The long range goal is to understand the mechanisms that regulate the biosynthesis of phosphatidylcholine and the 1,2 - dipalmitoyl species in the lung and during lung development. The dipalmitoyl species of phosphatidylcholine is an important component of pulmonary surfactant, a substance necessary for lung function. Since the control of phosphatidylcholine biosynthesis is relatively unknown in any tissue, a comprehensive investigation of the basic regulatory mechanisms is fundamental to studies on the control during lung development. The research in this proposal is guided by the hypothesis that the biosynthesis of phosphatidylcholine is regulated by the supply of both diacylglycerol and CDP choline which in turn are coordinated by the reversible formation of a multienzyme complex on the endoplasmic reticulum. This complex would contain cytidylyltranserase, phosphatidate phosphohydrolase and choline-phosphotransferase. The present studies focus on the cytidylyltransferase component and will 1) provide detailed information about the molecular properties and subcelluar forms of cytidylyltransferase, 2) define the biochemistry of the translocation of cytidylyltransferase to membrane binding sites 3) characterize the binding domains on the membrane and 4) explore the possible existence of a multienzyme complex formed as a result of the translocation. Antibodies for cytidylyltranserase will be used to locate subcellular forms, to develope immunoassay for cytidylyltranserase and to isolate native forms of the enzyme. The mechanism which modulate translocation will be studied using in vitro systems and hepatocytes and HeLa cells in culture. An important part of these studies is to determine whether cytosolic or membrane components function in the regulation of the activity and subcellular location of cytidylyltransferase.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
2R01HD002871-18A3
Application #
3310169
Study Section
Biochemistry Study Section (BIO)
Project Start
1979-05-01
Project End
1992-11-30
Budget Start
1987-12-01
Budget End
1988-11-30
Support Year
18
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Type
Schools of Medicine
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Weinhold, P A; Feldman, D A (1995) Fatty acids promote the formation of complexes between choline-phosphate cytidylyltransferase and cytidylyltransferase binding protein. Arch Biochem Biophys 318:147-56
Weinhold, P A; Charles, L; Feldman, D A (1994) Regulation of CTP: phosphocholine cytidylyltransferase in HepG2 cells: effect of choline depletion on phosphorylation, translocation and phosphatidylcholine levels. Biochim Biophys Acta 1210:335-47
Feldman, D A; Weinhold, P A (1993) Identification of a protein complex between choline-phosphate cytidylyltransferase and a 112-kDa protein in rat liver. J Biol Chem 268:3127-35
Weinhold, P A; Charles, L; Rounsifer, M E et al. (1991) Control of phosphatidylcholine synthesis in Hep G2 cells. Effect of fatty acids on the activity and immunoreactive content of choline phosphate cytidylyltransferase. J Biol Chem 266:6093-100
Weinhold, P A; Charles, L G; Feldman, D A (1991) Microsomal CTP:choline phosphate cytidylyltransferase: kinetic mechanism of fatty acid stimulation. Biochim Biophys Acta 1086:57-62
Rooney, S A; Smart, D A; Weinhold, P A et al. (1990) Dexamethasone increases the activity but not the amount of choline-phosphate cytidylyltransferase in fetal rat lung. Biochim Biophys Acta 1044:385-9
Feldman, D A; Rounsifer, M E; Charles, L et al. (1990) CTP:phosphocholine cytidylyltransferase in rat lung: relationship between cytosolic and membrane forms. Biochim Biophys Acta 1045:49-57
Weinhold, P A; Rounsifer, M E; Charles, L et al. (1989) Characterization of cytosolic forms of CTP: choline-phosphate cytidylyltransferase in lung, isolated alveolar type II cells, A549 cell and Hep G2 cells. Biochim Biophys Acta 1006:299-310
Ide, H; Miller, J C; Weinhold, P A (1988) Ethanolaminephosphotransferase in rat lung: selectivity for endogenous and exogenous diacylglycerol. Biochim Biophys Acta 960:119-24