Engineering broad‐spectrum disease‐resistant rice by editing multiple susceptibility genes

Abstract

Rice blast and bacterial blight are important diseases of rice (Oryza sativa) caused by the fungus Magnaporthe oryzae and the bacterium Xanthomonas oryzae pv. oryzae (Xoo), respectively. Breeding rice varieties for broad‐spectrum resistance is considered the most effective and sustainable approach to controlling both diseases. Although dominant resistance genes have been extensively used in rice breeding and production, generating disease‐resistant varieties by altering susceptibility (S) genes that facilitate pathogen compatibility remains unexplored. Here, using CRISPR/Cas9 technology, we generated loss‐of‐function mutants of the S genes Pi21 and Bsr‐d1 and showed that they had increased resistance to M. oryzae. We also generated a knockout mutant of the S gene Xa5 that showed increased resistance to Xoo. Remarkably, a triple mutant of all three S genes had significantly enhanced resistance to both M. oryzae and Xoo. Moreover, the triple mutant was comparable to the wild type in regard to key agronomic traits, including plant height, effective panicle number per plant, grain number per panicle, seed setting rate, and thousand‐grain weight. These results demonstrate that the simultaneous editing of multiple S genes is a powerful strategy for generating new rice varieties with broad‐spectrum resistance.

Engineering of rice varieties with enhanced resistances to both blast and bacterial blight diseases via CRISPR/Cas9

“…More Xoo strains will be tested in these mutants to investigate their broad‐spectrum resistance to the bacterial blight. A recent study showed that CRISPR/Cas9‐mediated multiplex editing of the Pi21 , Bsr‐d1 as well as a well‐known Xoo ‐resistant gene Xa5 generated pi21‐bsr‐d1‐xa5 triple mutant which conferred resistant to both blast and bacterial blight (Tao et al ., 2021 ). In our study, on one hand, we further compared the blast resistance level among Bsr‐d1 , Pi21 and ERF922 single mutants and identified the erf922 mutant with the strongest blast resistance.…”

“…In our study, on one hand, we further compared the blast resistance level among Bsr‐d1 , Pi21 and ERF922 single mutants and identified the erf922 mutant with the strongest blast resistance. On the other hand, our study described the new roles of Pi21 as well as ERF922 in the resistance to Xoo , which could well explain why the pi21‐bsr‐d1‐xa5 triple mutant exhibited much stronger resistance to Xoo than xa5 single mutant in the previous study (Tao et al ., 2021 ).…”

“…So far, an atypical R gene Pi21 and two defense-related genes Bsr-d1 as well as ERF922 have attracted much more attention due to their broad-spectrum resistance. Bsr-d1 encodes a C 2 H 2 -type transcription factor that directly binds to the promoter of three peroxidase genes (Os01g73170, Os05g04470 and Os10g39170), thereby affecting H 2 O 2 accumulation and broad-spectrum resistance to M. oryzae (Li et al, 2017;Tao et al, 2021;Zhu et al, 2020). In addition, Bsr-d1 was shown to be transcriptionally regulated by MYSB1 through directly binding to its promoter and a single base change (SNP33-G) in the Bsr-d1 promoter leads to enhanced affinity with MYSB1, which suppresses Bsr-d1 gene expression.…”

“…Deletions of the proline-rich motifs cause a loss of function. Knockout of Pi21 confers non-race specific and durable blast resistance (Tao et al, 2021). ERF922 encodes an APETELA2/ ethylene response factor (AP2/ERF) type transcription factor and is strongly induced by M. oryzae (Liu et al, 2012).…”

Recent Progress in Rice Broad-Spectrum Disease Resistance

“…Editing the EBEs of S genes OsSWEET11 , OsSWEET13 , and OsSWEET14 in rice varieties japonica Kitaake and indica IR64 and Ciherang-sub1 resulted in loss of induction of these S genes by Xoo and broad-spectrum resistance against Xoo [ 123 , 124 , 125 , 126 ]. Similar strategies were used in the modification of S genes, Pi21 , Bsr-d1 , and Xa5 to obtain broad-spectrum resistance rice against Xoo and M. oryzae [ 127 ]. In addition, it was well summarized that editing the open reading frame of susceptibility defense regulators could obtain broad-spectrum resistance rice plants in the Wang’s review ( Figure 2 b) [ 128 ].…”