James W. Dennis PhD Canada Research Chair Senior Investigator & Professor Department of Molecular Genetics Samuel Lunenfeld Research Institute Mount Sinai Hospital 600 University Ave. R988 Toronto, ON. M5G 1X5 Phone 416-586-8233

Research Interests

Golgi and metabolic regulation of cell surface proteins

Cells present an external face covered with glycoproteins, including receptors for cytokines and adhesion, and molecular transporters. Cells respond to multiple extracellular cues with intensities dependent on the levels of surface receptors. The information is integrated by intracellular signaling pathways and directs cell proliferation and differentiation. Asn- X-Ser/Thr sites are N-glycosylated, and the N-glycans are modified on transit through the Golgi, resulting in large molecular population of glycoforms for each gene product.  The N-glycans bind to mammalian lectins at the cell surface forming a multivalent lattice that protects the glycoproteins from loss to endocytosis and caveolae. Glycoform distributions are highly variable, and depend on tissue-specific regulation of the Golgi enzymes as well as metabolite flux into sugar-nucleotides, the basic building blocks for glycans. We are studying the interaction between Golgi pathways and the nutrient environment that regulate surface receptors critical to cancer, T cells mediated autoimmunity, and type II diabetes. We are using mouse models, human genetic data, high content cell imaging, and high throughput interference RNA, as well as computational models to reveal new properties of the network controlling surface glycoproteins.

Polarity in C. Elegans embryo

Sperm binding and entry defines the anterior-posterior axis in nematode, C. elegans, and two of the earliest events following fertilization are secretion of the chitinous eggshell and completion of meiosis.  Developmental regulation of glucosamine-6-phosphate N-acetyltransferase-2 (GNA-2) is required for synthesis of UDP-GlcNAc, the substrate for eggshell chitin synthesis by chitin synthase-1 (CHS-1).  The eggshell is essential for movement of the sperm pronucleus to the cortex, polar body extrusion, and polarization of the embryo, essential actin-dependent early events. Eggshell integrity requires chitin-binding proteins CEJ-1 and B0280.5, and presumably other glycoconjugates that may signal and provide a scaffold to organizes membrane domains as required for initiation of polarity. We are using RNAi screening and classical genetics to reveal the transmembrane glycoconjugates linking the eggshell to specific intracellular signaling pathways for cell polarity.  Dr. Charles Warren initiated the C. elegans work in our lab as a post doctoral fellow, and was the first to showed that a worm glycosyltransferase, gly-2, could rescue an Mgat5 defeciency in mammalian cells. Charles was assistant Professor at University of New Hampshire before his death in a paragliding accident Aug 2005, and is sorely missed by many friends.

Plk4/Sak in cell cycle regulation and carcinogenesis

Plk4/Sak, a member of the polo gene family of protein kinases, is cell-cycle regulated and required for centriole duplication and cytokinesis. Plk4-/- embryos arrest after gastrulation at E7.5, with cells blocked in late anaphase or telophase. Delays in cell cycle transitions are associated with hepatocellular carcinogenesis in Plk4+/- animals, suggesting that reduced Plk4 gene dosage increases the probability of mitotic errors and cancer development. Moreover, loss of heterozygosity (LOH) occurs frequently (~60%) at polymorphic markers adjacent to the Plk4 locus in human hepatoma.  We are studying Sak substrates and interacting proteins to understand the role of Plk4 in maintaining mitotic stability and preventing cancer. We are also using in vivo tumour models and studying human hepatoma specimens to further define Plk4/Sak functions at a molecular and cellular level.