CHIMERIC-TRANSGENIC MICE REPRESENT A POWERFUL TOOL FOR STUDYING HOW THE PROLIFERATION AND DIFFERENTIATION PROGRAMS OF INTESTINAL EPITHELIAL-CELL LINEAGES ARE REGULATED

An in vivo system has been developed for examining the effects of wild -type or mutant proteins on cell fate determination in the mouse intes tinal epithelium or on the proliferation and differentiation programs of its component epithelial lineages. This system takes advantage of t he fact that at the conclusion of gut morphogenesis, each intestinal c rypt is composed of a monoclonal population of cells descended from a single active multipotent stem cell, each villus is supplied by severa l monoclonal crypts, and the four principal cell types of the intestin al epithelium differentiate during a rapid, geographically well-organi zed migration along the crypt-to-villus axis. Embryonic stem (ES) cell s (129/Sv origin) are initially transfected with recombinant DNAs cons isting of a reporter of interest linked to transcriptional regulatory elements that control the cell lineage-specific, differentiation-depen dent, and axial patterns of expression of fatty acid binding protein g enes in the gut. Stably transfected ES cells are subsequently introduc ed into host C57BL/6 blastocysts to generate chimeric-transgenic mice. At the borders of ES cell-derived and host blastocyst-derived epithel ium, intestinal villi are found that are supplied by both ES cell- and host blastocyst-derived crypts. These villi can be rapidly identified in fixed whole-mount preparations of intestine using the alpha-L-fuco se-specific Ulex europaeus agglutinin type I (UEA-I) lectin. They appe ar striped because UEA-I recognizes a cell-surface carbohydrate polymo rphism between the inbred strains used to generate the chimeric animal s. The strength of this system derives from the fact that two gut epit helial populations can be compared and contrasted that occupy virtuall y identical positions along the crypt-to-villus and duodenal-to-coloni c axes within the same animal and differ only by the presence or absen ce of a single gene product. The band of blastocyst-derived epithelium in these striped, polyclonal villi can be used as an internal control to assess the biological effect of the transfected gene product produ ced in the adjacent stripe of ES-derived cells. The system can be used for either gain-of-function or loss-of-function experiments.