This is the second competing renewal of CA65572, an investigation that is aimed at the long-term goal of understanding mammalian cell cycle control and tumorigenesis. Looking back at the journey of the last 10 years, we are pleased to see the contribution we made to the field and, more remarkably, the advancement achieved collectively by others working in this field. The focus of our own research has moved forward from determining the function and regulation of the newly discovered first CDK inhibitor, p21 in 1993, to the discovery and study of the Arf-Mdm2-p53 pathway during the second funding period. The research proposed in this application will focus on determining the cytoplasmic control of p53, an area that is emerging as a critical regulatory step in p53 regulation and tumor suppression. Leading to this proposal are three findings we and others made recently: (i) broad utilization of nuclear export by multiple stress checkpoint pathways in regulating p53;(ii) the discovery of ROC1 and ROC2 as the catalytic subunit of cullin family E3 ubiquitin ligases, and (iii) two ROC-dependent ligases, PARC and CUL7, bind to p53 and localize in the cytoplasm. Four specific aims are proposed to determine: (Aim I) The functional consequence of PARC-p53 interaction, (Aim II) The mechanisms of PARC and CUL7 function, (Aim HI) The function of DOC domain in PARC and CUL7, and (Aim IV) The function of RBR domain of PARC. This proposal combines our strength and success in four areas: genetic and tumor analysis in mouse with targeted mutations;biochemical characterization of ubiquitin ligases;cellular studies of p53 export and p53 checkpoint pathways, and IP-Mass Spec-based protein complex characterization. We anticipate that these comprehensive and multi-disciplinary studies will lead to a better understanding of cytoplasmic control of p53, the function and mechanism of the ubiquitin pathway and the process of tumor suppression.