Eric Wauson’s Thomas Collum Butler Fellowship Statement:
Department of Pharmacology
Application
Thomas Collum Butler Fellowship
(Deadline: November 23, 2005)
Name: Eric M. Wauson
Mentor: Lee M. Graves
Graduate Program/Entering Date: Pharmacology/2001
II. Brief State of Research Interests (1 page maximum):
My research interests are in determining how signal transduction pathways modulate CAD, a key metabolic enzyme in pyrimidine synthesis. Pyrimidine synthesis is necessary for cellular proliferation, as pyrimidines are used in the formation DNA, RNA, phospholipids, and pyrimidine sugars. CAD catalyzes the first rate-limiting step in the de novo pyrimidine synthesis pathway. The expression and activity of CAD are highly correlated with cell proliferation and the normal function of this enzyme is reported to be necessary for the early embryological development of specific tissues in zebrafish and drosophila. CAD activity is regulated by multiple mechanisms including allosteric modulation and phosphorylation. The goals of my research are to determine how CAD phosphorylation and protein-protein interactions regulate CAD activity, and how this affects pyrimidine metabolism at the cellular and organismal level. Ultimately, I want to determine how the function of CAD affects cell growth and mammalian development.
To investigate CAD phosphorylation, I have used 32P metabolic labeling, phospho-peptide mapping, and mass spectrometry to identify novel phosphorylated amino acids in CAD from intact cells. I have mutated specific residues to determine how phosphorylation at these sites affects enzyme activity and cellular growth. Preliminary results suggest the phosphorylation of a specific CAD residue may be necessary for cellular proliferation.
Genome wide protein-protein interaction studies suggest that CAD forms interactions with other proteins. I am interested in determining if protein-protein interactions with CAD affect its enzymatic activity and pyrimidine synthesis. In collaboration with Aziz Sancar and Laua Lindsy-Boltz (UNC), we discovered that a domain in CAD bound to a specific region the human checkpoint protein Rad9. I determined that there was an increase in CAD activity when in complex with Rad9, suggesting that this protein-protein interaction serves a physiological function. This work was published and we are now investigating whether the Rad9-induced increase in CAD activity influences the flux through the de novo pyrimidine synthesis pathway.
Future goals of my research are to investigate the role of CAD and pyrimidine metabolism during development by creating a mouse that contains a CAD gene deletion. Since de novo pyrimidine synthesis is critical for lymphocyte expansion, and CAD is important in normal development of drosophila and zebrafish, a mouse model is critical to determining the role of CAD and pyrimidine synthesis in mammalian development.
III. State How Your Work Relates to Developmental Disabilities (1 paragraph):
There are numerous genetic disorders of pyrimidine metabolism that result in developmental disabilities in humans. Disruption of normal pyrimidine homeostasis causes neurological abnormalities, anaemia, and immunodeficiency in humans. Since CAD is the first rate-limiting step in de novo pyrimidine biosynthesis, the mechanisms by which CAD is regulated are critical to understanding organismal development. This hypothesis is supported by data from model organisms where mutations in CAD give rise to severe developmental defects. A defect in drosphila called rudimentary,which causes wing malformation and female sterility, is known to be caused by a mutation in CAD. Additionally, a defect in zebra fish called perplexed, also caused by a mutation in CAD, is characterized by reduced eye size, abnormal retina morphogenesis, and disrupted development of the fin and jaw structures. Though CAD function has been studied in invertebrates and lower vertebrates, the role of CAD in mammalian development has not been investigated. Therefore our mouse CAD deletion project will provide valuable insights into the developmental role of CAD.
IV. List Publications, Abstracts, Awards:
PUBLICATIONS
*Lindsey-Boltz LA, *Wauson EM, Graves LM, Sancar A. (2004) The human Rad9 checkpoint protein stimulates the carbamoyl phosphate synthetase activity of the multifunctional protein CAD. Nucleic Acids Res. 32(15): 4524-30. *Co-first authors.
Wauson EM, Langan AS, Vorce RL. (2002) Sodium arsenite inhibits and reverses expression of adipogenic and fat cell-specific genes during in vitro adipogenesis. Toxicological Sciences 65(2): 211-219.
Ruan Y, Peterson MH, Wauson EM, Waes JG, Finnell RH, Vorce RL. (2000) Folic acid protects SWV/Fnn embryo fibroblasts against arsenic toxicity. Toxicological Letters 117(3): 129-37.
Trouba KJ, Wauson EM, Vorce RL. (2000) Sodium arsenite inhibits terminal differentiation of murine C3H-10T1/2 preadipocytes. Toxicol. Appl. Pharmacol. 169(1): 25-35.
Trouba KJ, Wauson EM, Vorce RL. (2000) Sodium arsenite-induced dysregulation of proteins involved in proliferative signaling. Toxicol. Appl. Pharmacol. 164(2): 161-70.