Proteomics

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Posttranslational modifications regulate cellular responses upon UV-induced DNA damage


ABSTRACT: SUMOylation is a posttranslational protein modification which is characterized by the covalent attachment of a small 11kDa protein, called Small Ubiquitin-like MOdifier (SUMO). SUMOylation plays a pivotal role in a multitude of cellular pathways including cellular responses upon DNA damage. Here, we identified multiple proteins which are SUMOylated in U2OS cells in response to ultraviolet light (UV) irradiation and ionizing radiation (IR). We show that the SUMOylation response upon UV irradiation was more pronounced compared to the response upon IR. The major SUMOylation target upon UV-irradiation was the transcription-coupled nucleotide excision repair (TC-NER) protein, Cockayne Syndrome B (CSB). This protein plays an important role in the repair of UV-induced lesions in actively transcribed genes. In a second proteomic approach we identified SUMOylation-dependent and independent protein interactors of the N-terminus of CSB. Here, we uncovered that the affinity of multiple RNA polymerase-associated proteins towards CSB is influenced by SUMOylation. Finally, we set out to identify ubiquitination events upon UV-irradiation which are influenced by the CSA-ubiquitin ligase complex, which is also involved in TC-NER and is closely connected to CSB, because mutations in either CSA or CSB result in the same phenotype, Cockayne syndrome. We found that RPB1, the major subunit of RNA polymerase II, was ubiquitinated in a CSA-dependent manner upon UV which finally led to its degradation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo sapiens  

TISSUE(S): Cell Culture

DISEASE(S): Not Available

SUBMITTER: Frauke Liebelt  

LAB HEAD: Alfred Vertegaal

PROVIDER: PXD010609 | Pride | 2019-10-11

REPOSITORIES: pride

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Publications

Transcription-coupled nucleotide excision repair is coordinated by ubiquitin and SUMO in response to ultraviolet irradiation.

Liebelt Frauke F   Schimmel Joost J   Verlaan-de Vries Matty M   Klemann Esra E   van Royen Martin E ME   van der Weegen Yana Y   Luijsterburg Martijn S MS   Mullenders Leon H LH   Pines Alex A   Vermeulen Wim W   Vertegaal Alfred C O ACO  

Nucleic acids research 20200101 1


Cockayne Syndrome (CS) is a severe neurodegenerative and premature aging autosomal-recessive disease, caused by inherited defects in the CSA and CSB genes, leading to defects in transcription-coupled nucleotide excision repair (TC-NER) and consequently hypersensitivity to ultraviolet (UV) irradiation. TC-NER is initiated by lesion-stalled RNA polymerase II, which stabilizes the interaction with the SNF2/SWI2 ATPase CSB to facilitate recruitment of the CSA E3 Cullin ubiquitin ligase complex. Howe  ...[more]

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