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1	PLANT-ENDOPHYTE INTERPLAY PROTECTS TOMATO AGAINST A VIRULENT VERTICILLIUM DAHLIAE Shittu, Hakeem Olalekan 05 October 2010 (has links) When tomato Craigella is infected with Verticillium dahliae Dvd-E6 (Dvd-E6), a tolerant state is induced with substantial pathogen load, but few symptoms. Unexpectedly, these plants are more robust and taller with Dvd-E6 behaving as an endophyte. Some endophytes can protect plants from virulent pathogens. This research was undertaken to improve understanding of the cellular and molecular nature of Verticillium tolerance in tomato, especially whether infection by Dvd-E6 can protect Craigella from virulent V. dahliae, race 1 (Vd1). To permit mixed infection experiments a restriction fragment length polymorphism (RFLP)-based assay was developed and used for differentiating Dvd-E6 from Vd1, when present in mixed infections. The results suggested that protection involves molecular interplay between Dvd-E6 and Vd1 in susceptible Craigella (CS) tomatoes, resulting in restricted Vd1 colonization. Further studies showed a dramatic reduction of Vd1 spores and mycelia. To examine genetic changes that account for these biological changes, a customized DNA chip (TVR) was used to analyze defense gene mRNA levels. The defense gene response was categorized into four groups. Group 1 was characterized by strong induction of defense genes followed by suppression. However, Vd1-induced gene suppression was blocked by Dvd-E6 in mixed infections. These genes included some transcription factors and PR proteins such as class IV chitinases and beta glucanases which are known to target fungal spores and mycelia. Experiments also were repeated with a Craigella resistant (CR) isoline containing a fully active Ve locus (Ve1+ and Ve2+). The biological results showed that the presence of the Ve1+ allele resulted in restricted Vd1 colonization and, in a mixed infection with Dvd-E6, Vd1 was completely eliminated from the plant stem. Surprisingly, there was no significant increase in defense gene mRNAs. Rather, elevated basal levels of defense gene products appeared sufficient to combat pathogen attack. To investigate functional effects of the genetic changes observed, an inducible RNAi knockdown vector for a defense gene (TUS15G8) with unknown function (pMW4-TUS15G8) as well as the Ve2 resistance gene (pMW-Ve2) was prepared as a initial step for future transformation analyses. Taken together the results reveal intriguing but complex biological and molecular changes in mixed infections, which remain a basis for future experiments and potential agricultural benefits. / Canadian Commonwealth Scholarship and Fellowship Plan Tomato-Verticillium response Pathogenesis related (PR) protein Resistant relationship Susceptible relationship Endophyte-induced protection Tolerant relationship Compatible interaction Incompatible interaction Defence protein Mycelia Fungal spores Mixed infection Agrobacterium Functional analysis RNAi construct pMW-Ve2 pMW4-TUS15G8 Gene expression Silencing Ve gene Susceptibility Resistance Defence genes plant-pathogen interaction RFLP Verticillium dahliae race 1 Dvd-E6 Craigella RT-PCR Tolerance Endophyte Plant transformation RNAi Microarray Tomato Verticillium

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PLANT-ENDOPHYTE INTERPLAY PROTECTS TOMATO AGAINST A VIRULENT VERTICILLIUM DAHLIAE