Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1074553 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR27936500_1.fastq.gz | Fastqsanger.gz | |||
| SRR27936500_2.fastq.gz | Fastqsanger.gz | |||
| SRR27936501_1.fastq.gz | Fastqsanger.gz | |||
| SRR27936501_2.fastq.gz | Fastqsanger.gz | |||
| SRR27936502_1.fastq.gz | Fastqsanger.gz |
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Project description:Transcription-coupled DNA repair (TCR) efficiently removes bulky DNA lesions from transcribed parts of the genome and constitutes a major defence against DNA damage by UV irradiation. TCR begins when RNA polymerase II (Pol II) stalls at a DNA lesion and recruits the Cockayne syndrome protein CsB, the E3 ubiquitin ligase complex CRL4CsA, and the UV-stimulated scaffold protein A (UVSSA). Here we provide five high-resolution structures of Pol II transcription complexes with bound TCR factors and complementary biochemical data. Together with published work, our results converge on a model for the molecular mechanism of transcription-DNA repair coupling. In this model, Pol II stalling triggers the formation of an alternative transcription elongation complex that we call ECact. In ECact, CsB has replaced the elongation factor DSIF, binds the PAF1 complex, and repositions upstream DNA such that it contacts the elongation factor SPT6. The CsB translocase now pulls on the upstream DNA template, thereby pushing Pol II forward. If the lesion cannot be bypassed, Pol II gets stably stalled. CRL4CsA spans over the Pol II clamp to reach the Pol II jaw and direct ubiquitination of RPB1 residue lysine-1268. This triggers the recruitment of TFIIH to UVSSA, which is located at downstream DNA, and enables nucleotide excision repair. Finally, large-scale conformational changes in CRL4CsA enable ubiquitination of CsB and the release of TCR factors, thereby allowing Pol II to resume elongation and continue transcription over repaired DNA.
2022-02-17 | PXD025328 | Pride
Project description:Transcription-coupled DNA repair (TCR) removes bulky DNA lesions impeding RNA polymerase II (RNAPII) transcription. Recent studies have outlined the stepwise assembly of TCR factors CSB, CSA, UVSSA, and TFIIH around lesion-stalled RNAPII. However, the mechanism and factors required for the transition to downstream repair steps, including RNAPII removal to provide repair proteins access to the DNA lesion, remain unclear. Here, we identify STK19 as a new TCR factor facilitating this transition. Loss of STK19 does not impact initial TCR complex assembly or RNAPII ubiquitylation but delays lesion-stalled RNAPII clearance, thereby interfering with the downstream repair reaction. Cryo-EM and mutational analysis reveal that STK19 associates with the TCR complex, positioning itself between RNAPII, UVSSA, and CSA. The structural insights and molecular modeling suggest that STK19 positions the ATPase subunits of TFIIH onto DNA in front of RNAPII. Together, these findings provide new insights into the factors and mechanisms required for TCR.
2024-09-02 | GSE266083 | GEO
Project description:Transcription-coupled DNA repair (TCR) removes bulky DNA lesions impeding RNA polymerase II (RNAPII) transcription. Recent studies have outlined the stepwise assembly of TCR factors CSB, CSA, UVSSA, and TFIIH around lesion-stalled RNAPII. However, the mechanism and factors required for the transition to downstream repair steps, including RNAPII removal to provide repair proteins access to the DNA lesion, remain unclear. Here, we identify STK19 as a new TCR factor facilitating this transition. Loss of STK19 does not impact initial TCR complex assembly or RNAPII ubiquitylation but delays lesion-stalled RNAPII clearance, thereby interfering with the downstream repair reaction. Cryo-EM and mutational analysis reveal that STK19 associates with the TCR complex, positioning itself between RNAPII, UVSSA, and CSA. The structural insights and molecular modeling suggest that STK19 positions the ATPase subunits of TFIIH onto DNA in front of RNAPII. Together, these findings provide new insights into the factors and mechanisms required for TCR.
2024-09-02 | GSE266085 | GEO
Project description:Transcription-coupled DNA repair (TCR) removes bulky DNA lesions impeding RNA polymerase II (RNAPII) transcription. Recent studies have outlined the stepwise assembly of TCR factors CSB, CSA, UVSSA, and TFIIH around lesion-stalled RNAPII. However, the mechanism and factors required for the transition to downstream repair steps, including RNAPII removal to provide repair proteins access to the DNA lesion, remain unclear. Here, we identify STK19 as a new TCR factor facilitating this transition. Loss of STK19 does not impact initial TCR complex assembly or RNAPII ubiquitylation but delays lesion-stalled RNAPII clearance, thereby interfering with the downstream repair reaction. Cryo-EM and mutational analysis reveal that STK19 associates with the TCR complex, positioning itself between RNAPII, UVSSA, and CSA. The structural insights and molecular modelling suggest that STK19 positions the ATPase subunits of TFIIH onto DNA in front of RNAPII. Together, these findings provide new insights into the factors and mechanisms required for TCR.
2024-11-26 | GSE272790 | GEO
Project description:The response to DNA damage-stalled RNA polymerase II (RNAPIIo) involves the assembly of the transcription-coupled repair (TCR) complex on actively transcribed strands. The function of the TCR proteins CSB, CSA and UVSSA and the manner in which the core DNA repair complex, including transcription factor IIH (TFIIH), is recruited is largely unknown. Here, we define the assembly mechanism of the TCR complex in human isogenic knockout cells. We show that TCR is initiated by RNAPIIo-bound CSB, which recruits CSA through a newly identified CSA-interaction domain (CID). Once recruited, CSA facilitates the association of UVSSA with stalled RNAPIIo. Importantly, we find that UVSSA is the key factor that regulates the assembly of the TFIIH complex in a manner that is stabilized by CSB and CSA. We identify the UV-specific and independent interactors of UVSSA
2020-04-14 | PXD013572 | Pride
Project description:STK19 accelerates the clearance of lesion-stalled RNAPII to facilitate transcription-coupled DNA repair
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Project description:STK19 accelerates the clearance of lesion-stalled RNAPII to facilitate transcription-coupled DNA repair
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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:In adaptive immunity, transcription-coupled damage is introduced into antibody genes by activation-induced cytidine deaminase (AID) to diversify antibody repertoire. However, the coordination between transcription and DNA damage/repair remains elusive. Here, we find that transcription elongation factor 1 (ELOF1) stabilizes paused RNA polymerase II (RNAPII) at transcription barriers, providing a platform for transcription-coupled DNA damage/repair. Using a genetic screen, we discover that ELOF1 is required for AID targeting and that ELOF1 deficiency results in defective antibody class switch recombination and somatic hypermutation in mice. While downstream transcription-coupled repair factors are dispensable for AID-damage, ELOF1 mechanistically facilitates both transcription-coupled damage and repair by stabilizing chromatin-bound RNAPII. In ELOF1-deficient cells, paused RNAPII tends to detach from chromatin and fails to recruit factors to induce or repair DNA damage. Our study places ELOF1 at the center of transcription-coupled DNA metabolism processes and suggests a transition of RNAPII from elongation to a DNA damage/repair scaffold.
2025-03-01 | GSE284306 | GEO
Project description:In adaptive immunity, transcription-coupled damage is introduced into antibody genes by activation-induced cytidine deaminase (AID) to diversify antibody repertoire. However, the coordination between transcription and DNA damage/repair remains elusive. Here, we find that transcription elongation factor 1 (ELOF1) stabilizes paused RNA polymerase II (RNAPII) at transcription barriers, providing a platform for transcription-coupled DNA damage/repair. Using a genetic screen, we discover that ELOF1 is required for AID targeting and that ELOF1 deficiency results in defective antibody class switch recombination and somatic hypermutation in mice. While downstream transcription-coupled repair factors are dispensable for AID-damage, ELOF1 mechanistically facilitates both transcription-coupled damage and repair by stabilizing chromatin-bound RNAPII. In ELOF1-deficient cells, paused RNAPII tends to detach from chromatin and fails to recruit factors to induce or repair DNA damage. Our study places ELOF1 at the center of transcription-coupled DNA metabolism processes and suggests a transition of RNAPII from elongation to a DNA damage/repair scaffold.
2025-03-01 | GSE284305 | GEO