Chitin is a major component of fungal cell wall and is synthesized by chitin synthases (Chs). Plant pathogenic fungi normally have multiple chitin synthase genes. To determine their roles in development and pathogenesis, we functionally characterized all seven CHS genes in Magnaporthe oryzae. Three of them, CHS1, CHS6, and CHS7, were found to be important for plant infection. While the chs6 mutant was non-pathogenic, the chs1 and chs7 mutants were significantly reduced in virulence. CHS1 plays a specific role in conidiogenesis, an essential step for natural infection cycle. Most of chs1 conidia had no septum and spore tip mucilage. The chs6 mutant was reduced in hyphal growth and conidiation. It failed to penetrate and grow invasively in plant cells. The two MMD-containing chitin synthase genes, CHS5 and CHS6, have a similar expression pattern. Although deletion of CHS5 had no detectable phenotype, the chs5 chs6 double mutant had more severe defects than the chs6 mutant, indicating that they may have overlapping functions in maintaining polarized growth in vegetative and invasive hyphae. Unlike the other CHS genes, CHS7 has a unique function in appressorium formation. Although it was blocked in appressorium formation by germ tubes on artificial hydrophobic surfaces, the chs7 mutant still produced melanized appressoria by hyphal tips or on plant surfaces, indicating that chitin synthase genes have distinct impacts on appressorium formation by hyphal tip and germ tube. The chs7 mutant also was defective in appressorium penetration and invasive growth. Overall, our results indicate that individual CHS genes play diverse roles in hyphal growth, conidiogenesis, appressorium development, and pathogenesis in M. oryzae, and provided potential new leads in the control of this devastating pathogen by targeting specific chitin synthases. Chitin is one of the major components of cell wall that plays vital roles in hyphal tip growth and fungal morphogenesis. Biosynthesis of chitin is catalyzed by chitin synthases, a well-known fungicide target. However, systematic characterization of chitin synthase genes has not yet been reported in plant pathogenic ascomycetes. To determine their roles in development and pathogenesis, in this study we functionally characterized all the seven chitin synthase genes (CHS1-CHS7) in Magnaporthe oryzae, a model for studying fungal development and pathogenesis. While CHS2, CHS3, CHS4, and CHS5 are dispensable for plant infection, CHS6 is essential for pathogenesis. The chs6 mutant failed to penetrate plant cells and develop infectious hyphae. Two other chitin synthase genes, CHS1 and CHS7, also are important for virulence. The chs1 and chs7 mutants caused only rare lesions on rice seedlings. Other than plant infection, CHS1 and CHS7 also play specific roles during conidiogenesis and appressorium formation, respectively. Interestingly, the chs7 mutant was blocked in appressorium formation by germ tubes on artificial hydrophobic surfaces but was normal by hyphal tips and on plant surfaces. Different chitin synthase genes may be involved during appressorium formation by different fungal tissues or on different surfaces in M. oryzae.

Summary: Lesion mimic mutants are used to elucidate mechanisms controlling plant responses to pathogen attacks and environmental stresses. Although dozens of genes had been functionally demonstrated to be involved in lesion mimic phenotype in several plant species, the molecular mechanisms underlying the hypersensitive response are largely unknown. Here, a rice (Oryza sativa) lesion mimic mutant natural blight leaf 3 (nbl3) was identified from T‐DNA insertion lines. The causative gene, OsNBL3, encodes a mitochondrion‐localized pentatricopeptide repeat (PPR) protein. The nbl3 mutant exhibited spontaneous cell death response and H₂O₂ accumulation, and displayed enhanced resistance to the fungal and bacterial pathogens Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae. This resistance was consistent with the up‐regulation of several defence‐related genes; thus, defence responses were induced in nbl3. RNA interference lines of OsNBL3 exhibited enhanced disease resistance similar to that of nbl3, while the disease resistance in overexpression lines did not differ from that of the wild type. In addition, nbl3 displayed improved tolerance to salt, accompanied by up‐regulation of several salt‐associated marker genes. OsNBL3 was found to mainly participate in the splicing of mitochondrial gene nad5 intron 4. Disruption of OsNBL3 leads to the reduction in complex I activity, the elevation of alternative respiratory pathways and the destruction of mitochondrial morphology. Overall, the results demonstrated that the PPR protein‐coding gene OsNBL3 is essential for mitochondrial development and functions, and its disruption causes the lesion mimic phenotype and enhances disease resistance and tolerance to salt in rice.

Endoplasmic reticulum (ER) homeostasis is essential for plants to manage responses under environmental stress. Plant immune activation requires the ER, but how ER homeostasis is associated with plant immune activation is largely unexplored. Here we find that transcription of an HVA22 family gene, OsHLP1 (HVA22-like protein 1), is induced by Magnaporthe oryzae infection. Overexpression of OsHLP1 significantly enhances blast disease resistance but impairs ER morphology in rice (Oryza sativa), resulting in enhanced sensitivity to ER stress. OsHLP1 interacts with the NAC (NAM, ATAF, and CUC) transcription factor OsNTL6 at the ER. OsNTL6 localizes to the ER and is relocated to the nucleus after cleavage of the transmembrane domain. OsHLP1 suppresses OsNTL6 protein accumulation, whereas OsNTL6 counteracts OsHLP1 by alleviating sensitivity to ER stress and decreasing disease resistance in OsHLP1 overexpression plants. These findings unravel a mechanism whereby OsHLP1 promotes disease resistance by compromising ER homeostasis when plants are infected by pathogens.

Summary: Brassinosteroids (BRs) play a crucial role in shaping the architecture of rice (Oryza sativa) plants. However, the regulatory mechanism of BR signalling in rice immunity remains largely unexplored. Here we identify a rice mutant dla, which exhibits decreased leaf angles and is insensitive to 24‐epiBL (a highly active synthetic BR), resembling the BR‐deficient phenotype. The dla mutation caused by a T‐DNA insertion in the OsDLA gene leads to downregulation of the causative gene. The OsDLA knockout plants display reduced leaf angles and less sensitivity to 24‐epiBL. In addition, both dla mutant and OsDLA knockout plants are more susceptible to rice blast compared to the wild type. OsDLA is a GRAS transcription factor and interacts with the BR signalling core negative regulator, GSK2. GSK2 phosphorylates OsDLA for degradation via the 26S proteasome. The GSK2 RNAi line exhibits enhanced rice blast resistance, while the overexpression lines thereof show susceptibility to rice blast. Furthermore, we show that OsDLA interacts with and stabilizes the WRKY transcription factor OsWRKY53, which has been demonstrated to positively regulate BR signalling and blast resistance. OsWRKY53 directly binds the promoter of PBZ1 and activates its expression, and this activation can be enhanced by OsDLA. Together, our findings unravel a novel mechanism whereby the GSK2‐OsDLA‐OsWRKY53 module coordinates blast resistance and plant architecture via BR signalling in rice.

The regulation of leaf senescence and disease resistance plays a crucial role in determining rice grain yield and quality, which are important to meet the ever-increasing food demands of the world. Here, we identified an atypical Dof transcriptional factor OsDes1 that contributes to the stay-green phenotype, grain yield, and disease resistance in rice. The expression level of OsDes1 is positively associated with stay-green in natural variations of japonica rice, suggesting that OsDes1 would be alternatively used in breeding programs. Mechanistically, OsDes1 targets the promoter of the Rieske FeS protein gene OsPetC to activate its expression and interacts with OsPetC to protect against its degradation, thus promoting stay-green and ultimately improving the grain yield. OsDes1 also binds to the promoter region of defense-related genes, such as OsPR1b , and activates their expression, leading to enhanced disease resistance. These findings offer a potential strategy for breeding rice to enhance grain yield and disease resistance.