GENETIC APPROACHES TO ELUCIDATE THE N GENE -TMV MEDIATED SIGNAL TRANSDUCTION PATHWAY IN TOMATO
C USTACHl, HH CHANG2, S WHITHAM3, L KNIGHTl, B BAKER2
lDepartment of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA; 2Plant Gene Expression Center, University of California and United States Department of Agriculture-Agriculture Research Service, Albany, CA 94710, USA; 3 Institute of Biological Chemistry, Washington State University, Pullman, WA, 99164-6340, USA.
Background and objectives
The N gene confers resistance to tobacco mosaic virus (TMV) in tobacco . It is hypothesized that N constitutes an early component of a signal transduction pathway which results in the hypersensitive response (HR), systemic acquired resistant (SAR), and pathogen inhibition. Little is known about how N participates in this signal transduction pathway. We are taking genetic approaches to elucidate the N-mediated signal transduction pathway involved in TMV resistance.
NwasisolatedfromtobaccousingthemaizetransposonAc[l]. Itisamember of the TIR-NBS-LRR class of R genes. A useful property of the N-mediated HR to TMV in tobacco is that it is reversibly inactivated at elevated temperatures. At temperatures of 28' and above, the N-mediated HR is suppressed and TMV moves systemically in the plant. HR is restored when the temperature is shifted to below 28' and necrosis occurs throughout the plant, resulting in massive cell death. In a mutational screen using the temperature shift assay, plants bearing a mutation in the N-mediated resistance response will survive the screen, while plants able to mount a normal resistance response will die. This temperature sensistive property was exploited to identify N loss-of function mutations in tobacco .
N confers resistance to TMV in N transgenic tomato and the temperature sensitive nature of N-mediated HR is conserved in NN tomato plants. Tomato is an excellent model system for genetic analysis because of its extensive genetic linkage map which includes morphological, biochemical and RFLP markers. Thus, the N homozygous transgenic tomato line was used in our mutational screen. We employed the temperature shift screen with EMS and fast neutron mutagenized homozygous N tomato plants to identify other loci that are possibly involved in N-mediated signal transduction.
Results and conclusions
Sixty-one EMS mutants have been isolated using the above described temperature shift screen. These 61 mutants fall into several phenotypic categories which possibly represent mutations in N-mediated signal transduction components or in plant host factors. These categories include: 1) asymptomatic for disease; 2) weak to severe mosaic symptoms; 3) weak resistance (HR with mosaic symptoms). We will present results of genetic and phenotypic characterization of these mutants.