OsMT1a and OsMT1b positively regulate rice resistance to M. oryzae
The above RNA-seq analysis suggested that 12 hpi is a critical time point for rice defense against M. oryzae. Among 4680 fungal upregulated DEGs at 12 hpi in Nipponbare, the type 1 metallothionein (OsMT1a , Os11g0704500 ) gene had the highest expression value (FPKM). OsMT1a was also one of the top four genes in terms of the FPKM value among 6808 upregulated DEGs at 12 hpi in Hui1586 (Supplemental dataset 1). In rice, OsMT1a has a very close protein homolog (with 84% identity and 89% positivity in Blastp) encoded by Os03g0288000 (hereafter named OsMT1b ), which was also upregulated by M. oryzae in Nipponbare and Hui1586 (Supplemental Dataset 1). To assess the involvement of OsMT1a andOsMT1b in rice blast resistance, we generated osmt1a osmt1b (osmt1a/b for short) double knockout lines in the Nipponbare background using CRISPR/Cas9 technology. We acquired theosmt1a/b-1 and osmt1a/b-2 double mutant, which contains a one-base deletion in OsMT1a and a five-base deletion or one-base deletion in OsMT1b (Fig. S3B).
We then inoculated osmt1a/b mutant and Nipponbare plants with the blast isolate Guy11. We found that the osmt1a/b plants were more susceptible to rice blast compared with Nipponbare (Figs. 9A and 9B). Consistent with the enhanced blast susceptibility in osmt1a/b,the expression of OsPR5 and OsPAL were reduced inosmt1a/b compared with Nipponbare plants after M. oryzaeinfection (Fig. 9C). Thus, we concluded that OsMT1a andOsMT1b positively contributed to rice resistance to blast disease.