Rice small secreted peptide, OsRALF26, recognized by FERONIA-like receptor 1 induces immunity in rice and Arabidopsis.

Rapid alkalinization factors (RALFs), belonging to a family of small secreted peptides, have been considered as important signaling molecules in diverse biological processes, including immunity. Current studies on RALF-modulated immunity mainly focus on Arabidopsis, but little is reported in crop plants. The rice immune receptor XA21 confers immunity to the bacterial blight pathogen, Xanthomonas oryzae pv. oryzae (Xoo). Here, we pursued functional characterization of rice RALF26 (OsRALF26) up-regulated by Xoo during XA21-mediated immune response. When applied exogenously as a recombinant peptide, OsRALF26 induced a series of immune responses, including pathogenesis-related genes (PRs) induction, reactive oxygen species (ROS) production, and callose deposition in rice and/or Arabidopsis. Transgenic rice and Arabidopsis overexpressing OsRALF26 exhibited significantly enhanced resistance to Xoo and Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), respectively. In yeast two-hybrid, pull-down assays, and co-immunoprecipitation analyses, rice FER-like receptor 1 (OsFLR1) was identified as a receptor of OsRALF26. Transient expression of OsFLR1 in Nicotiana benthamiana leaves displayed significantly increased ROS production and callose deposition after OsRALF26 treatment. Together, we propose that OsRALF26 induced by Xoo in an XA21-dependent manner is perceived by OsFLR1 and may play a novel role in the enforcement of XA21-mediated immunity.

Rice NLR protein XinN1, induced by a pattern recognition receptor XA21, confers enhanced resistance to bacterial blight.

KEY MESSAGE: Rice CC-type NLR XinN1, specifically induced by a PRR XA21, activates defense pathways against Xoo. Plants have evolved two layers of immune systems regulated by two different types of immune receptors, cell surface located pattern recognition receptors (PRRs) and intracellular nucleotide-binding domain leucine-rich repeat-containing receptors (NLRs). Plant PRRs recognize conserved molecular patterns from diverse pathogens, resulting in pattern-triggered immunity (PTI), whereas NLRs are activated by effectors secreted by pathogens into plant cells, inducing effector-triggered immunity (ETI). Rice PRR, XA21, recognizes a tyrosine-sulfated RaxX peptide (required for activation of XA21-mediated immunity X) as a molecular pattern secreted by Xanthomonas oryzae pv. oryzae (Xoo). Here, we identified a rice NLR gene, XinN1, that is specifically induced during the XA21-mediated immune response against Xoo. Transgenic rice plants overexpressing XinN1 displayed increased resistance to infection by Xoo with reduced lesion length and bacterial growth. Overexpression of autoactive mutant of XinN1 (XinN1D543V) also displayed increased resistance to Xoo, accompanied with severe growth retardation and cell death. In rice protoplast system, overexpression of XinN1 or XinN1D543V significantly elevated reactive oxygen species (ROS) production and cytosolic-free calcium (Ca2+) accumulations. In addition, XinN1 overexpression additionally elevated the ROS burst caused by the interaction between XA21 and RaxX-sY and induced the transcription of PTI signaling components, including somatic embryogenesis receptor kinases (OsSERKs) and receptor-like cytoplasmic kinases (OsRLCKs). Our results suggest that XinN1 induced by the PRR XA21 activates defense pathways and provides enhanced resistance to Xoo in rice.

Oryza-Specific Orphan Protein Triggers Enhanced Resistance to Xanthomonas oryzae pv. oryzae in Rice.

All genomes carry lineage-specific orphan genes lacking homology in their closely related species. Identification and functional study of the orphan genes is fundamentally important for understanding lineage-specific adaptations including acquirement of resistance to pathogens. However, most orphan genes are of unknown function due to the difficulties in studying them using helpful comparative genomics. Here, we present a defense-related Oryza-specific orphan gene, Xio1, specifically induced by the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo) in an immune receptor XA21-dependent manner. Salicylic acid (SA) and ethephon (ET) also induced its expression, but methyl jasmonic acid (MeJA) reduced its basal expression. C-terminal green fluorescent protein (GFP) tagged Xio1 (Xio1-GFP) was visualized in the nucleus and the cytosol after polyethylene glycol (PEG)-mediated transformation in rice protoplasts and Agrobacterium-mediated infiltration in tobacco leaves. Transgenic rice plants overexpressing Xio1-GFP showed significantly enhanced resistance to Xoo with reduced lesion lengths and bacterial growth, in company with constitutive expression of defense-related genes. However, all of the transgenic plants displayed severe growth retardation and premature death. Reactive oxygen species (ROS) was significantly produced in rice protoplasts constitutively expressing Xio1-GFP. Overexpression of Xio1-GFP in non-Oryza plant species, Arabidopsis thaliana, failed to induce growth retardation and enhanced resistance to Pseudomonas syringae pv. tomato (Pst) DC3000. Our results suggest that the defense-related orphan gene Xio1 plays an important role in distinctive mechanisms evolved within the Oryza and provides a new source of Oryza-specific genes for crop-breeding programs.

Population Structure of Stagonosporopsis Species Associated with Cucurbit Gummy Stem Blight in Korea.

Gummy stem blight (GSB), a common and serious disease in cucurbits worldwide, is caused by three genetically distinct species: Stagonosporopsis cucurbitacearum (syn. Didymella bryoniae), S. citrulli, and S. caricae. In Korea, however, the three species of Stagonosporopsis have been barely characterized. In this study, 21 Stagonosporopsis isolates were recovered from watermelon (Citrullus lanatus) and muskmelon (Cucumis melo) leaves and stem showing blight symptoms collected from 43 fields in Korea. Sequence analysis performed with an internal transcribed spacer region was not competent to differentiate the Stagonosporopsis isolates. On the contrary, analysis of ß-tubulin (TUB) genes and three microsatellite markers, Db01, Db05, and Db06, successfully differentiated Stagonosporopsis isolates. Further sequence analysis identified two Stagonosporopsis species, S. citrulli and S. caricae, and one previously unknown species of Stagonosporopsis. Representative isolates from three species caused dark water-soaked lesions on the detached watermelon and muskmelon leaves with no significant differences in the aggressiveness. Our results indicate that the S. citrulli, S. caricae, and unknown Stagonosporopsis sp. are all causal agents of GSB for both watermelon and muskmelon. This is the first report of a new species and the population structure of Stagonosporopsis species causing GSB in Korea.

Incidence of Alternaria Species Associated with Watermelon Leaf Blight in Korea.

Alternaria leaf blight is one of the most common diseases in watermelon worldwide. In Korea, however, the Alternaria species causing the watermelon leaf blight have not been investigated thoroughly. A total of 16 Alternaria isolates was recovered from diseased watermelon leaves with leaf blight symptoms, which were collected from 14 fields in Korea. Analysis of internal transcribed spacer (ITS) region, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and RNA polymerase II second largest subunit (RPB2) were not competent to differentiate the Alternaria isolates. On the contrary, analysis of amplicon size of the histone H3 (HIS3) gene successfully differentiated the isolates into three Alternaria subgroups, and further sequence analysis of them identified three Alternaria spp. Alternaria tenuissima, A. gaisen, and A. alternata. Representative Alternaria isolates from three species induced dark brown leaf spot lesions on detached watermelon leaves, indicating that A. tenuissima, A. gaisen, and A. alternata are all causal agents of Alternaria leaf blight. Our results indicate that the Alternaria species associated watermelon leaf blight in Korea is more complex than reported previously. This is the first report regarding the population structure of Alternaria species causing watermelon leaf blight in Korea.

Gene expression profile of Th1 and Th2 cytokines and their receptors in human and nonhuman primates.

BACKGROUND: To date comparative knowledge concerning gene expression profiles of T-helper 1(Th1)/Th2 cytokines and their receptors between human and non-human primates is scarce. METHODS: We assessed the gene expression level of both Th1 [interleukin-4(IL-4)] and Th2 [IL-12, interferon-gamma(IFN-gamma)] cytokines and the receptors (IL-4Ralpha, IFN-gammaR1, IFN-gammaR2) in peripheral blood mononuclear cells (PBMC) from humans, chimpanzee, baboon, and macaque by a quantitative real-time reverse transcriptase-polymerase chain reaction(RT-PCR). RESULTS: The expression level of the IFN-gamma gene was markedly lower in humans than that in non-human primates. The IL-4 gene expression was significantly higher, whereas that of IL-12 was distinctly lower, in human/chimpanzee than in baboon/macaque. The IFN-gammaR2 gene expression was especially higher in the macaque than in the other three primates. CONCLUSIONS: These results indicate distinct gene expression of Th1/Th2 cytokines and their receptors in primates. These also suggest characteristic differences in Th1/Th2 immune responses affecting host defense and/or disease susceptibility among these primates.