Modulation of cellular signaling and genetic regulation by imidopeptides, amino acids and their metabolites were studied at the level of: a) Imidodipeptides and prolidase, b) Pyrroline 5-carboxylate (P5C) as a signaling and regulatory molecule and c) Enzymes mediating the formation and degradation of P5C. a. Imidodipeptides and prolidase. Imidodipeptides containing proline or hydroxyproline originate from either tissue matrix degradation or from dietary protein. They are not substrate for generic peptidases but instead circulate to tissues where they are hydrolyzed by prolidase. Our studies showed that the level of cellular prolidase is regulated by extracellular collagen acting through integrin receptors. Thus, the hydrolysis of imidodipetides, degradative products of matrix collagen and dietary proteins, is responsive to cellular interaction with extracellular matrix. In addition, imidodipeptides bind to certain members of the family of Peptidyl prolyl cis-trans isomerases (Rotamases) which have a wide spectrum of function from immunoregulation to apoptosis and cell cycle control. Interaction between imidodipeptides and rotamases are being studied. b. P5C as a signaling and regulatory molecule. P5C stimulates PRPP and purine ribonucleotide synthesis synergistically with certain growth factors and increases the entry of cells into S-phase. Recent studies suggest that P5C is synergistic with hypoxia in inducing expression of the gene for Nitric oxide synthase II (NOSII). Since NO can serve as an ubiquitous signaling molecule, some of the previously observed P5C effects may be mediated by NO. Preliminary results suggest that inhibitors of rotamase activity can block the P5C effect. c. Enzymes mediating the synthesis and degradation of P5C Because P5C has the aforementioned regulatory effects, the synthesis and degradation of P5C is of interest. Specifically, ornithine aminotransferase, P5C reductrase and P5C dehydrogenase are being studied. Using Northern blots, we are studying the expression of these enzymes in animals and in cultured cells. We found that the regulation of hepatic ornithine aminotrasferase by diet requires the presence of p53. In transgenic p53 knockout mice, regulation by dietary carbohydrate is attenuated.
