Project Summary Nucleotide excision repair (NER) is the major pathway to remove bulky DNA lesions induced by UV irradiation, environmental mutagens, and chemotherapeutic agents. Deficiency of genes involved in NER has been linked to Xeroderma Pigmentosum (XP) and skin cancer. There are two mechanisms to detect DNA damage by NER, one is global genome NER(GG-NER) and another is transcription-coupled NER(TC-NER). GG-NER occurs anywhere in the genome, whereas TC-NER is responsible for the accelerated repair of lesions in the transcribed strand of active genes. The both pathways are divided into early and late steps. The early step is the sequential actions, in which XP proteins recognize, unwind, and incise the DNA lesion. The latter step is identical in both mechanisms and is characterized by gap-filling repair synthesis, in which DNA replication proteins fill in the ~30 nucleotide gap, followed by ligation. Compared to the well-characterized early step, the molecular mechanism regulating the activation of gap-filling DNA synthesis at late step remains largely unknown. Even less is known about the physiological impact of such a regulatory pathway. And-1 is an acidic nucleoplasmic DNA-binding protein and its yeast ortholog, Ctf4, was originally identified as a critical gene for chromosome stability. Interestingly, yeast cells with depletion of Ctf4 gene are hypersensitive to UV lights, suggesting a role of And-1 in UV-induced DNA damage response. However, how And-1 regulates NER remains largely unknown. In this study, we now have extensive preliminary data demonstrating that And-1 is critical for NER by regulating gap-filling DNA synthesis. Our hypothesis is that And-1 regulates DNA polymerase activity at UV-lesion sites to activate gap-filling DNA synthesis at the late stage of NER. To test this hypothesis, we plan to pursue three specific aims. Aim 1: Determine the mechanism by which And-1 regulates DNA polymerase activity in NER. Aim 2: Determine the unique mechanism by which And-1 is recruited to UV-lesion sites. Aim 3: Determine the role of And-1 in NER and skin tumorigenesis using And-1 deficient mouse models. The completion of proposed studies will not only advance the field by uncovering a novel And-1-mediated pathway to regulate NER, but also provide us with the in vivo evidence to elucidate a novel role of And-1 in NER and skin tumor.