Differentiation and Development in Gastric Mucosa

Variations on a common pattern of self-renewal are found throughout gastrointestinal mucosa. Rapidly dividing, immature cells occur in localized proliferative zones within the crypts in intestine and the glands in stomach. The majority of the cells produced in these zones migrate upward toward the surface, differentiate into one or more mature cell types, and are shed after lifetimes of a few days. A smaller number of cells migrate downward into the base of the gland or crypt, where they differentiate into longer-lived cell types. The mechanisms by which growth is regulated are unknown.

We are currently using transgenic mouse models to investigate lineage and trophic relationships among cells of the gastric mucosa. Using the promoter from the gastric H,K-ATPase beta subunit gene, we have generated a line of transgenic mice which express the herpes simplex thymidine kinase (TK) gene in parietal cells. Administration of ganciclovir to these animals ablates the parietal cells and produces profound disorganization of mucosal structure. Following removal of the drug, the mucosa regenerates and regains its normal morphology. By investigating cell loss and recovery in this system, we hope to gain an understanding of the role of parietal cells in these processes.

A second area of interest is the relationship between structure and function of membrane transport proteins, particularly Na,K-ATPase. Na,K-ATPase is the ubiquitous transport ATPase responsible for maintaining the sodium and potassium gradients necessary for action potentials in excitable cells, sodium-dependent transport across epithelia, and cell volume homeostasis. The enzyme is composed of an alpha subunit, which has multiple transmembrane spans, and a glycosylated beta subunit containing a single membrane span. We are applying a variety of genetic and biochemical approaches to the issues of structure and function of the alpha subunit. In one approach, we have constructed a panel of Na,K-ATPase alpha subunit cDNAs which contain insertions encoding an epitope recognized by an antibody. It is possible to map the transmembrane topology of the alpha subunit by analyzing the accessibility of the added epitope in permeabilized or intact cells. The epitope tagged polypeptides can also be used as a tool to monitor the assembly of alpha and beta subunits. We are currently attempting to map the sodium-binding domains of the Na,K-ATPase alpha subunit using a panel of chimeric cDNAs which incorporate segments of the gastric H,K-ATPase. The basis of these experiments is that the Na,K- and H,K-ATPases are highly similar mechanistically and structurally, but that Na,K-ATPase transports sodium ions, while H,K-ATPase transports protons in exchange for potassium ions.