The Role And Mechanism Of Mitotic Regulator Nlp In DNA Damage Repair

Cellular response to DNA damage, induced by ionizing radiation (IR), UV radiation and chemical carcinogens is critical for maintenance of genomic fidelity. Defects of DNA repair often result in genomic instability and malignant cell transformation, leading to generation of tumor. Current achievements in research on DNA repair provide a large number of basis for cancer treatment. The mechanisms of DNA repair are complicated, and have been one of the hot research topic.Nlp (ninein-like protein) has been characterized as an centrosomal protein, which is related to BRCA1. A lot of research work reveals that Nlp is involved in a variety of biological processes, including microtubules orgnization, spindle formation, centrosome maturation and cytokinesis. Although Nlp is an important cell cycle regulator that is required for proper mitotic progression, other aspects of function of Nlp is not clear.In previous study, after H1299 cells treated with UV-irradiation, we found that the subcellular localization of Nlp was changed obviously, suggesting that Nlp might be involved in DNA damage response induced by UV radiation. In order to verify the hypothesis, host cell reactivation (HCR) activity and removal efficiency of CPDs were detected. We demonstrate that Nlp is able to improve nuclear excision repair (NER) activity. To further reveal the function of Nlp, we used two kinds of cell lines, a pair of MEFs and newborn mice skin derived from Nlp trans genic and wild type mice. Through several assays, including cell viability, colony formation, apoptosis and in situ apoptosis assay, we find that Nlp could protect cells against UV radiation. To explore the related mechanisms, H1299 cells underwent UV local irradiation, which reveals Nlp positioning on the sites of DNA damage after UV irradiation. Then, many experiments were conducted, including nuclear protein extraction, western blotting, co-IP, GST pull-down. The results illustrate that upon UV radiation, Nlp is translocated to nucleus, which is mediated by NPM1 in a p53-independent manner. And C-terminal (1030-1382) of Nip is sufficient and necessary for its interaction with NPM1 and nuclear import. Furthermore, Nlp interacts with XPA and ERCC1 respectively, and enhances their association. In addition, in view of a clear correlation between UV radiation and the incidence of skin cancer, we find Nlp expression level in skin cancer samples is significantly lower than the normal counterpart with immunohistochemical assay, indicating that dysregulated Nlp is related to the development of human skin cancers.Taken together, this study identifies mitotic regulator Nlp as a new and important member of NER pathway, and thus provides novel insights into understanding on regulatory machinery involved in NER...