32001:

Activation tagging approach for identification of genes involved in plant disease resistance responses.

Authors:	Xia, Yiji(A)Borevitz, Justin(B)Guo, Zejian(B)Blount, Jack(A)Dixon, Richard(A)Lamb, Chris(B)
Affiliations:	(A): The Noble Foundation (B): The Salk Institute (C): University of Edinburgh
Presenter:	Xia, Yiji , yxia@noble.org

A gain-of-function mutant screen was employed to uncover genes involved in plant disease resistance pathways. We created a mutant population by transforming Arabidopsis with an agrobacterium harboring the activation tagging binary vector pSKI15. The pSKI15 has four copies of 35S enhancers next to the T-DNA right border. Insertion of the 35 enhancers into the plant genome is expected to result in ectopic over-expression of adjacent plant genes, causing gain-of-function mutations. We have screened approximately 5,000 T-DNA activation tagged lines for dominant mutants that exhibit either enhanced resistance or enhanced susceptibility to infection by virulent Pseudomonas syringe pv. tomato (Pst) or P. syringae pv. maculicola (Psm). We have identified, among others, cdr1-D (constitutive disease resistance) and asr1-D (activation-tagged suppresser of disease resistance) mutants. The cdr1-D mutant is resistant to both Pst and Psm. In the cdr1-D mutant, defense responses are constitutively activated. Firstly, the mutant plants constitutively express high level of defense-related genes, including the PR genes and RbohA that encodes a subunit of the oxidative burst NADPH oxidase. Secondly, the mutant accumulates high levels of salicylic acid (SA). In addition, the cdr1-D mutant develops spontaneous micro-bursts of H2O2 and microscopic lesions. Analysis of cdr1-D NahG plants revealed that SA is a necessary component in the cdr1-D-mediated induction of defense responses. The CDR1 gene has been cloned. The deduced CDR1 protein shares sequence homology with aspartic proteases. Using the transgenic plants harboring a construct in which the expression of CDR1 gene is under the control of an inducible promoter, it was found that transient induction of the CDR1 makes these plants highly resistant to subsequent pathogen infection. CDR1 protein has been found to be extracellular. Interestingly, the intercellular fluid isolated from the CDR1-overexpressing transgenic plants can strongly induce PR gene expression. We propose that the CDR1 protein might be involved in the generation of a polypeptide signal (elicitor) which could be a mobile SAR signal. The asr1-D mutant was originally identified by its enhanced susceptibility to the virulent Pst. Further studies revealed that the gene-for-gene disease resistance is also compromised in the mutant. Cloning of the ASR1 gene should broaden our understanding on regulation of disease resistance pathways. .