Molecular Mechanism Study Of Tomato Immune Receptor Protein Sw-5b Confers Broad Spectrum Resistance To Tospovirusand Genetic Diversity Analysis Of Tsw ResistanceBreaking Isolates Of TSWV In China

Tospovirus contains many serious quarantine virus,Tomato spotted wilt vruS(TSWV)is the representative species,distributes all over the world and mainly concentrates in the southwest area in China.The breeding of resistant varieties is an effective way to combat the disease.At present,there are only two kinds of resistance genes Sw-5b and TSwfor TSWV.1.Molecular mechanism study of tomato Sw-5b NLR confers broad-spectrum resistance to TospovirusesPlant and animal intracellular nucleotide-binding leucine-rich repeat(nucleotide-binding oligomerization domain-like receptors,NLRs)receptors play crucial roles in detecting and preventing infection from various pathogens.Unlikeanimal NLRs typically provide the resistance by recognizing conserved pathogen-associated molecular patterns(pathogen-associated molecular patterns,PAMPs),plant NLRs confer the resistance by recognizing pathogen race-specific effector.Here,we demonstrate that tomato Sw-5b NLR confers broad-spectrum resistance against diverse American type tospoviruses by recognizing a conserved 21-amino-acid peptide within the viral movement protein NSm(NSm21).Full-length Sw-5b and NB-ARC-LRR domain directly associate with NSm21 in vitro and in planta.Site-directed mutagenesis and structure modeling analyses identified that Sw-5b LRR R927 site is an important site for interaction with NB-ARC,and mutation at the R927 site can convert Sw-5b from an inhibitory state to an active state.Thus,recognition of NSm21 by Sw-5b likely weakens the intramolecular interaction between LRR and NB-ARC domains,resulting in the activation of Sw-5b.Results described here provide the first example of plant NLR that recognize a small conserved pathogen effector-derived peptide to conferbroad-spectrum resistance.2.Genetic diversity analysis of Tsw resistance breaking isolates of TSWV in ChinaWidely-used resistant peppers(Capsicum spp.)bearing the Tsw locus triggered the rapid emergence of resistance breaking(RB)isolates ofTSWV around the world.However,no RB isolates have been reported so far in China.TSWV-induced diseases rapidly increased in Yunnan,Southwest China,in recent years.No information is available about the diversity of TSWV isolates in Yunnan.In this study,we report the occurrence of natural TSWVRB variants among isolates collected in Yunnan.We initially collected a TSWV isolate from asparagus lettuce(TSWV-LE)in Yunnan in 2012.Surprisingly,this isolate of TSWV induced systemic necrosis on pepper carrying Tsw-resistance gene.We then collected novel TSWV isolates in 2015 and found that a tomato-isolate(TSWV-YN18)and a tobacco-isolate(TSWV-YN53)also overcome the Tsw-mediated resistance.TSWV-YN18 induced systemic ringspots,whereas YN53 caused systemic chlorotic mottle.Variations in the TSWV NSs protein are the key determinants associated with Tsw resistance-breaking isolates.We found that TSWV-LE NSs retains the hypersensitive response(HR)induction,whereas TSWV-YN18 and-YN53 NSs are unable to induce HR.However,the NSs of all three RB isolates suppress RNA silencing.Sequence analysis of NSs revealed that RB isolates of Yunnan have no amino acid mutation sites common to other previously reported RB isolates.However two amino acids(F74 and K272)on LE-NSs make it distinct from YN18 and YN53 RB isolates.The occurrence of different RB isolates and the failure of Tsw-mediated resistance control pose serious threats to domestic pepper crops in Southwest China.