IDENTIFICATION OF THE TMV REPLICASE SEQUENCE THAT ACTIVATES THE N GENE-MEDIATED HYPERSENSITIVE RESPONSE

The N gene-mediated hypersensitive response (HR) in tobacco provides a high degree of resistance against most tobamoviruses by halting the p rogress of infection at the site of inoculation, A previous report ind icated a role for the 126/183-kDa replicase in induction of the HR in tobacco containing the N gene (H. S. Padgett and R. N. Beachy, Plant C ell 5:577-586, 1993). Chimeric virus genomes were constructed in which the genes encoding the 126/183-kDa proteins of the HR-eliciting patho gen, tobacco mosaic virus (TMV), and the resistance breaking tobamovir us, Ob, were exchanged, Inoculation of the chimeric viruses to leaves of Nicotiana tabacum cv. Xanthi NN confirmed that either the replicase protein of TMV or its mRNA was responsible for induction of HR. An ex pression vector based on the Ob virus was used to express fragments of various replicase genes. With this approach, it was determined that t he HR is caused by a portion of the replicase protein extending from a mino acid 692 to 1116. Consistent with this result, Ob mutants that in duce the HR on NN tobacco were found to carry mutations within the sam e portion of the replicase gene. The N gene-mediated HR is inactive at high temperatures, yet these mutants were able to overcome the HR at significantly lower temperatures than could TMV, indicating that the t emperature sensitivity of the N gene response is manifested at the lev el of interaction between the virus and the defense response mechanism .