The long range goal of these studies is to understand the molecular organization, diversity and regulation of tight junctions of the gastrointestinal tract and liver. The tight junction forms the molecular seal limiting movement of water and solutes between epithelial cells thus is a fundamental determinant of transport properties. Dynamic changes in its sealing properties occur during development, during crypt to villus differentiation of enterocytes, and in response to physiologic stimuli and hormones. A breakdown in junction integrity contributes to numerous pathologic processes including some forms of hepatic cholestasis and diarrheal disease. There is evidence that the tight junction also serves as a barrier to maintain the separation of biochemically distinct apical and basolateral membrane domains. Despite its obvious importance to epithelial function in the liver and gut, molecular characterization of the junction has been impossible until the identification of its first protein component, ZO-1. ZO-1 appears to be a very important component of the junction since it is present in all junction, is tightly associated with the cytoplasmic surface of membrane contacts and there is a correlation between ZO-1 isoforms and junction morphology. ZO-1 is not abundant and has proven difficult to isolate and study thus, the proposed approaches rely on molecular biologic techniques using recombinant fusion proteins and cultured epithelial cell transfection studies to circumvent these limitation. The specific goals are to increase basic knowledge of ZO-1 by sequencing its full length human cDNA. The proteins structural/functional organization will be defined by seeking membrane binding activity is fusion proteins as well as transfected sequences. We will test the hypothesis that an intact junction is required to maintain cell polarity by disrupting the junction in cultured cells with transfected ZO-1 sequences. Finally we will investigate the hypothesis that ZO-1 isoforms, generated by alternate RNA splicing, contribute to junction diversity throughout the GI tract and are altered in extrahepatic cholestasis which is accompanied by junction disruption.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK045134-04
Application #
2144374
Study Section
General Medicine A Subcommittee 2 (GMA)
Project Start
1992-02-01
Project End
1996-01-31
Budget Start
1995-02-01
Budget End
1996-01-31
Support Year
4
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Yale University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
082359691
City
New Haven
State
CT
Country
United States
Zip Code
06520
Van Itallie, Christina M; Mitic, Laura L; Anderson, James M (2011) Claudin-2 forms homodimers and is a component of a high molecular weight protein complex. J Biol Chem 286:3442-50
Van Itallie, Christina M; Fanning, Alan S; Holmes, Jennifer et al. (2010) Occludin is required for cytokine-induced regulation of tight junction barriers. J Cell Sci 123:2844-52
Anderson, James M; Van Itallie, Christina M (2009) Physiology and function of the tight junction. Cold Spring Harb Perspect Biol 1:a002584
Van Itallie, Christina M; Holmes, Jennifer; Bridges, Arlene et al. (2009) Claudin-2-dependent changes in noncharged solute flux are mediated by the extracellular domains and require attachment to the PDZ-scaffold. Ann N Y Acad Sci 1165:82-7
Van Itallie, Christina M; Fanning, Alan S; Bridges, Arlene et al. (2009) ZO-1 stabilizes the tight junction solute barrier through coupling to the perijunctional cytoskeleton. Mol Biol Cell 20:3930-40
Van Itallie, Christina M; Holmes, Jennifer; Bridges, Arlene et al. (2008) The density of small tight junction pores varies among cell types and is increased by expression of claudin-2. J Cell Sci 121:298-305
Van Itallie, Christina M; Betts, Laurie; Smedley 3rd, James G et al. (2008) Structure of the claudin-binding domain of Clostridium perfringens enterotoxin. J Biol Chem 283:268-74
Jovov, Biljana; Van Itallie, Christina M; Shaheen, Nicholas J et al. (2007) Claudin-18: a dominant tight junction protein in Barrett's esophagus and likely contributor to its acid resistance. Am J Physiol Gastrointest Liver Physiol 293:G1106-13
Robertson, Susan L; Smedley 3rd, James G; Singh, Usha et al. (2007) Compositional and stoichiometric analysis of Clostridium perfringens enterotoxin complexes in Caco-2 cells and claudin 4 fibroblast transfectants. Cell Microbiol 9:2734-55
Van Itallie, Christina M; Anderson, James M (2006) Claudins and epithelial paracellular transport. Annu Rev Physiol 68:403-29

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