Persistent TFIIH binding to non-excised DNA damage causes cell and developmental failure

Muniesa-Vargas, Alba; Davó-Martínez, Carlota; Ribeiro-Silva, Cristina; van der Woude, Melanie; Thijssen, Karen L.; Haspels, Ben; Häckes, David; Kaynak, Ülkem U.; Kanaar, Roland; Marteijn, Jurgen A.; Theil, Arjan F.; Kuijten, Maayke M. P.; Vermeulen, Wim; Lans, Hannes

doi:10.1038/s41467-024-47935-9

Cited by 3 publications

(1 citation statement)

References 112 publications

(201 reference statements)

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“…Another possibility would be that these rare phenotypes of XP/CS, despite showing normal clearance of RNAPII, are caused by the accumulation of other repair intermediates. Indeed, recent work revealed that the absence of ERCC1-XPF or XPG, but not XPA, leads to persistent binding of TFIIH to non-excised DNA damage, correlating with developmental arrest and neuronal dysfunction in nematodes 41 .…”

Section: Discussionmentioning

confidence: 99%

Clearance of DNA damage-arrested RNAPII is selectively impaired in Cockayne syndrome cells

van der Meer,

Yakoub,

Nakazawa

et al. 2024

Preprint

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Arrest of elongating RNA polymerase II (RNAPII) at DNA lesions initiates transcription-coupled repair (TCR), involving the concerted action of specific TCR factors, followed by downstream nucleotide excision repair steps. Remarkedly, only congenital defects in the CSA or CSB genes cause the neurodegenerative disorder Cockayne syndrome, which is not observed with other TCR genes, despite their equal importance in TCR. An explanation for this discrepancy has been lacking. In this study, we developed an assay to track the fate of elongating RNAPII at sites of UV-induced DNA lesions. Employing this method on an isogenic collection of TCR knockout cells reveals a selective RNAPII clearance defect in cells defective in CSA or CSB, in contrast to knockouts of other TCR genes. Our findings provide evidence that a deficiency in RNAPII processing and prolonged transcription arrests in response to DNA damage, rather than compromised DNA repair, may underlie the Cockayne syndrome-like neurodegenerative phenotype.

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Section: Discussionmentioning

confidence: 99%

Clearance of DNA damage-arrested RNAPII is selectively impaired in Cockayne syndrome cells

van der Meer,

Yakoub,

Nakazawa

et al. 2024

Preprint

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Differential processing of RNA polymerase II at DNA damage correlates with transcription-coupled repair syndrome severity

Gonzalo-Hansen,

Steurer,

Janssens

et al. 2024

Nucleic Acids Research

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DNA damage severely impedes gene transcription by RNA polymerase II (Pol II), causing cellular dysfunction. Transcription-Coupled Nucleotide Excision Repair (TC-NER) specifically removes such transcription-blocking damage. TC-NER initiation relies on the CSB, CSA and UVSSA proteins; loss of any results in complete TC-NER deficiency. Strikingly, UVSSA deficiency results in UV-Sensitive Syndrome (UVSS), with mild cutaneous symptoms, while loss of CSA or CSB activity results in the severe Cockayne Syndrome (CS), characterized by neurodegeneration and premature aging. Thus far the underlying mechanism for these contrasting phenotypes remains unclear. Live-cell imaging approaches reveal that in TC-NER proficient cells, lesion-stalled Pol II is swiftly resolved, while in CSA and CSB knockout (KO) cells, elongating Pol II remains damage-bound, likely obstructing other DNA transacting processes and shielding the damage from alternative repair pathways. In contrast, in UVSSA KO cells, Pol II is cleared from the damage via VCP-mediated proteasomal degradation which is fully dependent on the CRL4CSA ubiquitin ligase activity. This Pol II degradation might provide access for alternative repair mechanisms, such as GG-NER, to remove the damage. Collectively, our data indicate that the inability to clear lesion-stalled Pol II from the chromatin, rather than TC-NER deficiency, causes the severe phenotypes observed in CS.

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Novel DNA Repair Inhibitors Targeting XPG to Enhance Cisplatin Therapy in Non-Small Cell Lung Cancer: Insights from In Silico and Cell-Based Studies

Manguinhas,

Serra,

Gil

et al. 2024

Cancers

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NSCLC is marked by low survival and resistance to platinum-based chemotherapy. The XPG endonuclease has emerged as a promising biomarker for predicting the prognosis of cisplatin-treated patients and its downregulation having been reported to increase cisplatin efficacy. This study presents an integrated strategy for identifying small molecule inhibitors of XPG to improve cisplatin therapy in NSCLC. A structure-based virtual screening approach was adopted, including a structural and physicochemical analysis of the protein, and a library of small molecules with reported inhibitory activities was retrieved. This analysis identified Lys84 as a crucial residue for XPG activity by targeting its interaction with DNA. After molecular docking and virtual screening calculations, 61 small molecules were selected as potential XPG inhibitors, acquired from the ChemBridge database and then validated in H1299 cells, a NSCLC cell line exhibiting the highest ERCC5 expression. The MTS assay was performed as a first screening approach to determine whether these potential inhibitors could enhance cisplatin-induced cytotoxicity. Overall, among the eight compounds identified as the most promising, three of them revealed to significantly increase the impact of cisplatin. The inherent cytotoxicity of these compounds was further investigated in a non-tumoral lung cell line (BEAS-2B cells), which resulted in the identification of two non-cytotoxic candidates to be used in combination with cisplatin in order to improve its efficacy in NSCLC therapy.

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Persistent TFIIH binding to non-excised DNA damage causes cell and developmental failure

Cited by 3 publications

References 112 publications

Clearance of DNA damage-arrested RNAPII is selectively impaired in Cockayne syndrome cells

Clearance of DNA damage-arrested RNAPII is selectively impaired in Cockayne syndrome cells

Differential processing of RNA polymerase II at DNA damage correlates with transcription-coupled repair syndrome severity

Novel DNA Repair Inhibitors Targeting XPG to Enhance Cisplatin Therapy in Non-Small Cell Lung Cancer: Insights from In Silico and Cell-Based Studies

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