The long-term goal of this project is to investigate the regulation of hepatic folate-dependent one-carbon (1-C) metabolism; specifically to investigate the supply and oxidation of 1-C units from serine, glycine, choline, histidine and formate. A model is proposed for the unidirectional flow of 1-C units generated from serine, glycine and choline in mitochondria via 10-formyltetrahydrofolate (10-HCO-H4PteGlu) and delivered to the cytosol as formate, in adult liver. The key enzyme in the mitochondrial conversion of folate-linked 1-C units to formate is a mitochondrial form of C1-tetrahydrofolate synthase (mC1-THFS), and enzyme that has not been purified or characterized from mitochondria. Formate is the predominant source of 1-C units in cytosol and is assimilated into the cytosolic folate pool by conversion to 10-HCO-H4PteGlu and conversion to other folate forms by cytosolic (c) C1-THFS. Cytosolic and mitochondrial 10- HCO-H4PteGlu pools are reservoirs of 1-C units that supply the biosynthetic folate-dependent reactions or are oxidized to H4PteGlu and CO2 by isozymes of 10-formyltetrahydrofolate dehydrogenase (FDH). NEUT2 homozygous mice lack both cytosolic and mitochondrial isozymes of FDH and are unable to oxidize 1-C units as 10-HCO-H4PteGlu to CO2 and H4PteGlu. Lack of FDH in homozygous NEUT2 mice results in expanded 10-HCO-H4PteGlu pools and diminished H4PteGlu pools. The changes in the H4PteGlu pools correlates with changes in the protein levels of liver cC1-THFS. It is hypothesized that expression of cC1-THFS is regulated at the transcriptional level by the cytosolic level of H4PteGlu. NEUT2 mice offer a unique opportunity for the study of 1-C metabolism in a system where 1-C units as, 10-HCO-H4PteGlu, cannot be oxidized.
The specific aims are; 1) purification, characterization and cloning of mC1-THFS; 2) investigation of the promoter region of the cC1-THFS gene and 3), cloning and function of mFDH.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK049563-06
Application #
6524157
Study Section
Special Emphasis Panel (ZRG1-SSS-T (02))
Program Officer
May, Michael K
Project Start
1996-03-01
Project End
2004-07-31
Budget Start
2002-08-01
Budget End
2003-07-31
Support Year
6
Fiscal Year
2002
Total Cost
$226,500
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212