The objective of this proposal is to determine the mechanism by which the newly identified intracellular signaling molecules, the retro-retinoids, regulate cell growth and cell death. The retro-retinoid 14-hydroxy-retro-retinol (14-HRR) is induced in activated lymphocytes and supports their proliferation. A second retro-retinoid, anhydroretinol (AR), a retinol metabolite found in liver and lung, is a competitive inhibitor of 14-HRR, and prevents activation of T lymphocytes and proliferation of activated B and T lymphocytes. Dr. Buck has recently determined that the intracellular retro-retinoid pathway is not restricted to lymphoid cells. In general, AR has been shown to block activation and cause cell death in retinol-dependent cell lines, i.e., NIH 3T3 cells, and both effects can be prevented by 14-HRR in a dose-dependent manner. AR-induced cell death cannot be blocked by transcriptional or translational inhibitors but can be prevented by the protein tyrosine kinase inhibitor, herbimycin A, implying the 14-HRR/AR signaling pathway involves the regulation of one or more tyrosine kinase(s). Drs. Li and Cohen at Stanford University developed the method of Random Homozygous KnockOut in Mammalian Cells to identify previously unknown genes encoding proteins involves in intracellular signaling pathways. Using this methodology, the applicant isolated in a preliminary screen two not yet characterized AR-resistant clones whose selected gene products involved in anhydroretinol-induced cell death using Random Homozygous KnockOut. The genes and their corresponding proteins will be characterized by genetic, biochemical and immunological methods.
| Pakhomova, S; Luz, J G; Kobayashi, M et al. (2000) Crystallization of retinol dehydratase from Spodoptera frugiperda: improvement of crystal quality by modification by ethylmercurythiosalicylate. Acta Crystallogr D Biol Crystallogr 56:1641-3 |
| Chen, Y; Buck, J (2000) Cannabinoids protect cells from oxidative cell death: a receptor-independent mechanism. J Pharmacol Exp Ther 293:807-12 |
| Sinclair, M L; Wang, X Y; Mattia, M et al. (2000) Specific expression of soluble adenylyl cyclase in male germ cells. Mol Reprod Dev 56:11-Jun |
| Buck, J; Sinclair, M L; Schapal, L et al. (1999) Cytosolic adenylyl cyclase defines a unique signaling molecule in mammals. Proc Natl Acad Sci U S A 96:79-84 |
| Chen, Y; Buck, J; Derguini, F (1999) Anhydroretinol induces oxidative stress and cell death. Cancer Res 59:3985-90 |
