Rice microRNA156/529-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE7/14/17 modules regulate defenses against bacteria.

Rice (Oryza sativa L.) microRNA156/529-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE7/14/17 (miR156/529-SPL7/14/17) modules have pleiotropic effects on many biological pathways. OsSPL7/14 can interact with DELLA protein SLENDER RICE1 (SLR1) to modulate gibberellin acid (GA) signal transduction against the bacterial pathogen Xanthomonas oryzae pv. oryzae. However, whether the miR156/529-OsSPL7/14/17 modules also regulate resistance against other pathogens is unclear. Notably, OsSPL7/14/17 functioning as transcriptional activators, their target genes, and the corresponding downstream signaling pathways remain largely unexplored. Here, we demonstrate that miR156/529 play negative roles in plant immunity and that miR156/529-regulated OsSPL7/14/17 confer broad-spectrum resistance against 2 devastating bacterial pathogens. Three OsSPL7/14/17 proteins directly bind to the promoters of rice Allene Oxide Synthase 2 (OsAOS2) and NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1 (OsNPR1) and activate their transcription, regulating jasmonic acid (JA) accumulation and the salicylic acid (SA) signaling pathway, respectively. Overexpression of OsAOS2 or OsNPR1 impairs the susceptibility of the osspl7/14/17 triple mutant. Exogenous application of JA enhances resistance of the osspl7/14/17 triple mutant and the miR156 overexpressing plants. In addition, genetic evidence confirms that bacterial pathogen-activated miR156/529 negatively regulate pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) responses, such as pattern recognition receptor Xa3/Xa26-initiated PTI. Our findings demonstrate that bacterial pathogens modulate miR156/529-OsSPL7/14/17 modules to suppress OsAOS2-catalyzed JA accumulation and the OsNPR1-promoted SA signaling pathway, facilitating pathogen infection. The uncovered miR156/529-OsSPL7/14/17-OsAOS2/OsNPR1 regulatory network provides a potential strategy to genetically improve rice disease resistance.

Optimizing nutrient transporters to enhance disease resistance in rice.

Fertilizers and plant diseases contribute positively and negatively to crop production, respectively. Macro- and micronutrients provided by the soil and fertilizers are transported by various plant nutrient transporters from the soil to the roots and shoots, facilitating growth and development. However, the homeostasis of different nutrients has different effects on plant disease. This review is aimed at providing insights into the interconnected regulation between nutrient homeostasis and immune responses, and it highlights strategies to enhance disease resistance by optimal manipulation of nutrient transporters in rice. First, we highlight the essential roles of six macronutrients (nitrogen, phosphorus, potassium, sulfur, calcium, magnesium) and eight micronutrients (iron, manganese, zinc, copper, boron, molybdenum, silicon, nickel), and summarize the diverse effects of each on rice diseases. We then systematically review the molecular mechanisms of immune responses modulated by nutrient transporters and the genetic regulatory pathways that control the specific nutrient-mediated immune signaling that is regulated by the pathogens and the host plant. Finally, we discuss putative strategies for breeding disease-resistant rice by genetic engineering of nutrient transporters.

The versatile functions of OsALDH2B1 provide a genic basis for growth-defense trade-offs in rice.

In plants, enhanced defense often compromises growth and development, which is regarded as trade-offs between growth and defense. Here we identified a gene, OsALDH2B1, that functions as a master regulator of the growth-defense trade-off in rice. OsALDH2B1 has its primary function as an aldehyde dehydrogenase and a moonlight function as a transcriptional regulator. Loss of function of OsALDH2B1 greatly enhanced resistance to broad-spectrum pathogens, including fungal blast, bacterial leaf blight, and leaf streak, but caused severe phenotypic changes such as male sterility and reduced plant size, grain size, and number. We showed that its primary function as a mitochondrial aldehyde dehydrogenase conditions male fertility. Its moonlight function of transcriptional regulation, featuring both repressing and activating activities, regulates a diverse range of biological processes involving brassinolide, G protein, jasmonic acid, and salicylic acid signaling pathways. Such regulations cause large impacts on the morphology and immunity of rice plants. The versatile functions of OsALDH2B1 provide an example of the genic basis of growth-defense trade-offs in plants.

The group I GH3 family genes encoding JA-Ile synthetase act as positive regulator in the resistance of rice to Xanthomonas oryzae pv. oryzae.

Plant GH3 genes are key components of the hormonal mechanism regulating growth and development, responding to biotic and abiotic stress. GH3 proteins are involved in hormonal homeostasis through conjugation to amino acids of the free form of salicylic acid, jasmonic acid (JA) or indole-3-acetic acid (IAA). Our previous work has uncovered that two GH3 genes encoding IAA-amido synthetase play important roles in the resistance to bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) in rice, however, whether other rice GH3 genes play roles in resistance to Xoo is unclear. Here, we validated that GH3.3, GH3.5, GH3.6 and GH3.12, four members of group I GH3 family, are the functional JA-Ile synthetases by catalyzing the conversion of free JA into active form of JA-Ile in vitro and in vivo. The overexpressing plants of four genes individually accumulated less JA but more JA-Ile than the wild type plants. Conversely, the corresponding suppressing plants of four genes contained comparable JA and JA-Ile concentrations, but the triple and quadruple suppressing plants had lower level of JA-Ile compared with wild type plants, suggesting functional redundancy of the same clade of GH3 family. Furthermore, the group I GH3 family genes positively mediated rice resistance to bacterial pathogen Xoo through modulating JA homeostasis and regulating transcription pattern of JA-responsive genes.

Cadmium accumulation regulated by a rice heavy-metal importer is harmful for host plant and leaf bacteria.

INTRODUCTION: Cadmium (Cd), one of the major toxic heavy metals, causes severe deleterious effects on all living organisms from prokaryotes to eukaryotes. Cadmium deposition affects bacterial diversity and bacterial population in soil. Cadmium accumulation in plants is mainly controlled by transporters and the resulting Cd enrichment gives rise to phytotoxicity. OBJECTIVE: This study aimed to mine transporters that control Cd import or accumulation in rice and uncover the underlying mechanisms that how accumulated Cd poses risks to host plant and leaf bacteria. METHODS: RNA-seq analysis, histochemical assays, and elemental quantification were carried out to reveal the biological roles of OsABCG43 for Cd import. Pathogen inoculation, IC50 value, and bacterial virulence assays were conducted to disclose the effects of Cd on leaf bacteria. RESULTS: OsABCG43 is characterized as a Cd importer controlling Cd accumulation in rice. OsABCG43 was induced under Cd stress and specifically expressed in the vasculature of leaves and roots. Overexpression of OsABCG43 caused Cd accumulation which inhibits photosynthesis and development and alters the antioxidant system, resulting in phytotoxicity. Moreover, overexpression of OsABCG43 resulted in retarded plant growth and enhanced rice sensitivity to Cd stress. Numerous differentially expressed genes were identified via RNA-seq analysis between the OsABCG43-overexpressing plants and wild type, which functioned in Cd or reactive oxygen species (ROS) homeostasis. In addition, OsABCG43 transcripts were induced by leaf bacteria Xanthomonas oryzae pv. oryzicola (Xoc) and X. oryzae pv. oryzae (Xoo). The enriched Cd directly impaired the formation of virulence factors for the leaf bacteria, preventing colonization or proliferation of Xoc or Xoo in rice leaves. CONCLUSION: This work reveals that OsABCG43 is expressed specifically in the vascular and plasma membrane-localized OsABCG43 functions as a Cd importer. OsABCG43-mediated import of Cd is harmful for both rice and the corresponding leaf bacteria.

An MKP-MAPK protein phosphorylation cascade controls vascular immunity in plants.

Global crop production is greatly reduced by vascular diseases. These diseases include bacterial blight of rice and crucifer black rot caused by Xanthomonas oryzae pv. oryzae (Xoo) and Xanthomonas campestris pv. campestris (Xcc). The molecular mechanisms that activate vascular defense against such pathogens remains underexplored. Here, we show that an Arabidopsis MAPK phosphatase 1 (MKP1) mutant has increased host susceptibility to the adapted pathogen Xcc and is compromised in nonhost resistance to the rice pathogen Xoo. MKP1 regulates MAPK-mediated phosphorylation of the transcription factor MYB4 that negatively regulates vascular lignification through inhibiting lignin biosynthesis. Induction of lignin biosynthesis is, therefore, an important part of vascular-specific immunity. The role of MKP-MAPK-MYB signaling in lignin biosynthesis and vascular resistance to Xoo is conserved in rice, indicating that these factors form a tissue-specific defense regulatory network. Our study likely reveals a major vascular immune mechanism that underlies tissue-specific disease resistance against bacterial pathogens in plants.

miR395-regulated sulfate metabolism exploits pathogen sensitivity to sulfate to boost immunity in rice.

MicroRNAs (miRNAs) play important roles in plant physiological activities. However, their roles and molecular mechanisms in boosting plant immunity, especially through the modulation of macronutrient metabolism in response to pathogens, are largely unknown. Here, we report that an evolutionarily conserved miRNA, miR395, promotes resistance to Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc), two destructive bacterial pathogens, by regulating sulfate accumulation and distribution in rice. Specifically, miR395 targets and suppresses the expression of the ATP sulfurylase gene OsAPS1, which functions in sulfate assimilation, and two sulfate transporter genes, OsSULTR2;1 and OsSULTR2;2, which function in sulfate translocation, to promote sulfate accumulation, resulting in broad-spectrum resistance to bacterial pathogens in miR395-overexpressing plants. Genetic analysis revealed that miR395-triggered resistance is involved in both pathogen-associated molecular pattern-triggered immunity and R gene-mediated resistance. Moreover, we found that accumulated sulfate but not S-metabolites inhibits proliferation of pathogenic bacteria, revealing a sulfate-mediated antibacterial defense mechanism that differs from sulfur-induced resistance. Furthermore, compared with other bacteria, Xoo and Xoc, which lack the sulfate transporter CysZ, are sensitive to high levels of extracellular sulfate. Accordingly, miR395-regulated sulfate accumulation impaired the virulence of Xoo and Xoc by decreasing extracellular polysaccharide production and biofilm formation. Taken together, these results suggest that rice miR395 modulates sulfate metabolism to exploit pathogen sensitivity to sulfate and thereby promotes broad-spectrum resistance.

Rice group I GH3 gene family, positive regulators of bacterial pathogens.

Plant GH3 genes play pivotal roles in biotic stress through involving in hormonal homeostasis by conjugation to amino acids of the free-form of salicylic acid, jasmonic acid (JA) or indole-3-acetic acid. We recently showed that rice group I GH3 gene family, with four members, are the functional JA-Ile synthetases and positively mediated rice resistance to Xanthomonas oryzae pv. oryzae (Xoo). Here, we further found that these four genes are also positive regulators conferring resistance to Xanthomonas oryzae pv. oryzicola (Xoc), the devastating bacterial pathogen of rice. The transcript of these four genes were all activated upon Xoc invasion. The overexpressing plants showed less lesion length in comparison with wild type plant accompanying with higher pathogenesis-related genes accumulation, while the triple and quadruple suppressing plants showed susceptible to Xoc with less pathogenesis-related genes accumulation. Previous and present work demonstrate that rice group I GH3 family genes act as positive regulators in the resistance to Xoo and Xoc.

TALE-carrying bacterial pathogens trap host nuclear import receptors for facilitation of infection of rice.

Many plant-pathogenic Xanthomonas rely on the secretion of virulence transcription activator-like effector (TALE) proteins into plant cells to activate plant susceptibility genes to cause disease. The process is dependent on the binding of TALEs to specific elements of host target gene promoters in the plant nucleus. However, it is unclear how TALEs, after injection into host cells, are transferred from the plant cytoplasm into the plant nucleus, which is the key step of successful pathogen infection. Here, we show that the host plant cytoplasm/nuclear shuttle proteins OsImpα1a and OsImpα1b are key components for infection by the TALE-carrying bacterial pathogens Xanthomonas oryzae pv. oryzae (Xoo) and Xanthomonas oryzae pv. oryzicola (Xoc), the causal agents of bacterial leaf blight and bacterial leaf streak, respectively, in rice. Direct interaction between the second nuclear localization signal of TALEs of Xoo or Xoc and OsImpα1a or OsImpα1b is required for the transportation of TALEs into the nucleus. Conversely, suppression of the expression of OsImpα1a and OsImpα1b genes attenuates the shuttling of TALEs from the cytoplasm into the nucleus and the induction of susceptibility genes, thus improving the broad-spectrum disease resistance of rice to Xoo and Xoc. These results provide an applicable strategy for the improvement of resistance to TALE-carrying pathogens in rice by moderate suppression of the expression of plant nuclear import receptor proteins.

Jasmonic Acid-Involved OsEDS1 Signaling in Rice-Bacteria Interactions.

BACKGROUND: The function of Arabidopsis enhanced disease susceptibility 1 (AtEDS1) and its sequence homologs in other dicots have been extensively studied. However, it is unknown whether rice EDS1 homolog (OsEDS1) plays a role in regulating the rice-pathogen interaction. RESULTS: In this study, a OsEDS1-knouckout mutant (oseds1) was characterized and shown to have increased susceptibility to Xanthomonas oryzae pv. oryzae (Xoo) and Xanthomonas oryzae pv. oryzicola (Xoc), suggesting the positive role of OsEDS1 in regulating rice disease resistance. However, the following evidence suggests that OsEDS1 shares some differences with AtEDS1 in its way to regulate the host-pathogen interactions. Firstly, OsEDS1 modulates the rice-bacteria interactions involving in jasmonic acid (JA) signaling pathway, while AtEDS1 regulates Arabidopsis disease resistance against biotrophic pathogens depending on salicylic acid (SA) signaling pathway. Secondly, introducing AtEDS1 could reduce oseds1 mutant susceptibility to Xoo rather than to Xoc. Thirdly, exogenous application of JA and SA cannot complement the susceptible phenotype of the oseds1 mutant, while exogenous application of SA is capable of complementing the susceptible phenotype of the ateds1 mutant. Finally, OsEDS1 is not required for R gene mediated resistance, while AtEDS1 is required for disease resistance mediated by TIR-NB-LRR class of R proteins. CONCLUSION: OsEDS1 is a positive regulator in rice-pathogen interactions, and shares both similarities and differences with AtEDS1 in its way to regulate plant-pathogen interactions.

The host basal transcription factor IIA subunits coordinate for facilitating infection of TALEs-carrying bacterial pathogens in rice.

Many plant-pathogenic Xanthomonas rely largely on secreting virulence transcription activator-like effectors (TALEs) proteins into plant nucleus to activate host susceptibility genes to cause disease, the process is dependent on pathogen TALEs association with host plants basal transcription factor IIA small subunit TFIIAγ. TFIIAγ together with large subunit TFIIAαß constitute as a key component of RNA polymerase II complex for transcriptome initiation. However, whether TFIIAαß coordinates or competes with pathogen TALEs for interaction with TFIIAγ to activate transcript of TALEs-targeting genes is unclear. Here, we showed that TALE-carrying bacterial pathogens Xanthomonas oryzae pv. oryzae (Xoo) and Xanthomonas oryzae pv. oryzicola (Xoc), the causal agent for bacterial leaf blight and bacterial leaf streak in rice, using their major virulence TALEs to physically associate with N-terminal of OsTFIIAγ5. OsTFIIAα and OsTFIIAß which are post-translationally mature proteins of OsTFIIAαß separately bound to N- and C-terminal of OsTFIIAγ5. OsTFIIAα coordinated with TALEs for binding with OsTFIIAγ5 to upregulate rice susceptibility genes to cause disease. Conversely, suppression of OsTFIIAαß attenuated TALEs-targeting genes transcription, thus improved broad-spectrum disease resistance of rice to Xoo and Xoc. These results provide an applicable strategy for improving resistance to TALE-carrying pathogens in rice by appropriate suppression of plant basal transcription factors expression.

Dynamic phytohormone profiling of rice upon rice black-streaked dwarf virus invasion.

Rice black-streaked dwarf virus (RBSDV) is the causal agent of rice black-streaked dwarf disease, a serious constraint to rice production. A great deal of effort has been made to elucidate the transcriptome and proteome changes of rice upon virus inoculation. However, the relationship between RBSDV invasion and rice endogenous phytohormone profiling is largely unclear. Here, we surveyed the dynamic content profiling of endogenous phytohormones, which were severely disturbed by RBSDV invasion. The levels of abscisic acid (ABA) and cytokinins (CTKs) increased, while indole-3-acetic acid (IAA), gibberellins (GAs), jasmonic acid (JA), and salicylic acid (SA) decreased, accompanied by changes in the transcripts of genes participating in phytohormone metabolism and signalling pathways. Moreover, exogenously supplied GA3 could rescue the typical dwarfing symptom, and pre-application of SA largely decreased the occurrence of RBSDV disease in rice. The results partially suggest that RBSDV successfully invaded host rice by modulating the expression patterns of phytohormone metabolism to upset the balance of plant endogenous phytohormones.

Xanthomonas oryzae pv. oryzae Inoculation and Growth Rate on Rice by Leaf Clipping Method.

Bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most serious bacterial diseases and a major impediment to the increase of rice yield. Appropriate methods for inoculation of Xoo and disease scoring are necessary to investigate the nature of the disease and the mechanism of plant resistance to the pathogen. As the most-widely grown crop in the worldwide, rice yield plays an important role in food security. Uncovering mechanisms of plant-pathogen interaction of rice and Xoo will help develop rice plants that are more resistant to disease caused by Xoo. Here we describe our validated and efficient methods for inoculation of Xoo and disease scoring.

A Conserved Basal Transcription Factor Is Required for the Function of Diverse TAL Effectors in Multiple Plant Hosts.

Many Xanthomonas bacteria use transcription activator-like effector (TALE) proteins to activate plant disease susceptibility (S) genes, and this activation contributes to disease. We recently reported that rice basal transcription factor IIA gamma subunit, OsTFIIAγ5, is hijacked by TALE-carrying Xanthomonas oryzae infecting the plants. However, whether TFIIAγs are also involved in TALE-carrying Xanthomonas-caused diseases in other plants is unknown. Here, molecular and genetic approaches were used to investigate the role of TFIIAγs in other plants. We found that TFIIAγs are also used by TALE-carrying Xanthomonas to cause disease in other plants. The TALEs of Xanthomonas citri pv. citri (Xcc) causing canker in citrus and Xanthomonas campestris pv. vesicatoria (Xcv) causing bacterial spot in pepper and tomato interacted with corresponding host TFIIAγs as in rice. Transcriptionally suppressing TFIIAγ led to resistance to Xcc in citrus and Xcv in pepper and tomato. The 39th residue of OsTFIIAγ5 and citrus CsTFIIAγ is vital for TALE-dependent induction of plant S genes. As mutated OsTFIIAγ5V 39E, CsTFIIAγV 39E, pepper CaTFIIAγV 39E, and tomato SlTFIIAγV 39E also did not interact with TALEs to prevent disease. These results suggest that TALE-carrying bacteria share a common mechanism for infecting plants. Using TFIIAγV 39E-type mutation could be a general strategy for improving resistance to TALE-carrying pathogens in crops.