Nucleotide excision repair (NER) is a highly conserved DNA repair pathway that functions to protect the genome from the detrimental effects of DNA lesions introduced by endogenous and environmental sources, including UV light. Defects in the NER system result in genomic instability and the development of cancers. The initial step in NER requires lesion recognition, and is performed by the heterodimeric DNA damage binding proteins XPC-HR23B and UV-DDB (composed of the p48 and p127 proteins). The DDB2 (encoding the p48 protein) and XPC genes are transcriptionally induced by the p53 tumor suppressor gene following DNA damage. In addition, both p48 and XPC are post-translationally regulated by the ubiquitin-proteasome degradation pathway. Covalent attachment of mono- and polyubiquitin chains to cellular proteins has emerged as a predominant cellular regulatory mechanism with roles in cell division, signal transduction, endocytic trafficking, and DNA repair. Thus, the DNA damage recognition step of NER is highly regulated through transcriptional and post-transcriptional mechanisms. The long-term objectives of this project are to understand how the ubiquitin-proteasome pathway regulates human NER. The hypothesis is that UV-induced ubiquitination of the NER proteins t>48 and XPC specifically regulate their degradation, protein- and DNA damage binding activities, and affect on NER. These studies will have broad applicability to understanding the role of DNA repair in health and disease, and provide important insight into the role of NER in the etiology of human cancer. The project will be approached through the following specific aims. 1. To determine if the human DNA damage recognition factors p48 and XPC are ubiquitinated and if this process is regulated by HR23B. 2. To determine whether ubiquitination of these NER proteins is required for DNA damage binding activity, in vivo. 3. To determine the functional consequences of ubiquitination on NER activity in human cells.