Molecular control of powdery mildew resistance in grapevine

Functional characterisation of NBS-LRR genes involved powdery mildew resistance in grapevine

Supervisors: Dr Ian Dry (08 8303 8632)
Location: Adelaide
Financial support: To be confirmed

Plants are protected from disease by specific recognition of diverse effectors presented by invading pathogens. This recognition capacity is encoded by plant resistance (R) genes that, in turn, activate a multitude of innate defense responses. The most prevalent class of plant R genes encode putative intracellular receptors containing highly conserved motifs including an N-terminal coiled coil (CC) or Toll/Interleukin-1 receptor-like (TIR) domain, a nucleotide binding site (NBS), and C-terminal, leucine-rich repeats (LRR). Typically, one member of a multigene family specifies resistance through direct or indirect recognition of a cognate pathogen avirulence (Avr) gene product. However, it has been shown that two functional members of a gene family can confer the same specificity or conversely, that one gene encodes recognition of multiple distinct pathogen signals In addition, genetic diversity within host-pathogen systems may also exist in the requirements for downstream components of disease response pathways.

Some details of the host components that regulate fundamental aspects of R protein triggered responses are now being uncovered. Resistance mediated by the barley Mla resistance protein is dependent on the presence of Rar1, Sgt1 and Hsp90 which appear to be crucial for the formation and activation of R protein-containing recognition complexes as well as for regulating downstream signalling processes. More recent results suggest that activation of the Mla protein, in the presence of its cognate fungal avirulence (Avr) protein, occurs via interaction of the N-terminal CC domain with a specific nuclear WRKY-box transcription factor, which regulates induction of the defense response. 

Powdery mildew (Erysiphe necator) is the most economically important grapevine disease worldwide. Using a map-based cloning approach, we have identified a major resistance locus (Run1) from a wild North American grape species that confers resistance to this obligate biotrophic pathogen.  Sequencing of this locus has revealed the presence of up to 6 full-length TIR-NBS-LRR type proteins that are very similar in structure to the Mla powdery mildew resistance genes isolated from barley.

Aims: This project aims to study the activation of each of the putative grapevine powdery mildew R gene candidates following powdery mildew infection to determine the relative specificities of each family member.  It will also investigate the regulation of signal transduction from the R gene candidates and identify host proteins which interact with and mediate the induction of the defense response. 

Experimental approach: Full-length transcripts will be isolated from a cDNA library constructed from powdery mildew infected grapevine leaves. Gene-specific primers will then be used in Real time-PCR experiments to study activation of individual R gene family members along with induction of Rar1 and Sgt1 genes in response to powdery mildew infection. Yeast 2 hybrid techniques will be employed to identify host factors which interact with the grapevine powdery mildew R proteins to mediate downstream signal transduction.

References:

• Barker CB et al. (2003) Use of molecular techniques for the transfer of powdery mildew resistance from a wild American grapevine into elite winegrape cultivars.  Australian & New Zealand Grapegrower & Winemaker 473: 97-99.
• Halterman DA et al. (2003) Powdery mildew-induced Mla mRNAs are alternatively spliced and contain multiple upstream open reading frames. Plant Physiology 131: 558-67.
• Shirasu K and Schulze-Lefert P (2003) Complex formation, promiscuity and multi-functionality: protein interactions in disease-resistance pathways Trends in Plant Science 8: 252-258.
• McHale L et al. (2006) Plant NBS-LRR proteins: adaptable guards. Genome Biology 7: 212.