The C-terminal Region and SUMOylation of Cockayne Syndrome Group B Protein Play Critical Roles in Transcription-coupled Nucleotide Excision Repair

Abstract

Cockayne syndrome (CS) is a recessive disorder that results in deficiencies in transcription-coupled nucleotide excision repair (TC-NER), a subpathway of nucleotide excision repair, and cells from CS patients exhibit hypersensitivity to UV light. CS group B protein (CSB), which is the gene product of one of the genes responsible for CS, belongs to the SWI2/SNF2 DNA-dependent ATPase family and has an ATPase domain and an ubiquitinbinding domain (UBD) in the central region and the C-terminal region, respectively. The C-terminal region containing the UBD is essential for the functions of CSB. In this study, we generated several CSB deletion mutants and analyzed the functions of the C-terminal region of CSB in TC-NER. Not only the UBD but also the C-terminal 30-amino acid residues were required for UV light resistance and TC-NER. This region was needed for the interaction of CSB with RNA polymerase II, the translocation of CS group A protein to the nuclear matrix, and the association of CSB with chromatin after UV irradiation. CSB was modified by small ubiquitin-like modifier 2/3 in a UV light-dependent manner. This modification was abolished in a CSB mutant lacking the C-terminal 30 amino acid residues. However, the substitution of lysine residues in this region with arginine did not affect SUMOylation or TC-NER. By contrast, substitution of a lysine residue in the N-terminal region with arginine decreased SUMOylation and resulted in cells with defects in TC-NER. These results indicate that both the most C-terminal region and SUMOylation are important for the functions of CSB in TC-NER.

The Cellular Response to Transcription-Blocking DNA Damage

“…CUL4 DDB2 –XPC and CUL4 CSA –CSB may thus represent functionally related enzyme–substrate constellations required for GG-NER and TC-NER, respectively. Both XPC and CSB also appear to be deubiquitylated by USP7 30 , 39 , 40 (see below) and regulated by SUMOylation 37 , 41 , further underscoring the analogies ( Figure 2 ).

Figure 2 Analogous Regulation of Transcription-Coupled Nucleotide Excision Repair (TC-NER) and General Genome Repair (GG-NER) by Ubiquitin.

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ATM and CDK2 control chromatin remodeler CSB to inhibit RIF1 in DSB repair pathway choice

“…The WHD is a versatile domain that is implicated in protein-DNA and protein–protein interactions 50 . Recent studies suggest that the last 30 amino acids of CSB is necessary for its interaction with RNAPII in transcription-coupled UV repair 51 . Our finding that CSB interacts with RIF1 through its WHD in DSB repair supports the notion that the CSB WHD acts as a protein–protein interaction module to mediate its interaction with different partners depending upon the type of DNA repair process.…”

Transcription-coupled repair: an update

“…CSB recruits the CSA complex, NER factors and chromatin remodelers such as p300 to sites of arrested RNAPII, and it has been considered the master coordinator of TCR in humans, with roles similar to those carried out by Mfd in E. coli . The C-terminal region of CSB, essential for TCR, is required for interaction with RNAPII and for translocation of CSA to the nucleus, and is modified by small ubiquitin-like modifier (SUMO) 2/3 (Sin et al 2016). CSB colocalizes with lesions other than those induced by UV, such as interstrand crosslinks induced by trioxalen, angelicin monoadducts, double-strand breaks, and oxidative damage (Iyama and Wilson 2016), and is involved in the repair of endogenously generated cyclopurines in mouse tissues (Kirkali et al 2009).…”