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ERC-ID Report Summary

Project ID: 340988
Funded under: FP7-IDEAS-ERC
Country: Netherlands

Mid-Term Report Summary - ERC-ID (Excision Repair and chromatin interaction dynamics)

Already within the first half (2,5 years) of this ERC-granted project, named: “Excision Repair Chromatin Interaction Dynamics (Acronym: ERC-ID)”, exciting new progress has been made on the dissection on how the vital anti-aging and cancer-circumventing DNA repair process ‘Nucleotide Excision Repair’ (NER) is being regulated and controlled. The basic mechanism of NER is rather well known, however, how this process is controlled within the context of chromatin in the living cell, and living different tissues with respect to DNA damage signaling is basically unknown. The main results are summarized as follows:
1) New interactors identified of CSA. CSA is an essential factor for transcription-coupled NER (TC-NER). TC-NER is cricual process to circumvent cellular senescence and cell death induced by DNA lesions that block transcription. Inherited defects in TC-NER are associated with severe development and progeroid (extreme fast aging) symptoms in Cockayne syndrome patients, illustrating the importance and significance of the TC-NER process to clear endogenously (by e.g. metabolites) induced DNA damage. The new identified interacting proteins (i.e. molecular chaperone complex) are important for proper folding and localization of CSA. Other novel interactions will be further investigated, and has opened new research avenues.
2) We have identified a new gene implicated in non-photosensitive Trichthiodystrophy (TTD) in two non-related TTD patients. We showed that these mutations do not affect NER function, but specifically only interferes with proper transcription of terminally differentiated tissue.
Manuscripts describing the above are in an advance stage and will be soon submitted
3) We have dissected the role of the newly identified SUMO-dependent Ubiquitin ligase RNF111 in its function to modify the GG-NER DNA damage recognition complex XPC. We showed that RNF111-dpeendent ubiquitylation is required for efficient progression of the NER reaction (van Cuijk, Nature Comm., 2015)
4) We have successfully generated (by CRISPR/CAS9-mediated homologous gene-targeting) mouse embryonic stem cells that express from both alleles GFP-tagged RNA polymerase II’s (RNAPII) largest subunit (RPB1). These cells allow for the first time to quantitatively determine the fate of RNAPII in response to transcription-blocking DNA damage in living cells under physiological conditions. These ES are currently being used to generate GFP-tagged RPB1 mice, allowing analysis of the vital TC-NER process in different tissues.
5) We have generated transgenic nematodes C.elegans worms that express GFP-tagged NER factors. Next we developed procedures to locally inflict DNA damage in sub-nuclear area in individual cells in the living worm. This unique and unprecedented procedure allows us now for the first time to quantitatively determine NER activity in living cells within a living organism.
6) We have dissected the function of the ISWI chromatin remodeler SMARCA5 (SNF2H) in TC-NER and showed that SNF2H is essential for proper TC-NER complex assembly (Aydin, NAR, 2014).

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