Our research is concentrated on gaining fundamental knowledge of the biology of cells in normal and disease settings. We have chosen to focus on the mechanisms underlying immune responses and tumorigenesis. With this broad agenda in mind, we have initiated several complementary programs. Many of these projects have evolved from the production and analysis of genetically engineered mouse strains.
Biology of the Immune System
The various compartments of the immune system make up what is perhaps the most intriguing and intricate cellular network aside from the nervous system. Through the targeted mutation of individual genes, our laboratory has endeavoured to dissect the function of its various components, one molecule at a time. To achieve this goal, we have generated mice lacking the key receptors, co-receptors and signaling molecules that participate in T cell development, activation and differentiation. To understand signal transduction through the TCR complex, animals lacking the tyrosine kinase Lck, the tyrosine phosphatase CD45, or the adaptors Bcl-10 or MALT1 have been generated. Bcl10 and MALT1 appear to cooperate in a signaling complex that specifically links antigen receptor engagement to activation of the cell survival transcription factor NF-kB. The activation of NF-kB depends on the degradation of an inhibitory binding protein called IkB. The role that Bcl-10 and MALT1 play in the signaling of a wide variety of other non-antigen specific receptors are being investigated.
Biology of Programmed
Apoptosis is the process of programmed cell death essential for normal embryonic development, resolution of immune responses, and defence against tumorigenesis. We have generated mice with defects in several molecules associated with apoptotic pathways, including TRAF-2, FADD, Apaf-1, caspase-3 and caspase-9. Analyses of these mutant animals have revealed roles for these molecules in lymphocyte ontogeny, activation, effector function and cell death, as well as for susceptibility to autoimmune diseases. In addition, it has become clear that multiple apoptotic pathways exist that are stimulus- and tissue-specific. These pathways show differential requirements for Apaf-1 and the caspases. Study of mice deficient for TNFR revealed that, depending on the adaptors recruited to the cytoplasmic tail of this receptor, signaling for either survival or apoptosis can be transduced. We have also examined other molecules involved in the regulation of apoptosis, e.g. TRADD.
The Pathogenesis of Cancer
Genetic susceptibility factors are associated with most types of cancer, but few of these genes have been identified. We have generated mutant mouse strains with engineered modifications of genes shown to be strongly linked to cancer. We have also adopted comparison genomic hybridization array screening as well as a Drosophila genetic modifier screening methods to identify novel genes linked to cancer.
One of our areas of interest is DNA repair. We studied mutant mice lacking MSH2, a component of the DNA mismatch repair machinery. MSH2-deficient mice spontaneously develop tumours early in life, making them a useful model for the study of tumorigenesis, carcinogens and anti-cancer agents. Studies of mice lacking the breast cancer susceptibility genes Brca1 or Brca2 suggest that these proteins have roles in pathways controlling DNA damage repair and the maintenance of genome stability.
One of the most important tumour suppressor genes (TSG) for human cancer is p53, a multi- functional protein that induces apoptosis or cell cycle arrest in cells with damaged DNA. We identified Chk2 as a kinase that phosphorylates p53 and stabilizes it, allowing it to carry out its anti-tumorigenic functions. We also generated mice lacking HIPK1, a kinase that binds to p53 and appears to modulate its transcriptional activity. HIPK1 may promote oncogenesis if dysregulated.
Our work on TSG has recently focussed on PTEN and genes with which it interacts. Studies of mice lacking PTEN indicated that PTEN negatively regulates the PKB/AKT cell survival pathway. PTEN-deficient cells are resistant to agents inducing apoptosis, becoming immortal and thus susceptible to acquiring additional mutations that lead to tumorigenesis. Loss of heterozygosity for PTEN in heterozygous mice results in a high incidence of tumours, especially T cell lymphomas. We have also undertaken to uncover genes that influence PTEN and p53 functions using phenotype rescue experiments in Drosophila. We identified PARK7 (DJ-1), an early onset Parkinson gene as a protein that regulates PTEN functions and oxidative stress. As well, we discoved that a rarely studied fatty acid transporter, CPT1C, regulates cell survival in tumour cells under metabolic stress. Finally, we are also studying the role of RhoC in metastasis. The functions of these genes are under active investigation in the hopes of identifying new targets for cancer drug therapy.
- David Cescon
- Christine Chio
- Isaac Harris
- Ying-ju Jang
- Val Lapin
- Amy Lin
- David McIlwain
- Elize Shirdel
- Stephanie Sue
Selected References:Link to Pubmed Publications
de la Pompa, J., Timmerman, L.A., Takimoto, H., Hoshida, H., Elia, A.J., Samper, E., Potter, J., Wakeham, A., Marengère, L., Langille, B.L., Crabtree, G.R. and Mak, T.W. 1998. Role of the NF-Atc transcriptiion factor in morphogenesis of cardiac valves and septum. Nature 392: 182-186.
Yeh, W-C, de la Pompa, J.L., McCurrach, M.E., Shu, H-B, Elia, A.J., Shahinian, A., Ng, M., Wakeham, A., Khoo, W., Mitchell, K., El-Deiry, W.S., Lowe, S.W., Goeddel, D.V. and Mak, T.W. 1998. FADD: Essential for Embryo Development and Signaling from Some, But Not All Inducers of Apoptosis. Science 279: 1954-1958.
Hakem, R., Hakem, A., Duncan, G.S., Henderson, J.T., Woo, M., Soengas, M.S., Elia, A., de la Pompa, J-L., Kagi, D., Khoo, W., Potter, J., Yoshida, R., Kaufman, S.A., Lowe, S.W., Penninger, J.M. and Mak, T.W. 1998. Differential Requirement for Caspase 9 in Apoptotic Pathways In Vivo. Cell 94: 339-352.
Yoshida, H., Kong, Y-Y., Yoshida, R., Elia, A.J., Hakem, A., Hakem, R., Penninger, J.M. and Mak, T.W. 1998. Apaf-1 is required for mitochondrial pathways of apoptosis and brain development, but not Fas-induced apoptosis. Cell 94: 739-750.
Stambolic, V., Suzuki, A., de la Pompa, J.L., Brothers, G.M., Mirtsos, C., Sasaki, T., Ruland, J., Penninger, J., Siderovski, D.P. and Mak, T.W. 1998. Negative regulation of PKB/Akt-dependent cell survival by the tumor suppressor PTEN. Cell 95: 29-39.
Hirao, A., Kong, Y-Y., Matsuoka, S., Wakeham, A., Ruland, J., Yoshida, H., Liu, D., Elledge, S.J. and Mak, T.W. 2000 DNA damage-induced activation of p53 by the checkpoint kinase chk2. Science 287: 1824-1827.
Tafuri, A., Shahinian, A., Bladt, F., Yoshinaga, S., Jordana, M., Wakeham, A., Boucher, L-M., Duncan, G., Bouchard, D., Chan, V., Odermatt, B., Ho, A., Itie, A., Horan, T., Whoriskey, J.S., Pawson, A.J., Penninger, J.M., Ohashi, P. and Mak, T.W. 2001. Essential Role of ICOS in Effective Helper T cell Responses. Nature 409: 105-109.
Ruland, J., Duncan, G.S., Elia, A., del Barco Barrantes, I., Nguyen, L., Plyte, S., Millar, D.G., Bouchard, D., Wakeham, A., Ohashi, P.S. and Mak, T.W. 2000. Bc110 is a positive regulator of antigen receptor-induced activation of NF-B and neural tube closure. Cell 104: 33-42.
Backman, S.A., Stambolic, V., Suzuki, A., Haight, J., Elia, A., Pretorius, J., Tsao, M-S., Shannon, P., Bolon, B., Ivy, G.O., and Mak, T.W. (2001) Deletion of Pten in mouse brain causes seizures, ataxia and defects in soma size resembling Lhermitte-Duclos disease. Nature Genetics 29: 396-403.
Hakem, A., Sanchez-Sweatman, O., You-Ten, A., Duncan, G., Wakeham, A., Khokha, R., and Mak, T.W. (2005) RhoC is dispensable for embryogenesis and tumor initiation but essential for metastasis. Genes & Dev. 19: 1974-1979.
Kim, R.H., Peters, M., Shi, W., Pintilie, M., DeLuca, C., Liepa, J., Zhou, L., Snow, B., Binari, R.C., Manoukian, A.S., Liu, F-F., Tsao, M-S. and Mak, T.W. (2005) DJ-1, a novel regulator of the tumor suppressor PTEN. Cancer Cell 7: 263-273.
Hao, Z., Duncan, G.S., Chang, C-C., Elia, A., Fang, M., Wakeham, A., Okada, H., Calzascia, T., Jang, Y-J., You-Ten, A., Yeh, W-C., Ohashi, P., Wang, X., and Mak, T.W. (2005) Specific Ablation of the Apoptotic Function of Cytochrome c in Mice Leads to Impaired Lymphocyte Homeostasis. Cell 121: 579-91.
You, H., Pellegrini, M., Tsuchihara, K., Yamamoto, K., Hacker, G., Erlacher, M., Villunger, A., and Mak, T.W. (2006) FOXO3a-dependent regulation of Puma in response to cytokine/growth factor withdrawal. J. Exp. Med. 203: 1657-63.
Bakker, W., Harris, I., and Mak, T.W. (2007) Stimulation of CITED2 transcription by FOXO3a inhibits HIF1-induced apoptosis during hypoxia. Mol Cell 28:941-953 Mak, T.W. (2007) Gene Targeting in Embryonic Stem Cells Scores a Knockout in Stockholm. Cell 131:1027-1031