NRG1 functions downstream of EDS1 to regulate TIR-NLR-mediated plant immunity in Nicotiana benthamiana.

Effector-triggered immunity (ETI) in plants involves a large family of nucleotide-binding leucine-rich repeat (NLR) immune receptors, including Toll/IL-1 receptor-NLRs (TNLs) and coiled-coil NLRs (CNLs). Although various NLR immune receptors are known, a mechanistic understanding of NLR function in ETI remains unclear. The TNL Recognition of XopQ 1 (Roq1) recognizes the effectors XopQ and HopQ1 from Xanthomonas and Pseudomonas, respectively, which activates resistance to Xanthomonas euvesicatoria and Xanthomonas gardneri in an Enhanced Disease Susceptibility 1 (EDS1)-dependent way in Nicotiana benthamiana In this study, we found that the N. benthamiana N requirement gene 1 (NRG1), a CNL protein required for the tobacco TNL protein N-mediated resistance to tobacco mosaic virus, is also essential for immune signaling [including hypersensitive response (HR)] triggered by the TNLs Roq1 and Recognition of Peronospora parasitica 1 (RPP1), but not by the CNLs Bs2 and Rps2, suggesting that NRG1 may be a conserved key component in TNL signaling pathways. Besides EDS1, Roq1 and NRG1 are necessary for resistance to Xanthomonas and Pseudomonas in N. benthamiana NRG1 functions downstream of Roq1 and EDS1 and physically associates with EDS1 in mediating XopQ-Roq1-triggered immunity. Moreover, RNA sequencing analysis showed that XopQ-triggered gene-expression profile changes in N. benthamiana were almost entirely mediated by Roq1 and EDS1 and were largely regulated by NRG1. Overall, our study demonstrates that NRG1 is a key component that acts downstream of EDS1 to mediate various TNL signaling pathways, including Roq1 and RPP1-mediated HR, resistance to Xanthomonas and Pseudomonas, and XopQ-regulated transcriptional changes in N. benthamiana

Heritable virus-induced germline editing in tomato.

Here, we report the successful implementation of heritable virus-induced genome editing (VIGE) in tomato (Solanum lycopersicum). We generated three transgenic tomato lines expressing Streptococcus pyogenes Cas9 (SpCas9) under the control of Cauliflower mosaic virus 35S (35S), S. lycopersicum ribosomal protein S5A (SlRPS5A), or S. lycopersicum YAO promoters (SlYAO). These three lines were tested for somatic and heritable editing using the tobacco rattle virus (TRV)-based system carrying guide RNAs (gRNAs) fused with mobile RNA sequences. TRV with gRNA targeted to Phytoene desaturase (SlPDS) and Downy mildew resistance 6 (SlDMR6) genes fused to mobile RNA sequences showed significant somatic editing efficiency in all three tomato lines expressing SpCas9. However, the progenies from the SlYAO promoter-driven SpCas9 tomato infected with TRV with gRNA targeted to SlDMR6 fused to the mobile RNA sequence resulted in monoallelic mutations with a frequency of 3%. Optimization of environmental conditions, such as reduced light intensity, significantly increased heritable editing frequencies, from 0% to 86% at the SlPDS and from 3% to 100% at the SlDMR6, including biallelic mutations. These findings underscore the use of appropriate promoters to express Cas nucleases and optimized environmental conditions to enhance heritable genome editing efficiency in tomato using VIGE. Furthermore, our method enables the generation of mutants without additional tissue culture or transformation once a SpCas9-expressing tomato line is established

Optimized Agrobacterium-mediated sorghum transformation protocol and molecular data of transgenic sorghum plants

Agrobacterium-mediated sorghum transformation frequency has been enhanced significantly via medium optimization using immature embryos from sorghum variety TX430 as the target tissue. The new transformation protocol includes the addition of elevated copper sulfate and 6-benzylaminopurine in the resting and selection media. Using Agrobacterium strain LBA4404, the transformation frequency reached over 10% using either of two different selection marker genes, moPAT or PMI, and any of three different vectors in large-scale transformation experiments. With Agrobacterium strain AGL1, the transformation frequencies were as high as 33%. Using quantitative PCR analyses of 1,182 T(0) transgenic plants representing 675 independent transgenic events, data was collected for T-DNA copy number, intact or truncated T-DNA integration, and vector backbone integration into the sorghum genome. A comparison of the transformation frequencies and molecular data characterizing T-DNA integration patterns in the transgenic plants derived from LBA4404 versus AGL1 transformation revealed that twice as many transgenic high-quality events were generated when AGL1 was used compared to LBA4404. This is the first report providing molecular data for T-DNA integration patterns in a large number of independent transgenic plants in sorghum. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11627-013-9583-z) contains supplementary material, which is available to authorized users

Stability and inheritance of endosperm-specific expression of two transgenes in progeny from crossing independently transformed barley plants

To study stability and inheritance of two different transgenes in barley, we crossed a homozygous T8 plant, having uidA (or gus) driven by the barley endosperm-specific B1-hordein promoter (localized in the near centromeric region of chromosome 7H) with a second homozygous T4 plant, having sgfp(S65T) driven by the barley endosperm-specific D-hordein promoter (localized on the subtelomeric region of chromosome 2H). Both lines stably expressed the two transgenes in the generations prior to the cross. Three independently crossed F1 progeny were analyzed by PCR for both uidA and sgfp(S65T) in each plant and functional expression of GUS and GFP in F2 seeds followed a 3:1 Mendelian segregation ratio and transgenes were localized by FISH to the same location as in the parental plants. FISH was used to screen F2 plants for homozygosity of both transgenes; four homozygous plants were identified from the two crossed lines tested. FISH results showing presence of transgenes were consistent with segregation ratios of expression of both transgenes, indicating that the two transgenes were expressed without transgene silencing in homozygous progeny advanced to the F3 and F4 generations. Thus, even after crossing independently transformed, homozygous parental plants containing a single, stably expressed transgene, progeny were obtained that continued to express multiple transgenes through generation advance. Such stability of transgenes, following outcrossing, is an important attribute for trait modification and for gene flow studies

Dosage differences in 12-OXOPHYTODIENOATE REDUCTASE genes modulate wheat root growth

Wheat, an essential crop for global food security, is well adapted to a wide variety of soils. However, the gene networks shaping different root architectures remain poorly understood. We report here that dosage differences in a cluster of monocot-specific 12-OXOPHYTODIENOATE REDUCTASE genes from subfamily III (OPRIII) modulate key differences in wheat root architecture, which are associated with grain yield under water-limited conditions. Wheat plants with loss-of-function mutations in OPRIII show longer seminal roots, whereas increased OPRIII dosage or transgenic over-expression result in reduced seminal root growth, precocious development of lateral roots and increased jasmonic acid (JA and JA-Ile). Pharmacological inhibition of JA-biosynthesis abolishes root length differences, consistent with a JA-mediated mechanism. Transcriptome analyses of transgenic and wild-type lines show significant enriched JA-biosynthetic and reactive oxygen species (ROS) pathways, which parallel changes in ROS distribution. OPRIII genes provide a useful entry point to engineer root architecture in wheat and other cereals.Fil: Gabay, Gilad. University of California at Davis; Estados UnidosFil: Wang, Hanchao. University of California at Davis; Estados Unidos. University Of Haifa; IsraelFil: Zhang, Junli. University of California at Davis; Estados UnidosFil: Moriconi, Jorge Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Burguener, Germán Federico. University of California at Davis; Estados UnidosFil: Gualano, Leonardo David. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Howell, Tyson. University of California at Davis; Estados UnidosFil: Lukaszewski, Adam. University of California; Estados UnidosFil: Staskawicz, Brian. University of California; Estados UnidosFil: Cho, Myeong-Je. University of California; Estados UnidosFil: Tanaka, Jaclyn. University of California; Estados UnidosFil: Fahima, Tzion. University Of Haifa; IsraelFil: Ke, Haiyan. University of California; Estados UnidosFil: Dehesh, Katayoon. University of California; Estados UnidosFil: Zhang, Guo-Liang. Fudan University; ChinaFil: Gou, Jin Ying. Beijing Key Laboratory Of Crop Genetic Improvement; China. Fudan University; ChinaFil: Hamberg, Mats. Karolinska Huddinge Hospital. Karolinska Institutet; SueciaFil: Santa Maria, Guillermo Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Dubcovsky, Jorge. University of California at Davis; Estados Unidos. Howard Hughes Medical Institute; Estados Unido

Dosage differences in 12-OXOPHYTODIENOATE REDUCTASE genes modulate wheat root growth

Wheat, an essential crop for global food security, is well adapted to a wide variety of soils. However, the gene networks shaping different root architectures remain poorly understood. We report here that dosage differences in a cluster of monocot-specific 12-OXOPHYTODIENOATE REDUCTASE genes from subfamily III (OPRIII) modulate key differences in wheat root architecture, which are associated with grain yield under water-limited conditions. Wheat plants with loss-of-function mutations in OPRIII show longer seminal roots, whereas increased OPRIII dosage or transgenic over-expression result in reduced seminal root growth, precocious development of lateral roots and increased jasmonic acid (JA and JA-Ile). Pharmacological inhibition of JA-biosynthesis abolishes root length differences, consistent with a JA-mediated mechanism. Transcriptome analyses of transgenic and wild-type lines show significant enriched JA-biosynthetic and reactive oxygen species (ROS) pathways, which parallel changes in ROS distribution. OPRIII genes provide a useful entry point to engineer root architecture in wheat and other cereals.Fil: Gabay, Gilad. University of California at Davis; Estados UnidosFil: Wang, Hanchao. University of California at Davis; Estados Unidos. University Of Haifa; IsraelFil: Zhang, Junli. University of California at Davis; Estados UnidosFil: Moriconi, Jorge Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Burguener, Germán Federico. University of California at Davis; Estados UnidosFil: Gualano, Leonardo David. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Howell, Tyson. University of California at Davis; Estados UnidosFil: Lukaszewski, Adam. University of California; Estados UnidosFil: Staskawicz, Brian. University of California; Estados UnidosFil: Cho, Myeong-Je. University of California; Estados UnidosFil: Tanaka, Jaclyn. University of California; Estados UnidosFil: Fahima, Tzion. University Of Haifa; IsraelFil: Ke, Haiyan. University of California; Estados UnidosFil: Dehesh, Katayoon. University of California; Estados UnidosFil: Zhang, Guo-Liang. Fudan University; ChinaFil: Gou, Jin Ying. Beijing Key Laboratory Of Crop Genetic Improvement; China. Fudan University; ChinaFil: Hamberg, Mats. Karolinska Huddinge Hospital. Karolinska Institutet; SueciaFil: Santa Maria, Guillermo Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Dubcovsky, Jorge. University of California at Davis; Estados Unidos. Howard Hughes Medical Institute; Estados Unido

Probiotics partially attenuate the severity of acute kidney injury through an immunomodulatory effect

Background A healthy microbiome helps maintain the gut barrier and mucosal immune tolerance. Previously, we demonstrated that acute kidney injury (AKI) provoked dysbiosis, gut inflammation, and increased permeability. Here, we investigated the renoprotective effects of the probiotic Bifidobacterium bifidum BGN4 and the underlying mechanisms thereof. Methods C57BL/6 mice were subjected to bilateral renal ischemia-reperfusion injury (IRI) or sham operation. In the probiotic-treated group, BGN4 was administered by gavage once daily, starting 2 weeks before injury. Results Administration of BGN4 significantly increased gut microbiome diversity and prevented expansion of the Enterobacteriaceae and Bacteroidetes that were the hallmarks of AKI-induced dysbiosis. Further, BGN4 administration also significantly reduced other IRI-induced changes in the colon microenvironment, including effects on permeability, apoptosis of colon epithelial cells, and neutrophil and proinflammatory macrophage infiltration. Mononuclear cells co-cultured with BGN4 expressed significantly increased proportions of CD103+/CD11c+ and CD4+ CD25+ Treg cells, suggesting a direct immunomodulatory effect. BGN4 induced Treg expansion in colon, mesenteric lymph nodes (MNL), and kidney. BGN4 also reduced CX3CR1intermediateLy6Chigh monocyte infiltration and interleukin (IL)-17A suppression in the small intestine, which may have attenuated AKI severity, kidney IL-6 messenger RNA expression, and AKI-induced liver injury. Conclusion Prior supplementation with BGN4 significantly attenuated the severity of IRI and secondary liver injury. This renoprotective effect was associated with increased Foxp3 and reduced IL-17A expression in the colon, MNL, and kidney, suggesting that BGN4-induced immunomodulation might contribute to its renoprotective effects. Probiotics may therefore be a promising strategy to reduce AKI severity and/or remote organ injury

Chromosome-level genome assembly of a regenerable maize inbred line A188.

BACKGROUND The maize inbred line A188 is an attractive model for elucidation of gene function and improvement due to its high embryogenic capacity and many contrasting traits to the first maize reference genome, B73, and other elite lines. The lack of a genome assembly of A188 limits its use as a model for functional studies. RESULTS Here, we present a chromosome-level genome assembly of A188 using long reads and optical maps. Comparison of A188 with B73 using both whole-genome alignments and read depths from sequencing reads identify approximately 1.1 Gb of syntenic sequences as well as extensive structural variation, including a 1.8-Mb duplication containing the Gametophyte factor1 locus for unilateral cross-incompatibility, and six inversions of 0.7 Mb or greater. Increased copy number of carotenoid cleavage dioxygenase 1 (ccd1) in A188 is associated with elevated expression during seed development. High ccd1 expression in seeds together with low expression of yellow endosperm 1 (y1) reduces carotenoid accumulation, accounting for the white seed phenotype of A188. Furthermore, transcriptome and epigenome analyses reveal enhanced expression of defense pathways and altered DNA methylation patterns of the embryonic callus. CONCLUSIONS The A188 genome assembly provides a high-resolution sequence for a complex genome species and a foundational resource for analyses of genome variation and gene function in maize. The genome, in comparison to B73, contains extensive intra-species structural variations and other genetic differences. Expression and network analyses identify discrete profiles for embryonic callus and other tissues

Author Correction: Battery-free, wireless soft sensors for continuous multi-site measurements of pressure and temperature from patients at risk for pressure injuries

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