooperation between CHD4 and RNF8 to create a local chromatin environment that is permissive to the assembly of checkpoint and repair machineries at DNA lesions.
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DNA damage-inducible SUMOylation of HERC2 promotes RNF8 binding via a novel SUMO-binding Zinc finger.
The Journal of cell biology, 2012Co-Authors: Jannie Rendtlew Danielsen, Simon Bekker-jensen, Bine H. Villumsen, Lou Klitgaard Povlsen, Werner Streicher, Jakob Nilsson, Mats Wikström, Niels MailandAbstract:Nonproteolytic ubiquitylation of chromatin surrounding deoxyribonucleic acid (DNA) double-strand breaks (DSBs) by the RNF8/RNF168/HERC2 ubiquitin ligases facilitates restoration of genome integrity by licensing chromatin to concentrate genome caretaker proteins near the lesions. In parallel, SUMOylation of so-far elusive upstream DSB regulators is also required for execution of this ubiquitin-dependent chromatin response. We show that HERC2 and RNF168 are novel DNA damage–dependent SUMOylation targets in human cells. In response to DSBs, both HERC2 and RNF168 were specifically modified with SUMO1 at DSB sites in a manner dependent on the SUMO E3 ligase PIAS4. SUMOylation of HERC2 was required for its DSB-induced association with RNF8 and for stabilizing the RNF8–Ubc13 complex. We also demonstrate that the ZZ Zinc finger in HERC2 defined a novel SUMO-specific binding module, which together with its concomitant SUMOylation and T4827 phosphorylation promoted binding to RNF8. Our findings provide novel insight into the regulatory complexity of how ubiquitylation and SUMOylation cooperate to orchestrate protein interactions with DSB repair foci.
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herc2 coordinates ubiquitin dependent assembly of dna repair factors on damaged chromosomes
Nature Cell Biology, 2010Co-Authors: Simon Bekkerjensen, Jiri Lukas, Jiri Bartek, Jannie Rendtlew Danielsen, Kasper Fugger, Irina Gromova, Annika Nerstedt, Claudia Lukas, Niels MailandAbstract:HERC2 regulates the retention of repair proteins 53BP1, RAP80 and BRCA1 on damaged chromosomes in response to ionizing radiation by forming a complex with the ubiquitin ligases RNF8 and RNF168. Hecr2 deficiency leads to radiosensitivity. Regulatory ubiquitylation is emerging as an important mechanism to protect genome integrity in cells exposed to DNA damage1,2,3,4,5,6,7,8,9. However, the spectrum of known ubiquitin regulators of the DNA damage response (DDR) is limited and their functional interplay is poorly understood. Here, we identify HERC2 as a factor that regulates ubiquitin-dependent retention of repair proteins on damaged chromosomes. In response to ionising radiation (IR), HERC2 forms a complex with RNF8, a ubiquitin ligase involved in the DDR3,4,5,6. The HERC2–RNF8 interaction requires IR-inducible phosphorylation of HERC2 at Thr 4827, which in turn binds to the forkhead-associated (FHA) domain of RNF8. Mechanistically, we provide evidence that HERC2 facilitates assembly of the ubiquitin-conjugating enzyme Ubc13 with RNF8, thereby promoting DNA damage-induced formation of Lys 63-linked ubiquitin chains. We also show that HERC2 interacts with, and maintains the levels of, RNF168, another ubiquitin ligase operating downstream of RNF8 (Refs 7, 8). Consequently, knockdown of HERC2 abrogates ubiquitin-dependent retention of repair factors such as 53BP1, RAP80 and BRCA1. Together with the increased radiosensitivity of HERC2-depleted cells, these results uncover a regulatory layer in the orchestration of protein interactions on damaged chromosomes and they underscore the role of ubiquitin-mediated signalling in genome maintenance.
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Nucleotide excision repair-induced H2A ubiquitination is dependent on MDC1 and RNF8 and reveals a universal DNA damage response
The Journal of cell biology, 2009Co-Authors: Jurgen A. Marteijn, Simon Bekker-jensen, Niels Mailand, Nico P. Dantuma, Jiri Lukas, Hannes Lans, Petra Schwertman, Audrey M. Gourdin, Wim VermeulenAbstract:Chromatin modifications are an important component of the of DNA damage response (DDR) network that safeguard genomic integrity. Recently, we demonstrated nucleotide excision repair (NER)–dependent histone H2A ubiquitination at sites of ultraviolet (UV)-induced DNA damage. In this study, we show a sustained H2A ubiquitination at damaged DNA, which requires dynamic ubiquitination by Ubc13 and RNF8. Depletion of these enzymes causes UV hypersensitivity without affecting NER, which is indicative of a function for Ubc13 and RNF8 in the downstream UV–DDR. RNF8 is targeted to damaged DNA through an interaction with the double-strand break (DSB)–DDR scaffold protein MDC1, establishing a novel function for MDC1. RNF8 is recruited to sites of UV damage in a cell cycle–independent fashion that requires NER-generated, single-stranded repair intermediates and ataxia telangiectasia–mutated and Rad3-related protein. Our results reveal a conserved pathway of DNA damage–induced H2A ubiquitination for both DSBs and UV lesions, including the recruitment of 53BP1 and Brca1. Although both lesions are processed by independent repair pathways and trigger signaling responses by distinct kinases, they eventually generate the same epigenetic mark, possibly functioning in DNA damage signal amplification.
Simon Bekker-jensen - One of the best experts on this subject based on the ideXlab platform.
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Histone H1 couples initiation and amplification of ubiquitin signalling after DNA damage
Nature, 2015Co-Authors: Tina Thorslund, Takeo Narita, Titia K. Sixma, Anita Ripplinger, Saskia Hoffmann, Thomas Wild, Michael Uckelmann, Bine H. Villumsen, Chunaram Choudhary, Simon Bekker-jensenAbstract:DNA double-strand breaks (DSBs) are highly cytotoxic DNA lesions that trigger non-proteolytic ubiquitylation of adjacent chromatin areas to generate binding sites for DNA repair factors. This depends on the sequential actions of the E3 ubiquitin ligases RNF8 and RNF168 (refs 1-6), and UBC13 (also known as UBE2N), an E2 ubiquitin-conjugating enzyme that specifically generates K63-linked ubiquitin chains. Whereas RNF168 is known to catalyse ubiquitylation of H2A-type histones, leading to the recruitment of repair factors such as 53BP1 (refs 8-10), the critical substrates of RNF8 and K63-linked ubiquitylation remain elusive. Here we elucidate how RNF8 and UBC13 promote recruitment of RNF168 and downstream factors to DSB sites in human cells. We establish that UBC13-dependent K63-linked ubiquitylation at DSB sites is predominantly mediated by RNF8 but not RNF168, and that H1-type linker histones, but not core histones, represent major chromatin-associated targets of this modification. The RNF168 module (UDM1) recognizing RNF8-generated ubiquitylations is a high-affinity reader of K63-ubiquitylated H1, mechanistically explaining the essential roles of RNF8 and UBC13 in recruiting RNF168 to DSBs. Consistently, reduced expression or chromatin association of linker histones impair accumulation of K63-linked ubiquitin conjugates and repair factors at DSB-flanking chromatin. These results identify histone H1 as a key target of RNF8-UBC13 in DSB signalling and expand the concept of the histone code by showing that posttranslational modifications of linker histones can serve as important marks for recognition by factors involved in genome stability maintenance, and possibly beyond.
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RNF8 and RNF168 but not HERC2 are required for DNA damage-induced ubiquitylation in chicken DT40 cells.
DNA repair, 2012Co-Authors: Vibe H. Oestergaard, Simon Bekker-jensen, Niels Mailand, Constanze Pentzold, Rune Troelsgaard Pedersen, Silviu Iosif, Arno F. Alpi, Michael LisbyAbstract:The ubiquitylation cascade plays an important role in the recruitment of repair factors at DNA double-strand breaks. The involvement of a growing number of ubiquitin E3 ligases adds to the complexity of the DNA damage-induced ubiquitin signaling. Here we use the genetically tractable avian cell line DT40 to investigate the role of HERC2, RNF8 and RNF168 in the DNA damage-induced ubiquitylation pathway. We show that formation of ubiquitin foci as well as cell survival after DNA damage depends on both RNF8 and RNF168. However, we find that RNF8 and RNF168 knockout cell lines respond differently to treatment with camptothecin indicating that they do not function in a strictly linear manner. Surprisingly, we show that HERC2 is required neither for survival nor for ubiquitin foci formation after DNA damage in DT40. Moreover, the E3 ubiquitin ligase activity of HERC2 is not redundant to that of RNF8 or RNF168.
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A new non-catalytic role for ubiquitin ligase RNF8 in unfolding higher-order chromatin structure
The EMBO journal, 2012Co-Authors: Martijn S. Luijsterburg, Klara Acs, Leena Ackermann, Wouter W. Wiegant, Simon Bekker-jensen, Dorthe Helena Larsen, Kum Kum Khanna, Haico Van Attikum, Niels Mailand, Nico P. DantumaAbstract:The ubiquitin ligases RNF8 and RNF168 orchestrate DNA damage signalling through the ubiquitylation of histone H2A and the recruitment of downstream repair factors. Here, we demonstrate that RNF8, but not RNF168 or the canonical H2A ubiquitin ligase RNF2, mediates extensive chromatin decondensation. Our data show that CHD4, the catalytic subunit of the NuRD complex, interacts with RNF8 and is essential for RNF8-mediated chromatin unfolding. The chromatin remodelling activity of CHD4 promotes efficient ubiquitin conjugation and assembly of RNF168 and BRCA1 at DNA double-strand breaks. Interestingly, RNF8-mediated recruitment of CHD4 and subsequent chromatin remodelling were independent of the ubiquitin-ligase activity of RNF8, but involved a non-canonical interaction with the forkhead-associated (FHA) domain. Our study reveals a new mechanism of chromatin remodelling-assisted ubiquitylation, which involves the cooperation between CHD4 and RNF8 to create a local chromatin environment that is permissive to the assembly of checkpoint and repair machineries at DNA lesions.
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DNA damage-inducible SUMOylation of HERC2 promotes RNF8 binding via a novel SUMO-binding Zinc finger.
The Journal of cell biology, 2012Co-Authors: Jannie Rendtlew Danielsen, Simon Bekker-jensen, Bine H. Villumsen, Lou Klitgaard Povlsen, Werner Streicher, Jakob Nilsson, Mats Wikström, Niels MailandAbstract:Nonproteolytic ubiquitylation of chromatin surrounding deoxyribonucleic acid (DNA) double-strand breaks (DSBs) by the RNF8/RNF168/HERC2 ubiquitin ligases facilitates restoration of genome integrity by licensing chromatin to concentrate genome caretaker proteins near the lesions. In parallel, SUMOylation of so-far elusive upstream DSB regulators is also required for execution of this ubiquitin-dependent chromatin response. We show that HERC2 and RNF168 are novel DNA damage–dependent SUMOylation targets in human cells. In response to DSBs, both HERC2 and RNF168 were specifically modified with SUMO1 at DSB sites in a manner dependent on the SUMO E3 ligase PIAS4. SUMOylation of HERC2 was required for its DSB-induced association with RNF8 and for stabilizing the RNF8–Ubc13 complex. We also demonstrate that the ZZ Zinc finger in HERC2 defined a novel SUMO-specific binding module, which together with its concomitant SUMOylation and T4827 phosphorylation promoted binding to RNF8. Our findings provide novel insight into the regulatory complexity of how ubiquitylation and SUMOylation cooperate to orchestrate protein interactions with DSB repair foci.
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Nucleotide excision repair-induced H2A ubiquitination is dependent on MDC1 and RNF8 and reveals a universal DNA damage response
The Journal of cell biology, 2009Co-Authors: Jurgen A. Marteijn, Simon Bekker-jensen, Niels Mailand, Nico P. Dantuma, Jiri Lukas, Hannes Lans, Petra Schwertman, Audrey M. Gourdin, Wim VermeulenAbstract:Chromatin modifications are an important component of the of DNA damage response (DDR) network that safeguard genomic integrity. Recently, we demonstrated nucleotide excision repair (NER)–dependent histone H2A ubiquitination at sites of ultraviolet (UV)-induced DNA damage. In this study, we show a sustained H2A ubiquitination at damaged DNA, which requires dynamic ubiquitination by Ubc13 and RNF8. Depletion of these enzymes causes UV hypersensitivity without affecting NER, which is indicative of a function for Ubc13 and RNF8 in the downstream UV–DDR. RNF8 is targeted to damaged DNA through an interaction with the double-strand break (DSB)–DDR scaffold protein MDC1, establishing a novel function for MDC1. RNF8 is recruited to sites of UV damage in a cell cycle–independent fashion that requires NER-generated, single-stranded repair intermediates and ataxia telangiectasia–mutated and Rad3-related protein. Our results reveal a conserved pathway of DNA damage–induced H2A ubiquitination for both DSBs and UV lesions, including the recruitment of 53BP1 and Brca1. Although both lesions are processed by independent repair pathways and trigger signaling responses by distinct kinases, they eventually generate the same epigenetic mark, possibly functioning in DNA damage signal amplification.
Lili Liang - One of the best experts on this subject based on the ideXlab platform.
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direct binding of RNF8 to sumo2 3 promotes cell survival following dna damage
Molecular Medicine Reports, 2017Co-Authors: Lili Liang, Zhenzhu Zhang, Likui Wang, Wenlin Huang, Shijuan GaoAbstract:Ring finger protein 8 (RNF8), an FHA/RING domain containing E3 ubiquitin ligase, is critical in supporting genome integrity by facilitating the assembly of multiple DNA repair proteins at DNA lesions following DNA damage. In the present study, a search for novel binding partners of RNF8 was performed using a yeast two‑hybrid screening assay, and small ubiquitin‑like modifier (SUMO)2/3 was identified as one of the major RNF8‑binding candidates. GST pull‑down and immunoprecipitation assays revealed that RNF8 bound directly and noncovalently to SUMO2/3, but not to SUMO1, and that the FHA domain of RNF8 was required for the binding to SUMO2/3. Furthermore, RNF8 co‑localized with SUMO2/3 at sites of DNA lesions in response to ionizing radiation, as revealed by immunofluorescence assay. Survival assay indicated that the depletion of RNF8 and SUMO2/3 resulted in decreased cellular resistance to genotoxic stress. These data suggested that the binding of RNF8 to SUMO2/3 promoted the response to DNA damage.
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Direct binding of RNF8 to SUMO2/3 promotes cell survival following DNA damage.
Molecular medicine reports, 2017Co-Authors: Lili Liang, Zhenzhu Zhang, Likui Wang, Wenlin Huang, Shijuan GaoAbstract:Ring finger protein 8 (RNF8), an FHA/RING domain containing E3 ubiquitin ligase, is critical in supporting genome integrity by facilitating the assembly of multiple DNA repair proteins at DNA lesions following DNA damage. In the present study, a search for novel binding partners of RNF8 was performed using a yeast two‑hybrid screening assay, and small ubiquitin‑like modifier (SUMO)2/3 was identified as one of the major RNF8‑binding candidates. GST pull‑down and immunoprecipitation assays revealed that RNF8 bound directly and noncovalently to SUMO2/3, but not to SUMO1, and that the FHA domain of RNF8 was required for the binding to SUMO2/3. Furthermore, RNF8 co‑localized with SUMO2/3 at sites of DNA lesions in response to ionizing radiation, as revealed by immunofluorescence assay. Survival assay indicated that the depletion of RNF8 and SUMO2/3 resulted in decreased cellular resistance to genotoxic stress. These data suggested that the binding of RNF8 to SUMO2/3 promoted the response to DNA damage.
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RNF8 negatively regulates NF-kappaB signaling by targeting IkappaB kinase: implications for the regulation of inflammation signaling.
Biochemical and biophysical research communications, 2017Co-Authors: Shijuan Gao, Lili LiangAbstract:Persistent or excess activation of NF-κB leads to cancer, autoimmune and inflammatory diseases. Therefore, activated NF-κB needs to be terminated after induction, which highlights the physiological significance of NF-κB-negative regulators. However, the molecular mechanisms that negatively regulate NF-κB are not well understood. Here, we report that Ring Finger Protein 8 (RNF8), an E3 ubiquitin ligase, inhibits TNFα-mediated NF-κB activation by targeting IκB kinase (IKK). Upon TNFα stimulation, RNF8 binds to the catalytic subunits of IKK complex, resulting in inhibition of IKKα/β phosphorylation and subsequent NF-κB activation. RNF8 targets the IKK complex in a manner independent of its RING domain. We further provide evidence that the silencing of RNF8 results in enhanced TNFα-induced IKK activation, and an increase expression of NF-κB-induced inflammatory cytokine IL-8. Our study identifies a previously unrecognized role for RNF8 in the negative regulation of NF-κB activation by targeting and deactivating the IKK complex.
Azad Bonni - One of the best experts on this subject based on the ideXlab platform.
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RNF8/UBC13 ubiquitin signaling suppresses synapse formation in the mammalian brain
Nature communications, 2017Co-Authors: Pamela Valnegri, Ju Huang, Tomoko Yamada, Yue Yang, Luis A. Mejia, Ha Y. Cho, Anna Oldenborg, Azad BonniAbstract:Although ubiquitin ligases have been implicated in autism, their roles and mechanisms in brain development remain incompletely understood. Here, we report that in vivo knockdown or conditional knockout of the autism-linked ubiquitin ligase RNF8 or associated ubiquitin-conjugating enzyme UBC13 in rodent cerebellar granule neurons robustly increases the number of parallel fiber presynaptic boutons and functional parallel fiber/Purkinje cell synapses. In contrast to the role of nuclear RNF8 in proliferating cells, RNF8 operates in the cytoplasm in neurons to suppress synapse differentiation in vivo. Proteomics analyses reveal that neuronal RNF8 interacts with the HECT domain protein HERC2 and scaffold protein NEURL4, and knockdown of HERC2 or NEURL4 phenocopies the inhibition of RNF8/UBC13 signaling on synapse differentiation. In behavior analyses, granule neuron-specific knockout of RNF8 or UBC13 impairs cerebellar-dependent learning. Our study defines RNF8 and UBC13 as components of a novel cytoplasmic ubiquitin-signaling network that suppresses synapse formation in the brain. Ubiquitin ligases play critical roles in neuronal connectivity in the brain. Here, Valnegri and colleagues show that ubiquitin ligase RNF8 and ubiquitin-conjugating enzyme UBC13 regulate synapse number in cerebellar granule neurons and rodent cerebellar learning.
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RNF8 ubc13 ubiquitin signaling suppresses synapse formation in the mammalian brain
Nature Communications, 2017Co-Authors: Pamela Valnegri, Ju Huang, Tomoko Yamada, Yue Yang, Luis A. Mejia, Ha Y. Cho, Anna Oldenborg, Azad BonniAbstract:Although ubiquitin ligases have been implicated in autism, their roles and mechanisms in brain development remain incompletely understood. Here, we report that in vivo knockdown or conditional knockout of the autism-linked ubiquitin ligase RNF8 or associated ubiquitin-conjugating enzyme UBC13 in rodent cerebellar granule neurons robustly increases the number of parallel fiber presynaptic boutons and functional parallel fiber/Purkinje cell synapses. In contrast to the role of nuclear RNF8 in proliferating cells, RNF8 operates in the cytoplasm in neurons to suppress synapse differentiation in vivo. Proteomics analyses reveal that neuronal RNF8 interacts with the HECT domain protein HERC2 and scaffold protein NEURL4, and knockdown of HERC2 or NEURL4 phenocopies the inhibition of RNF8/UBC13 signaling on synapse differentiation. In behavior analyses, granule neuron-specific knockout of RNF8 or UBC13 impairs cerebellar-dependent learning. Our study defines RNF8 and UBC13 as components of a novel cytoplasmic ubiquitin-signaling network that suppresses synapse formation in the brain. Ubiquitin ligases play critical roles in neuronal connectivity in the brain. Here, Valnegri and colleagues show that ubiquitin ligase RNF8 and ubiquitin-conjugating enzyme UBC13 regulate synapse number in cerebellar granule neurons and rodent cerebellar learning.
Shijuan Gao - One of the best experts on this subject based on the ideXlab platform.
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direct binding of RNF8 to sumo2 3 promotes cell survival following dna damage
Molecular Medicine Reports, 2017Co-Authors: Lili Liang, Zhenzhu Zhang, Likui Wang, Wenlin Huang, Shijuan GaoAbstract:Ring finger protein 8 (RNF8), an FHA/RING domain containing E3 ubiquitin ligase, is critical in supporting genome integrity by facilitating the assembly of multiple DNA repair proteins at DNA lesions following DNA damage. In the present study, a search for novel binding partners of RNF8 was performed using a yeast two‑hybrid screening assay, and small ubiquitin‑like modifier (SUMO)2/3 was identified as one of the major RNF8‑binding candidates. GST pull‑down and immunoprecipitation assays revealed that RNF8 bound directly and noncovalently to SUMO2/3, but not to SUMO1, and that the FHA domain of RNF8 was required for the binding to SUMO2/3. Furthermore, RNF8 co‑localized with SUMO2/3 at sites of DNA lesions in response to ionizing radiation, as revealed by immunofluorescence assay. Survival assay indicated that the depletion of RNF8 and SUMO2/3 resulted in decreased cellular resistance to genotoxic stress. These data suggested that the binding of RNF8 to SUMO2/3 promoted the response to DNA damage.
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Direct binding of RNF8 to SUMO2/3 promotes cell survival following DNA damage.
Molecular medicine reports, 2017Co-Authors: Lili Liang, Zhenzhu Zhang, Likui Wang, Wenlin Huang, Shijuan GaoAbstract:Ring finger protein 8 (RNF8), an FHA/RING domain containing E3 ubiquitin ligase, is critical in supporting genome integrity by facilitating the assembly of multiple DNA repair proteins at DNA lesions following DNA damage. In the present study, a search for novel binding partners of RNF8 was performed using a yeast two‑hybrid screening assay, and small ubiquitin‑like modifier (SUMO)2/3 was identified as one of the major RNF8‑binding candidates. GST pull‑down and immunoprecipitation assays revealed that RNF8 bound directly and noncovalently to SUMO2/3, but not to SUMO1, and that the FHA domain of RNF8 was required for the binding to SUMO2/3. Furthermore, RNF8 co‑localized with SUMO2/3 at sites of DNA lesions in response to ionizing radiation, as revealed by immunofluorescence assay. Survival assay indicated that the depletion of RNF8 and SUMO2/3 resulted in decreased cellular resistance to genotoxic stress. These data suggested that the binding of RNF8 to SUMO2/3 promoted the response to DNA damage.
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RNF8 negatively regulates NF-kappaB signaling by targeting IkappaB kinase: implications for the regulation of inflammation signaling.
Biochemical and biophysical research communications, 2017Co-Authors: Shijuan Gao, Lili LiangAbstract:Persistent or excess activation of NF-κB leads to cancer, autoimmune and inflammatory diseases. Therefore, activated NF-κB needs to be terminated after induction, which highlights the physiological significance of NF-κB-negative regulators. However, the molecular mechanisms that negatively regulate NF-κB are not well understood. Here, we report that Ring Finger Protein 8 (RNF8), an E3 ubiquitin ligase, inhibits TNFα-mediated NF-κB activation by targeting IκB kinase (IKK). Upon TNFα stimulation, RNF8 binds to the catalytic subunits of IKK complex, resulting in inhibition of IKKα/β phosphorylation and subsequent NF-κB activation. RNF8 targets the IKK complex in a manner independent of its RING domain. We further provide evidence that the silencing of RNF8 results in enhanced TNFα-induced IKK activation, and an increase expression of NF-κB-induced inflammatory cytokine IL-8. Our study identifies a previously unrecognized role for RNF8 in the negative regulation of NF-κB activation by targeting and deactivating the IKK complex.
