Studies conducted in the Colon Molecular and Cell Biology Laboratory are designed to identify and characterize the properties of genes and pathways involved in the development and progression of cancer of the colon and stomach. The laboratory has developed a gene discovery program in zebrafish to identify genes that govern intestinal development and is also using reverse genetic approaches in mice to evaluate findings in the context of mammalian models of gastrointestinal neoplasia.

The overall aim of the Colon Molecular and Cell Biology Laboratory is to provide novel insights into the genetic basis of cancer of the gastrointestinal tract. A number of cellular characteristics, including proliferation, widespread migration, vigorous angiogenesis and apoptosis are shared by cells and tissues in developing organisms and in cancers. We study the genetic regulation of the mechanisms underlying these important cellular processes in two vertebrate model systems.

We use reverse genetics in mice to establish a causal role between a predetermined mutation in a gene and its encoded protein and neoplastic development in the gastrointestinal tract. We focus on components of the Stat3, TGF β and Wnt pathways that are known to play important roles in development and cancer. Using mice carrying mutations in the interleukin 6 receptor, gp130, we have shown that insufficient signalling by the latent transcription factor Stat3 results in disruption of epithelial wound healing in the colon. In contrast, exaggerated Stat3 activation results in neoplasia in the stomach. We have demonstrated that this is in part a consequence of Stat3 hyper-activation of the Smad7 gene which makes gastric epithelial cells less responsive to the cytostatic effect of TGF β .

We can deliberately mis-express genes of interest in the epithelial cell layer of the small and large intestines using gene targeting. We employ a knock-in strategy to hijack the regulatory elements of an intestinal epithelium-specific locus (the A33 antigen gene). We find that over-expression of a constitutively active component of the Wnt signalling pathway ( D N- β cat) results in increased adenomatous polyp formation. In A33 antigen knock-in mice over-expressing the DNA methyltransferase, Dnmt3a, epigenetic suppression of a negative regulator of the Wnt pathway (Sfrp5) also leads to increased polyp formation. In order to mimic genetic mutations that occur randomly in somatic cells of the adult, we have developed a mouse strain expressing an inducible version of the bacterial DNA recombinase, Cre. When these mice are crossed with mice harbouring conditional (loxP-flanked) mutant genes the resulting mice can be used to determine the consequences of deliberately activating or inactivating those genes in the intestinal epithelium.

We use forward genetics in zebrafish to identify novel genes involved in endoderm migration and intestinal epithelium development, differentiation and death. Our studies focus on a panel of zebrafish mutants that emerged from a focused genetic screen conducted in the laboratory of Professor Didier Stainier at the University of California , San Francisco (UCSF). This screen was built around a unique transgenic line (gutGFP) in which GFP expression is restricted to the developing digestive organs. This background facilitated the identification of abnormalities in the size and morphology of the intestine, liver and pancreas in transparent zebrafish embryos. We then used histology to fully characterise the defects in intestinal morphology, differentiation and apoptosis and identified a panel of eleven mutants for further analysis. Positional cloning has resulted in the identification of the mutant genes responsible for the abnormal phenotypes in three of these interesting mutants

• Identifying genes that play a role in vertebrate intestinal development
•  Studying the oncogenic role of Stat3 in gastric cancer
•  Using conditional mice to model human colorectal cancer

Bollrath J, Phesse TJ, von Burstin VA, Putoczki T, Bennecke M, Bateman T, Nebelsiek T, Lundgren-May T, Canli O, Schwitalla S, Matthews V, Schmid RM, Kirchner T, Arkan MC, Ernst M, and Greten FR gp130-mediated Stat3 activation in enterocytes regulates cell survival and cell-cycle progression during colitis-associated tumorigenesis. Cancer cell. 15(2): 91-102 (2009).

de Jong-Curtain TA, Parslow AC, Trotter AJ, Hall NE, Verkade H, Tabone T, Christie EL, Crowhurst MO, Layton JE, Shepherd IT, Nixon SJ, Parton RG, Zon LI, Stainier DY, Lieschke GJ, and Heath JK Abnormal nuclear pore formation triggers apoptosis in the intestinal epithelium of elys-deficient zebrafish. Gastroenterology. 136(3): 902-11 (2009).

Ng A.N., de Jong-Curtain T.A., Mawdsley D.J., White S.J., Shin J, Appel B., Dong P.D., Stainier D.J., and Heath J.K. Formation of the digestive system in zebrafish: III Intestinal epithelium morphogenesis. Developmental Biology, 2005 Aug 23; [Epub ahead of print]. (2005)

Mawdsley D.J., Cooper H.M., Hogan B.M., Cody S.H., Lieschke G.J., and Heath J.K. The Netrin receptor Neogenin is required for neural tube formation and somitogenesis in zebrafish. Developmental Biology. 269: 302-15 (2004) Jenkins BJ et al Nature Medicine 2005

 Jenkins B.J., Grail D., Nheu T., Najdovska M., Wang B, Inglese M., McLoughlin R.M., Jones S.A., Topley N., Baumann H., Judd L.M., Giraud A.S., Boussioutas A., Zhu H-J., and Ernst M. Hyperactivation of Stat3 in gp130 mutant mice promotes gastric hyperproliferation and desensitizes TGF-beta signaling. Nature Medicine. Jul 24; [Epub ahead of print]: (2005)

 Tebbutt N.C., Giraud A.S., Inglese M., Jenkins B.J., Waring P.M., Clay F.J., Malki S., Grail D., Hollande F., Heath J.K., and Ernst M. Reciprocal regulation of gastrointestinal homeostasis by SHP2 and STAT-mediated trefoil gene activation in gp130 mutant mice. Nature Medicine. 8 (10): 1089-97 (2002)