Large-scale identification and functional analysis of NLR genes in blast resistance in the Tetep rice genome sequence

Abstract

Tetep is a rice cultivar known for broad-spectrum resistance to blast, a devastating fungal disease. The molecular basis for its broad-spectrum resistance is still poorly understood. Is it because Tetep has many moreNLRgenes than other cultivars? Or does Tetep possess multiple majorNLRgenes that can individually confer broad-spectrum resistance to blast? Moreover, are there many interactingNLRpairs in the Tetep genome? We sequenced its genome, obtained a high-quality assembly, and annotated 455 nucleotide-binding site leucine-rich repeat (NLR) genes. We cloned and tested 219NLRgenes as transgenes in 2 susceptible cultivars using 5 to 12 diversified pathogen strains; in many cases, fewer than 12 strains were successfully cultured for testing. Ninety clonedNLRs showed resistance to 1 or more pathogen strains and each strain was recognized by multipleNLRs. However, fewNLRs showed resistance to >6 strains, so multipleNLRs are apparently required for Tetep’s broad-spectrum resistance to blast. This was further supported by the pedigree analyses, which suggested a correlation between resistance and the number of Tetep-derivedNLRs. In developing a method to identifyNLRpairs each of which functions as a unit, we found that >20% of theNLRs in the Tetep and 3 other rice genomes are paired. Finally, we designed an extensive set of molecular markers for rapidly introducing clustered and pairedNLRs in the Tetep genome for breeding new resistant cultivars. This study increased our understanding of the genetic basis of broad-spectrum blast resistance in rice.

Fitness benefit plays a vital role in the retention of thePi-tasusceptible alleles

“…Expression levels can alter the penetrance of phenotypes (Raj, et al 2010), and alternation of the expression of Pi-ta can lead to non-specific activation of the hypersensitive response (HR) or even lethality, if expression levels are too high (Wang, et al 2019). To elucidate the molecular components involved in fitness decline in Pi-ta knockout mutants, we first evaluated the expression of the Pi-ta gene by qRT-PCR in the Pi-ta − (X1, X2, X3, X4 and X6), Pi-ta WT (X7), and Pi-ta 0 (X0) lines in the absence of relevant pathogens (Figure 3A).…”

A molecular roadmap to the plant immune system

“…Many different types of proteins have been found as IDs in plant NLRs, and likely function in direct perception of effectors ( 193 – 197 ). The integrated HMA domains of the sensor NLRs of the rice Pik and Pia pairs are exemplars of IDs and serve as model systems for understanding the principles of effector perception by these domains ( 11 , 29 , 31 , 178 ).…”

Applications of CRISPR/Cas to Improve Crop Disease Resistance: Beyond Inactivation of Susceptibility Factors

“…However, R-genes such as nucleotide-binding site leucine-rich repeat (NLR) genes often function as pairs consisting of a helper NLR (activating the defense signaling) and a sensor NLR recognizing the pathogen effector and controlling the helper to prevent autoimmunity in the absence of the pathogen (Wu et al, 2017). Transferring R-genes to crops and expressing them in response to biotic stress could improve pathogen resistance, but the number of suitable R-genes is limited and may first require the identification of interacting helper NLRs (Wang et al, 2019). Furthermore, dominant R-gene-mediated resistance is less durable than recessive resistance and is usually restricted to a few race-specific isolates, which can quickly overcome this resistance by their higher mutation rate (Dangl et al, 2013;Pandolfi et al, 2017;Kourelis and van der Hoorn, 2018).…”