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ARS Home » Southeast Area » Raleigh, North Carolina » Plant Science Research » Research » Publications at this Location » Publication #302728

Research Project: Genetic Analysis of Complex Traits in Maize

Location: Plant Science Research

Title: Molecular and functional analyses of a maize autoactive NB-LRR protein identify precise structural requirements for activity

Author
item WANG, GUAN-FENG - North Carolina State University
item JI, JIABING - Purdue University
item JOHAL, GURI - Purdue University
item Balint-Kurti, Peter
item EL-KASMI, FARID - University Of North Carolina
item DANGL, JEFFERY - University Of North Carolina

Submitted to: PLoS Pathogens
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/1/2015
Publication Date: 2/26/2015
Citation: Wang, G., Ji, J., Johal, G., Balint Kurti, P.J., El-Kasmi, F., Dangl, J. 2015. Molecular and functional analyses of a maize autoactive NB-LRR protein identify precise structural requirements for activity. PLoS Pathogens. 11(2): e1004674. doi: 10.1371/journal.ppat.1004674.

Interpretive Summary: In this study we have analyzed a mutant maize gene with causes an autoimmune type response. We have identified several factors important for this response and from this can gain a better understanding of aspects of normal resistance gene function. We have identified precise structural requirements for activity in the protein. We have found a previously unidentified conserved region found in many similar resistance genes and have shown that it is important. Finally we have shown that the protein needs to be present in both the cytoplasm and nucleus of the cell and be able to move from one compartment to the other in order to confer the autoimmune response.

Technical Abstract: Plant disease resistance is often mediated by nucleotide binding-leucine rich repeat (NB-LRR or NLR) proteins, which trigger a hypersensitive response (HR), a rapid, localized cell death upon recognition of specific pathogens. The maize NLR-encoding Rp1-D21 gene is the result of an intergenic recombination event between two NLR genes, Rp1-D and Rp1-dp2 and confers an autoactive HR. We report systematic structural and functional analyses of Rp1-D21 in maize and N. benthamiana. A maize genetic screen identified 32 mutations that abolished Rp1-D21-induced autoactive HR. Transient expression of Rp1-D21 and its derivatives in N. benthamiana recapitulated the observed maize phenotypes. In this system, the N-terminal coiled-coil domain from either Rp1-D21 (CCD21) or Rp1-D was sufficient to induce HR, while the NB domain could suppress this phenotype. In Rp1-D21 but not in Rp1-D, the LRR domain relieved the NB-induced suppression. A systematic structural analysis through domain swaps between Rp1-dp2 and Rp1-D determined that the combination of amino acids (AAs) 370-651 from Rp1-dp2 with the C-terminal LRR region from Rp1-D (especially N1184 and the C-terminal 16 AAs) was associated with the autoactive HR. A novel motif conserved in NLRs was identified and named MHD2. Mutation of MHD2 in Rp1-dp2 resulted in autoactivity. Rp1-D21 was predominantly localized in cytoplasm with a small amount in the nucleus. Nucleocytoplasmic trafficking was required for Rp1-D21- and CCD21-triggered HR. This work reports several novel insights into structural and subcellular-localization requirements for NLR function and informs efforts towards utilizing these proteins for engineering disease resistance.