Summary: Abscisic acid (ABA) and jasmonic acid (JA) both inhibit seed germination, but their interactions during this process remain elusive. Here, we report the identification of a ‘SAPK10‐bZIP72‐ AOC ’ pathway, through which ABA promotes JA biosynthesis to synergistically inhibit rice seed germination. Using biochemical interaction and phosphorylation assays, we show that SAPK10 exhibits autophosphorylation activity on the 177 ^th serine, which enables it to phosphorylate bZIP72 majorly on 71 ^st serine. The SAPK10‐dependent phosphorylation enhances bZIP72 protein stability as well as the DNA‐binding ability to the G‐box cis ‐element of AOC promoter, thereby elevating the AOC transcription and the endogenous concentration of JA. Blocking of JA biosynthesis significantly alleviated the ABA sensitivity on seed germination, suggesting that ABA‐imposed inhibition partially relied on the elevated concentration of JA. Our findings shed a novel insight into the molecular networks of ABA–JA synergistic interaction during rice seed germination.

Jumonji C (JmjC) domain-containing proteins are a group of functionally conserved histone lysine demethylases in Eukaryotes. Growing evidences have shown that JmjCs epigenetically regulate various biological processes in plants. However, their roles in plant biotic stress, especially in rice bacterial blight resistance have been barely studied so far. In this study, we found that the global di- and tri-methylation levels on multiple lysine sites of histone three were dramatically altered after being infected by bacterial blight pathogen Xanthomonas oryzae pv. oryzae (Xoo). Xoo infection induced the transcription of 15 JmjCs, suggesting these JmjCs are involved in rice bacterial blight defense. Further functional characterization of JmjC mutants revealed that JMJ704 is a positive regulator of rice bacterial blight resistance as the jmj704 became more susceptible to Xoo than the wild-type. In jmj704, the H3K4me2/3 levels were significantly increased; suggesting JMJ704 may be involved in H3K4me2/3 demethylation. Moreover, JMJ704 suppressed the transcription of the rice defense negative regulator genes, such as NRR, OsWRKY62 and Os-11N3, by reducing the activation marks H3K4me2/3 on them. JMJ704 may be a universal switch controlling multiple genes of the bacterial blight resistance pathway. JMJ704 positively regulates rice defense by epigenetically suppressing master negative defense regulators, presenting a novel mechanism distinct from its homolog JMJ705 which also positively regulates rice defense but via activating positive defense regulators.

Rice is a major crop worldwide. Bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) has become one of the most devastating diseases for rice. It has been clear that phosphorylation plays essential roles in plant disease resistance. However, the role of phosphorylation is poorly understood in rice-Xoo system. Here, we report the first study on large scale enrichment of phosphopeptides and identification of phosphosites in rice before and 24 h after Xoo infection. We have successfully identified 2367 and 2223 phosphosites on 1334 and 1297 representative proteins in 0 h and 24 h after Xoo infection, respectively. A total of 762 differentially phosphorylated proteins, including transcription factors, kinases, epi-genetic controlling factors and many well-known disease resistant proteins, are identified after Xoo infection suggesting that they may be functionally relevant to Xoo resistance. In particular, we found that phosphorylation/dephosphorylation might be a key switch turning on/off many epi-genetic controlling factors, including HDT701, in response to Xoo infection, suggesting that phosphorylation switch overriding the epi-genetic regulation may be a very universal model in the plant disease resistance pathway. The phosphosites identified in this study would be a big complementation to our current knowledge in the phosphorylation status and sites of rice proteins. This research represents a substantial advance in understanding the rice phosphoproteome as well as the mechanism of rice bacterial blight resistance.

In autumn 2020, leaf blight was observed on rice (Oryza sativa L., variety Zhongzao39, Yongyou9, Yongyou12, Yongyou15, Yongyou18, Yongyou1540, Zhongzheyou8, Jiafengyou2, Xiangliangyou900 and Jiyou351) in the fields of 17 towns in Zhejiang and Jiangxi Provinces, China. The disease incidence was 45%-60%. Initially, water-soaked, linear, light brown lesions emerged in the upper blades of the leaves, and then spread down to leaf margins, which ultimately caused leaf curling and blight during the booting-harvest stage (Fig. S1). The disease symptoms were assumed to be caused by Xanthomonas oryzae pv. oryzae (Xoo), the pathogen of rice bacterial blight. 63 isolates were obtained from the collected diseased leaves as previously described (Hou et al. 2020). All isolates showed circular, smooth-margined, yellow colonies when cultured on peptone sugar agar (PSA) medium for 24h at 28℃. The cells were all gram-negative and rod-shaped with three to six peritrichous flagella; positive for catalase, indole, glucose fermentation and citrate utilization, while negative for oxidase, alkaline, phenylalanine deaminase, urease, and nitrate reductase reactions. 16S rRNA gene sequence analysis from the 6 isolates (FY43, JH31, JH99, TZ20, TZ39 and TZ68) revealed that the amplified fragments shared 98% similarity with Pantoea ananatis type strain LMG 2665T (GenBank JFZU01) (Table S3). To further verify P. ananatis identity of these isolates, fragments of three housekeeping genes including gyrB, leuS and rpoB from the 6 isolates were amplified and sequenced, which showed highest homology to LMG 2665T with a sequence similarity of 95%-100% (Table S3). Primers (Brady et al. 2008) and GenBank accession numbers of gene sequences from the 6 isolates are listed in Table S1 and Table S2. Phylogenetic analysis of gyrB, leuS and rpoB concatenated sequences indicated that the 6 isolates were clustered in a stable branch with P. ananatis (Fig. S2). Based on the above morphological, physiological, biochemical and molecular data, the isolates are identified as P. ananatis. For pathogenicity tests, bacterial suspension at 108 CFU/mL was inoculated into flag leaves of rice (cv. Zhongzao39) at the late booting stage using clipping method. Water was used as a negative control. The clipped leaves displayed water-soaked lesions at 3 to 5 days after inoculation (DAI); then the lesion spread downward and turned light brown. At about 14 DAI, blight was shown with similar symptoms to those samples collected from the rice field of Zhejiang and Jiangxi provinces (Fig. S1). In contrast, the control plants remained healthy and symptomless. The same P. ananatis was re-isolated in the inoculated rice plants, fulfilling Koch's postulates. In the past decade, P. ananatis has been reported to cause grain discoloration in Hangzhou, China (Yan et al. 2010) and induce leaf blight as a companion of Enterobacter asburiae in Sichuan province, China (Xue et al. 2020). Nevertheless, to the best of our knowledge, this is the first report of P. ananatis as the causative agent of rice leaf blight in southeast China. This study raises the alarm that the emerging rice bacterial leaf blight in southeast China might be caused by a new pathogen P. ananatis, instead of Xoo as traditionally assumed. Further, the differences of occurrence, spread and control between two rice bacterial leaf blight diseases caused by P. ananatis and Xoo, respectively need to be determined in the future.

As core components of ABA signaling pathway, SnRK2s (Sucrose nonfermenting1–Related protein Kinase 2) bind to and phosphorylate AREB/ABF (ABA responsive element binding protein/ABRE-binding factor) transcriptional factors, particularly bZIPs (basic region-leucine zipper), to participate in various biological processes, including flowering. Rice contains 10 SnRK2 members denoted as SAPK1-10 (Stress-Activated Protein Kinase) and dozens of bZIPs. However, which of the SAPKs and bZIPs pair and involve in ABA signaling remains largely unknown. In this study, we carried out a systematical protein-protein interactomic analysis of 10 SAPKs and 9 ABA-inducible bZIPs using yeast-two-hybrid technique, and identified 14 positive interactions. The reliability of Y2H work was verified by in vitro pull-down assay of the key flowering regulator bZIP77 with SAPK9 and SAPK10, respectively. Moreover, SAPK10 could phosphorylate bZIP77 in vitro. Over-expression of SAPK10 resulted in earlier flowering time, at least partially through regulating the FAC-MADS15 pathway. Conclusively, our results provided an overall view of the SAPK-bZIP interactions, and shed novel lights on the mechanisms of ABA-regulated rice flowering.

The mycotoxin fumonisin is known to be harmful to humans and animals, and thus it is desirable to reduce fumonisin content in crop products. We explored the functions of several genes that function in fumonisin biosynthesis (FUM1, FUM6, FUM8, FUM19, and FUM21) in Fusarium proliferatum and found that deletion of FUM1, FUM6, FUM8, or FUM21 results in a severe reduction in fumonisin biosynthesis, while loss of FUM19 does not. In addition, fumonisin-deficient strains display significantly decreased pathogenicity. Co-cultivation of the ΔFUM1, ΔFUM6, ΔFUM8, and ΔFUM19 mutants restores fumonisin synthesis. However, co-cultivation was unable to restore fumonisin synthesis in the ΔFUM21 strain. The relative expression levels of three key FUM genes (FUM1, FUM6, and FUM8) differed significantly in each mutant strain; notably, the expression levels of these three genes were significantly down-regulated in the ΔFUM21 strain. Taken together, our results demonstrate that FUM1, FUM6, FUM8, and FUM21 are essential for fumonisin synthesis, and FUM19 is non-essential. Partial mutants lost the ability to synthesize fumonisin, the co-culture of the mutants was able to restore fumonisin biosynthesis. While the pathogenicity of F. proliferatum is affected by many factors, inhibition of the synthesis of the mycotoxin fumonisin will weaken the pathogenicity of rice spikelet rot disease (RSRD).