The underlying cause of the most common inherited disease among Caucasians, cystic fibrosis, is a mutation in a cAMP-dependent Cl- channel. In addition to the respiratory problems associated with this disease, some patients also suffer from cholestasis (decreased bile flow resulting in Cirrhosis). In turn, this suggests that other cholestatic liver diseases such as primary biliary cirrhosis, primary sclerosing Cholangitis, sarcoidosis, liver transplant rejection and AIDS Cholangiopathy may also occur from or cause alterations in Cl- secretion. In biliary epithelial cells (BECs), another type of Cl- channel is regulated by Ca2+, in part, through the actions of Ca2+/calmodulin- dependent protein kinase II (CaMKII). The long-term goals of this project are to understand the mechanisms by which Ca2+ signals arising from extracellular stimuli are transduced into CaMKII activation and regulate Ca2+ dependent Cl- secretion in both normal and diseased states. In BECs, multiple isoforms of CaMKII are expressed which differ in their association domains. These domains are involved primarily in holoenzyme formation and subcellular localization. The hypothesis of this proposal is that the CaMKII subunits primary structures will determine the holoenzymes' structural and functional characteristics and that only a subset of these holoenzymes will regulate Cl secretion. To test this hypothesis the specific aims of this grant are to: (1) characterize the activities and holoenzyme structures of CaMKII isoforms overexpressed in a biliary epithelial cell line; (2) using subcellular fractionation and immunocytochemical methods, determine the subcellular localizations of CaMKII isoforms; and (3) using [125I] efflux and patch clamp recordings, determine which CaMKII isoforms regulate Ca2+ dependent Cl- secretion. These experiments are unique since they will identify and characterize specific CaMKII isoforms involved in regulating Ca2+-dependent Cl- secretion.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29DK051080-05
Application #
6177509
Study Section
Nutrition Study Section (NTN)
Program Officer
Serrano, Jose
Project Start
1996-04-01
Project End
2001-03-31
Budget Start
2000-04-01
Budget End
2001-03-31
Support Year
5
Fiscal Year
2000
Total Cost
$110,273
Indirect Cost
Name
Indiana University-Purdue University at Indianapolis
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
603007902
City
Indianapolis
State
IN
Country
United States
Zip Code
46202
McGill, J M; Yen, M S; Cummings, O W et al. (2001) Interleukin-5 inhibition of biliary cell chloride currents and bile flow. Am J Physiol Gastrointest Liver Physiol 280:G738-45
Kwiatkowski, A P; McGill, J M (2000) Alternative splice variant of gamma-calmodulin-dependent protein kinase II alters activation by calmodulin. Arch Biochem Biophys 378:377-83
McGill, J M; Kwiatkowski, A P (1998) Cholestatic liver diseases in adults. Am J Gastroenterol 93:684-91