Charles Spruck, Ph.D., Assistant Professor
Tumor Cell Biology Program

Dr. Spruck's Active Grants
Dr. Spruck's Publications
cspruck@skcc.org

Phone: (858)-450-5990, Ext. 387

Laboratory staff: Yingmeei Tan Ph.D., Zdenka Ellederova Ph.D., Sonia del Rincon Ph.D., Stefan Grotegut Ph.D., Dahui Sun

Research Interests:

Cell Cycle Regulation, Proteolysis, Cdk regulation, SCF ubiquitin ligases

Cancer occurs when the processes that regulate normal cell division become defective. Our laboratory is concerned with deciphering how cell division is regulated in normal cells and how alteration of these processes results in cancer.

Cyclin E. Cyclin E regulates the initiation of DNA synthesis in mammalian cells by activating a protein called cyclin-dependent kinase-2 (Cdk2). In normal cells, cyclin E protein levels are tightly regulated, peaking just prior to the initiation of DNA synthesis (G1/S phase boundary) and declining once DNA synthesis begins (S phase). However, in many human tumors cyclin E protein levels are elevated and/or deregulated relative to cell division, and this phenotype is associated with poor patient prognosis. Experimentally, deregulated cyclin E has been shown to induce a premature start of DNA synthesis and genetic instability in cultured cells, as well as tumor formation in mice. These studies suggest that deregulated cyclin E is oncogenic, but the mechanism by which this defect contributes to cancer is not understood. Our hypothesis is that the abnormal presence of cyclin E at inappropriate times during cell division could interfere with the execution of downstream events, thereby promoting cancer. Work in our laboratory is concerned with 1) determining how cyclin E becomes deregulated in human tumors, 2) identifying the cell division processes that are altered by deregulated cyclin E, and 3) identifying inappropriately phosphorylated substrates of deregulated cyclin E/Cdk2 kinase activity in human tumors.

Cks proteins. Cks proteins are small (9-18 kD), highly conserved proteins that bind Cdks (Cdk1, Cdk2, Cdk3). Studies in lower organisms (yeast and Xenopus) suggest that Cks proteins are essential for both the progression into and exit from mitosis. Although their precise function has remained elusive, studies in Xenopus suggests that Cks proteins may target cyclin/Cdk complexes to their appropriate substrates. In order to decipher the function of the two Cks orthologs in mammals (designated CKS1 and CKS2), mice deficient for each gene were created using targeted disruption strategies. These studies uncovered unique and unexpected roles for Cks proteins in both mammalian development and cell cycle regulation. Cks1 was found to be an essential cofactor in the degradation of a protein called p27 by the ubiquitin ligase SCFSkp2. Interestingly, p27 is a Cdk inhibitor that belongs to a class of proteins known as tumor suppressors and low levels of p27 in tumors have been shown to be a strong prognostic indicator of poor outcome. We are currently exploring the link between p27 and Cks1 in human tumors and determining whether the ubiquitination of other SCFSkp2 targets are also dependent on Cks1. On the other hand, Cks2 appears to play a specialized role in germ cell development. CKS2 deficient male and female mice were found to be sterile, with spermatocytes and oocytes arresting at metaphase I of the first meiotic division. Cks2 is a component of maturation promoting factor (MPF), consisting of cyclin B/Cdk1, which is the primary kinase complex that regulates meiosis in mammals. Our lab is attempting to determine the cause of the metaphase I arrest in CKS2 deficient germ cells by identifying potential substrates of MPF whose phosphorylation is dependent on Cks2.

Ubiquitin-mediated proteolysis and cancer. Ubiquitin is a small (76 amino acid), highly abundant protein that when covalently attached to other proteins in chains targets them for degradation. Ubiquitination is used by cells to regulate a variety of cellular process including differentiation, protein transport, DNA repair, as well as cell division. Our lab studies a ubiquitin ligase known as SCF, that has been implicated in cell division regulation. SCF ubiquitin ligases are multimeric protein complexes consisting of Cul1, Roc1, and Skp1 bound to one of approximately 50 different F-box proteins that provide substrate specificity. However, to date only a handful of F-box proteins have been extensively characterized. One well characterized F-box protein is hCdc4 that mediates the ubiquitination of cyclin E in mammalian cells. Interestingly, mutations in the hCDC4 gene have been found in a number of human tumor types suggesting that the deregulated cyclin E phenotype (see above) observed in tumors could be due to defective proteolysis. We are performing a comprehensive mutational and functional analysis of F-box proteins in human tumors to identify other ubiquitin-dependent proteolytic pathways that may contribute to human tumorigenesis.