Elongation factor ELOF1 drives transcription-coupled repairand prevents genome instability [DRB-TTchem-seq]
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE148844 | GEO | 2020/04/18
REPOSITORIES: GEO
Similar Datasets
Project description:DNA damage forms a major obstacle for gene transcription by RNA polymerase II (Pol II). Transcription-coupled nucleotide excision repair (TC-NER) efficiently eliminates transcription-blocking lesions (TBLs), thereby safeguarding accurate transcription, preserving correct cellular function and counteracting aging. TC-NER initiation involves the recognition of lesion-stalled Pol II by CSB, which recruits the CRL4CSA E3 ubiquitin ligase complex and UVSSA. TBL-induced Pol II ubiquitylation at lysine K1268 by CRL4CSA serves as a critical TC-NER checkpoint, governing Pol II stability and initiating TBL excision by TFIIH recruitment. However, the precise regulatory mechanisms of the CRL4CSA E3 ligase activity and TFIIH recruitment remains elusive. Here, we reveal Serine/Threonine Kinase 19 (STK19) as a novel TC-NER factor. Cryo-EM studies revealed that STK19 is an integral part of the Pol II-TC‐NER complex, bridging CSA with UVSSA, RPB1 and downstream DNA. Live-cell imaging and interaction proteomics shows that STK19 stimulates TC-NER complex stability and proper CRL4CSA complex assembly, resulting in Pol II (K1268) ubiquitylation and subsequent UVSSA and TFIIH recruitment. These findings underscore the crucial role of STK19 as a core component of the TC-NER machinery and its key involvement to the cellular responses to genomic injuries that interfere with transcription.
2024-10-16 | PXD051720 | Pride
Project description:Correct transcription is crucial for life. However, DNA damage severely impedes elongating RNA Polymerase II (Pol II), causing transcription inhibition and transcription-replication conflicts. Cells are equipped with intricate mechanisms to counteract the severe consequence of these transcription-blocking lesions (TBLs). However, the exact mechanism and factors involved remain largely unknown. Here, using a genome-wide CRISPR/cas9 screen, we identified elongation factor ELOF1 as an important new factor in the transcription stress response upon DNA damage. We show that ELOF1 has an evolutionary conserved role in Transcription-Coupled Nucleotide Excision Repair (TC-NER), where it promotes recruitment of the TC-NER factors UVSSA and TFIIH to efficiently repair TBLs and resume transcription. Additionally, ELOF1 modulates transcription to protect cells from transcription-mediated replication stress, thereby preserving genome stability. Thus, ELOF1 protects the transcription machinery from DNA damage via two distinct mechanisms.
2021-04-20 | PXD025304 | Pride
Project description:DNA-protein crosslinks (DPCs), arise from enzymatic intermediates, endogenous metabolism or exogenous chemicals like chemotherapeutics. DPCs are highly cytotoxic as they will impede DNA transacting processes such as replication, which is counteracted by proteolysis-mediated DPC removal by the protease SPRTN or the proteasome. However, how DPCs affect transcription and how transcription-blocking DPCs are repaired remains largely unknown. Here, we show that DPCs severely inhibit RNA polymerase II (Pol II)-mediated transcription, resulting in the recruitment of the transcription-coupled nucleotide excision repair (TC-NER) factors CSA and CSB. Interestingly, CSA and CSB are indispensable for transcription-coupled DPC (TC-DPC) repair, while the downstream TC-NER factors UVSSA and XPA are not, indicative of a non-canonical TC-NER mechanism. TC-DPC repair functions independent of SPRTN, but is mediated by the activities of the ubiquitin ligase CRL4CSA and the proteasome. Thus, DPCs are preferentially repaired in genes by a dedicated transcription-coupled repair mechanism, which is crucial to warrant correct transcription.
2024-02-09 | PXD041679 | Pride
Project description:Transcription-blocking DNA lesions are specifically targeted by transcription-coupled nucleotide excision repair (TC-NER), which removes a broad spectrum of DNA lesions to preserve transcriptional output and thereby cellular homeostasis to counteract aging. TC-NER is initiated by the stalling of RNA polymerase II at DNA lesions, which triggers the assembly of the TC-NER-specific proteins CSA, CSB and UVSSA. CSA, a WD40-repeat containing protein, is the substrate receptor subunit of a cullin-RING ubiquitin ligase complex composed of DDB1, CUL4A/B and RBX1 (CRL4CSA). Although ubiquitination of several TC-NER proteins by CRL4CSA has been reported, it is still unknown how this complex is regulated. To unravel the dynamic molecular interactions and the regulation of this complex, we applied a single-step protein-complex isolation coupled to mass spectrometry analysis and identified DDA1 as a CSA interacting protein. Cryo-EM analysis showed that DDA1 is an integral component of the CRL4CSA complex. Functional analysis revealed that DDA1 coordinates ubiquitination dynamics during TC-NER and is required for efficient turnover and progression of this process.
2024-05-02 | PXD051638 | Pride
Project description:Transcription coupled-nucleotide excision repair (TC-NER) repairs DNA lesions that stall RNA polymerase II (Pol II) transcription. Here, we show that the C-terminal domain (CTD) of elongation factor-1 (Elf1) plays a critical role in TC-NER in yeast. Analysis of genome-wide repair of UV-induced cyclobutane pyrimidine dimers (CPDs) using CPD-seq indicates that the Elf1 CTD is required for efficient Rad26-dependent and Rad26-independent TC-NER across the yeast genome. The Elf1-CTD is also important for TC-NER in rad16∆ cells deficient in GG-NER. Finally, we show that a mutant in the Elf1-CTD (elf1-Y99A) that disrupts binding to a subunit of TFIIH affects Rad26-independent repair in a rad26∆ mutant background.
2024-05-31 | GSE243603 | GEO
Project description:Transcription-blocking DNA lesions are removed by transcription-coupled nucleotide excision repair (TC-NER) to preserve cell viability. TC-NER is triggered by the stalling of RNA polymerase II at DNA lesions, leading to the recruitment of TC-NER-specific factors such as the CSA-DDB1-CUL4A-RBX1 cullin-RING ubiquitin ligase complex (CRLCSA). Despite its vital role in TC-NER, little is known about the regulation of the CRLCSA complex during TC-NER. Using conventional and cross-linking immunoprecipitations coupled to mass spectrometry, we uncover a stable interaction between CSA and the TRiC chaperonin. TRiC’s binding to CSA ensures its stability and DDB1-dependent assembly into the CRLCSA complex. Consequently, loss of TRiC leads to mislocalization and depletion of CSA, as well as impaired transcription recovery following UV damage, suggesting defects in TC-NER. Furthermore, Cockayne syndrome (CS)-causing mutations in CSA lead to increased TRiC binding and a failure to compose the CRLCSA complex. Thus, we uncover CSA as a novel TRiC substrate and reveal a role for TRiC in regulating CSA-dependent TC-NER and the development of CS.
2018-05-02 | PXD008868 | Pride
Project description:Transcription–blocking lesions are specifically targeted by transcription-coupled nucleotide excision repair (TC-NER), which removes a broad spectrum of DNA lesions to preserve transcriptional output and thereby cellular homeostasis to counteract aging. TC-NER is initiated by the physical stalling of RNA polymerase II by a DNA damage, which triggers the assembly of TC-NER-specific proteins, including CSA, a WD40 domain containing protein, that forms together with DDB1, CUL4A/B and RBX1, a cullin-RING ubiquitin ligase complex (CRL4CSA). Although ubiquitination of several TC-NER proteins by CSA has been reported, it is still unknown how this complex functions in TC-NER progression. To unravel the molecular mechanism underlying TC-NER, we applied a single-step protein-complex isolation coupled to mass spectrometry analysis and identified DDA1 as a CSA interacting protein. Cryo-EM analysis showed that DDA1 is an integral component of the CRL4CSA complex. Functional analysis revealed that DDA1 coordinates the step-wise ubiquitination during TC-NER and is required for efficient progression of this process by controlling the dynamic turnover of CRL4CSA complex.
2024-05-31 | PXD045415 | Pride
Project description:Transcription-coupled nucleotide excision repair (TC-NER) is promptly triggered by 'local' elongating RNAP II molecules encountering DNA adducts such as cyclobutane pyrimidine dimers (CPD) induced by ultraviolet light and speeds up removing and repair in transcribed loci. Genome-wide measurements have revealed a dramatic shutdown of global transcription with a few exceptions of individual genes that are consequently highly upregulated by DNA damage. Deciphering the underlying mechanisms of the multifaceted transcriptional response always comprises one of the most fascinating frontiers in DNA repair research. Recent reports have revealed that persistent RNAP II initiation at the transcription start site of active regulatory regions with the continuous synthesis of start-RNAs guarantee sufficient scanning and repair of the whole transcribed genome, although transcription elongation drastically slows down shortly after genotoxic stress. Furthermore, a recent study found that RNA is extremely rapidly m6A methylated by METTL3 in response to UV irradiation which is crucial for recruitment of Pol k to DNA damage sites and subsequently mediates TC-NER. Together, these findings substantiate the molecular processes underlying the transcription-coordinated cellular response upon genotoxic stress might be more complex than previously believed. Unfortunately, all these findings merely focused on the early phase of recovery (within 4 hr after DNA damage). It remains to be determined how accessible chromatin reconfigures and regulates transcriptional programs in UV-induced DNA damage repair. It is also unclear whether the dynamics of gene expression are associated with other epigenomic reconstruction events such as global DNA methylation and demethylation. Moreover, the roles of internal messenger RNA modifications like N6-methyladenosine in TC-NER are poorly understood. Our integrative analyses provide a comprehensive view of the spatio-temporal chromatin configuration and epigenetic characterizations that accompanies TC-NER.
2020-11-20 | GSE161792 | GEO
Project description:Transcription-coupled nucleotide excision repair (TC-NER) is promptly triggered by 'local' elongating RNAP II molecules encountering DNA adducts such as cyclobutane pyrimidine dimers (CPD) induced by ultraviolet light and speeds up removing and repair in transcribed loci. Genome-wide measurements have revealed a dramatic shutdown of global transcription with a few exceptions of individual genes that are consequently highly upregulated by DNA damage. Deciphering the underlying mechanisms of the multifaceted transcriptional response always comprises one of the most fascinating frontiers in DNA repair research. Recent reports have revealed that persistent RNAP II initiation at the transcription start site of active regulatory regions with the continuous synthesis of start-RNAs guarantee sufficient scanning and repair of the whole transcribed genome, although transcription elongation drastically slows down shortly after genotoxic stress. Furthermore, a recent study found that RNA is extremely rapidly m6A methylated by METTL3 in response to UV irradiation which is crucial for recruitment of Pol k to DNA damage sites and subsequently mediates TC-NER. Together, these findings substantiate the molecular processes underlying the transcription-coordinated cellular response upon genotoxic stress might be more complex than previously believed. Unfortunately, all these findings merely focused on the early phase of recovery (within 4 hr after DNA damage). It remains to be determined how accessible chromatin reconfigures and regulates transcriptional programs in UV-induced DNA damage repair. It is also unclear whether the dynamics of gene expression are associated with other epigenomic reconstruction events such as global DNA methylation and demethylation. Moreover, the roles of internal messenger RNA modifications like N6-methyladenosine in TC-NER are poorly understood. Our integrative analyses provide a comprehensive view of the spatio-temporal chromatin configuration and epigenetic characterizations that accompanies TC-NER.
2020-11-20 | GSE161791 | GEO
Project description:Transcription-coupled nucleotide excision repair (TC-NER) is promptly triggered by 'local' elongating RNAP II molecules encountering DNA adducts such as cyclobutane pyrimidine dimers (CPD) induced by ultraviolet light and speeds up removing and repair in transcribed loci. Genome-wide measurements have revealed a dramatic shutdown of global transcription with a few exceptions of individual genes that are consequently highly upregulated by DNA damage. Deciphering the underlying mechanisms of the multifaceted transcriptional response always comprises one of the most fascinating frontiers in DNA repair research. Recent reports have revealed that persistent RNAP II initiation at the transcription start site of active regulatory regions with the continuous synthesis of start-RNAs guarantee sufficient scanning and repair of the whole transcribed genome, although transcription elongation drastically slows down shortly after genotoxic stress. Furthermore, a recent study found that RNA is extremely rapidly m6A methylated by METTL3 in response to UV irradiation which is crucial for recruitment of Pol k to DNA damage sites and subsequently mediates TC-NER. Together, these findings substantiate the molecular processes underlying the transcription-coordinated cellular response upon genotoxic stress might be more complex than previously believed. Unfortunately, all these findings merely focused on the early phase of recovery (within 4 hr after DNA damage). It remains to be determined how accessible chromatin reconfigures and regulates transcriptional programs in UV-induced DNA damage repair. It is also unclear whether the dynamics of gene expression are associated with other epigenomic reconstruction events such as global DNA methylation and demethylation. Moreover, the roles of internal messenger RNA modifications like N6-methyladenosine in TC-NER are poorly understood. Our integrative analyses provide a comprehensive view of the spatio-temporal chromatin configuration and epigenetic characterizations that accompanies TC-NER.
2020-11-20 | GSE161790 | GEO