| Programmed cell death (PCD) is a genetically and biochemically regulated pathway used to target specific cells for orderly destruction. While plants have been demonstrated to undergo PCD during development and in response to biotic and abiotic stresses, our knowledge of plant PCD regulation is far from complete. The objective of this study was to identify plant PCD-regulating genes.; First, knowledge of animal PCD was employed to predict potentially conserved aspects of plant PCD. Sequence searches identified candidate plant death domain-containing proteins, through which the human Fas receptor could potentially induce plant PCD. Several attempts to induce plant PCD via human Fas proved intractable: transformation of the full-length Fas into plants; transient overexpression of the Fas death domain; and inducible expression of the Fas death domain using a copper-inducible system, characterized herein. An identified candidate plant XAF1 homolog failed to interact with human XIAP by yeast-2-hybrid, but may nevertheless be a TRAF zinc finger regulator of plant PCD. Finally, a staining technique was developed that revealed that mitochondrial permeability transition is an early event in osmotic shock-induced tobacco PCD.; Second, a screen was devised to isolate tomato cDNAs whose overexpression antagonizes plant PCD. A transgenic root population collectively transformed with a tomato cDNA library designed for in planta overexpression was produced. After treatment with Fumonisin B1 (FB1), a PCD-inducing mycotoxin, protective cDNAs were recovered from surviving roots by PCR. The protective cDNAs included metallothionein, glutathione S-transferase and geranylgeranyl pyrophosphate synthetase, whose products have been previously demonstrated to protect against oxidative stress. Treatment with ascorbic acid, an ROS scavenger, inhibited FB1-induced PCD, suggesting that oxidative stress may play a role in the complex pathway of FB1-induced plant PCD. The screen also identified a PR1 family member, a cDNA whose predicted protein shares similarity with plant disease resistance genes, and four cDNAs whose predicted proteins share no sequence homology to functionally characterized proteins. Because expression of animal and animal virus anti-PCD genes in plants has been shown to confer broad-spectrum disease resistance, each of the identified cDNAs can now be tested as a potential source of disease resistance. |
