Plasma membrane association and resistosome formation of plant helper immune receptors.

Intracellular plant immune receptors, termed NLRs (Nucleotide-binding Leucine-rich repeat Receptors), confer effector-triggered immunity. Sensor NLRs are responsible for pathogen effector recognition. Helper NLRs function downstream of sensor NLRs to transduce signaling and induce cell death and immunity. Activation of sensor NLRs that contain TIR (Toll/interleukin-1receptor) domains generates small molecules that induce an association between a downstream heterodimer signalosome of EDS1 (EnhancedDisease Susceptibility 1)/SAG101 (Senescence-AssociatedGene 101) and the helper NLR of NRG1 (NRequired Gene 1). Autoactive NRG1s oligomerize and form calcium signaling channels largely localized at the plasma membrane (PM). The molecular mechanisms of helper NLR PM association and effector-induced NRG1 oligomerization are not well characterized. We demonstrate that helper NLRs require positively charged residues in their N-terminal domains for phospholipid binding and PM association before and after activation, despite oligomerization and conformational changes that accompany activation. We demonstrate that effector activation of a TIR-containing sensor NLR induces NRG1 oligomerization at the PM and that the cytoplasmic pool of EDS1/SAG101 is critical for cell death function. EDS1/SAG101 cannot be detected in the oligomerized NRG1 resistosome, suggesting that additional unknown triggers might be required to induce the dissociation of EDS1/SAG101 from the previously described NRG1/EDS1/SAG101 heterotrimer before subsequent NRG1 oligomerization. Alternatively, the conformational changes resulting from NRG1 oligomerization abrogate the interface for EDS1/SAG101 association. Our data provide observations regarding dynamic PM association during helper NLR activation and underpin an updated model for effector-induced NRG1 resistosome formation.

Genomic characterization and expression profiles of stress-associated proteins (SAPs) in castor bean (Ricinus communis).

Stress-associated proteins (SAPs) are known as response factors to multiple abiotic and biotic stresses in plants. However, the potential physiological and molecular functions of SAPs remain largely unclear. Castor bean (Ricinus communis L.) is one of the most economically valuable non-edible woody oilseed crops, able to be widely cultivated in marginal lands worldwide because of its broad adaptive capacity to soil and climate conditions. Whether SAPs in castor bean plays a key role in adapting diverse soil conditions and stresses remains unknown. In this study, we used the castor bean genome to identify and characterize nine castor bean SAP genes (RcSAP). Structural analysis showed that castor bean SAP gene structures and functional domain types vary greatly, differing in intron number, protein sequence, and functional domain type. Notably, the AN1-C2H2-C2H2 zinc finger domain within RcSAP9 has not been often observed in other plant families. High throughput RNA-seq data showed that castor bean SAP gene profiles varied among different tissues. In addition, castor bean SAP gene expression varied in response to different stresses, including salt, drought, heat, cold and ABA and MeJA, suggesting that the transcriptional regulation of castor bean SAP genes might operate independently of each other, and at least partially independent from ABA and MeJA signal pathways. Cis-element analyses for each castor bean SAP gene showed that no common cis-elements are shared across the nine castor bean SAP genes. Castor bean SAPs were localized to different regions of cells, including the cytoplasm, nucleus, and cytomembrane. This study provides a comprehensive profile of castor bean SAP genes that advances our understanding of their potential physiological and molecular functions in regulating growth and development and their responses to different abiotic stresses.

Genomic insights into the origin, domestication and genetic basis of agronomic traits of castor bean.

BACKGROUND: Castor bean (Ricinus communis L.) is an important oil crop, which belongs to the Euphorbiaceae family. The seed oil of castor bean is currently the only commercial source of ricinoleic acid that can be used for producing about 2000 industrial products. However, it remains largely unknown regarding the origin, domestication, and the genetic basis of key traits of castor bean. RESULTS: Here we perform a de novo chromosome-level genome assembly of the wild progenitor of castor bean. By resequencing and analyzing 505 worldwide accessions, we reveal that the accessions from East Africa are the extant wild progenitors of castor bean, and the domestication occurs ~ 3200 years ago. We demonstrate that significant genetic differentiation between wild populations in Kenya and Ethiopia is associated with past climate fluctuation in the Turkana depression ~ 7000 years ago. This dramatic change in climate may have caused the genetic bottleneck in wild castor bean populations. By a genome-wide association study, combined with quantitative trait locus analysis, we identify important candidate genes associated with plant architecture and seed size. CONCLUSIONS: This study provides novel insights of domestication and genome evolution of castor bean, which facilitates genomics-based breeding of this important oilseed crop and potentially other tree-like crops in future.

Bulked segregant analysis reveals candidate genes responsible for dwarf formation in woody oilseed crop castor bean.

Plant dwarfism is a desirable agronomic trait in non-timber trees, but little is known about the physiological and molecular mechanism underlying dwarfism in woody plants. Castor bean (Ricinus communis) is a typical woody oilseed crop. We performed cytological observations within xylem, phloem and cambia tissues, revealing that divergent cell growth in all tissues might play a role in the dwarf phenotype in cultivated castor bean. Based on bulked segregant analyses for a F2 population generated from the crossing of a tall and a dwarf accession, we identified two QTLs associated with plant height, covering 325 candidate genes. One of these, Rc5NG4-1 encoding a putative IAA transport protein localized in the tonoplast was functionally characterized. A non-synonymous SNP (altering the amino acid sequence from Y to C at position 218) differentiated the tall and dwarf plants and we confirmed, through heterologous yeast transformation, that the IAA uptake capacities of Rc5NG4-1Y and Rc5NG4-1C were significantly different. This study provides insights into the physiological and molecular mechanisms of dwarfing in woody non-timber economically important plants, with potential to aid in the genetic breeding of castor bean and other related crops.

Changes and Associations of Genomic Transcription and Histone Methylation with Salt Stress in Castor Bean.

Soil salinity is a major source of abiotic plant stress, adversely affecting plant growth, development and productivity. Although the physiological and molecular mechanisms that underlie plant responses to salt stress are becoming increasingly understood, epigenetic modifications, such as histone methylations and their potential regulation of the transcription of masked genes at the genome level in response to salt stress, remain largely unclear. Castor bean, an important nonedible oil crop, has evolved the capacity to grow under salt stress. Here, based on high-throughput RNA-seq and ChIP-seq data, we systematically investigated changes in genomic transcription and histone methylation using typical histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 tri-methylated lysine 27 (H3K27me3) markers in castor bean leaves subjected to salt stress. The results showed that gain or loss of histone methylation was closely associated with activated or repressed gene expression, though variations in both transcriptome and histone methylation modifications were relatively narrow in response to salt stress. Diverse salt responsive genes and switched histone methylation sites were identified in this study. In particular, we found for the first time that the transcription of the key salt-response regulator RADIALIS-LIKE SANT (RSM1), a MYB-related transcription factor involved in ABA(abscisic acid)-mediated salt stress signaling, was potentially regulated by bivalent H3K4me3-H3K27me3 modifications. Combining phenotypic variations with transcriptional and epigenetic changes, we provide a comprehensive profile for understanding histone modification, genomic transcription and their associations in response to salt stress in plants.

Application of a high-resolution genetic map for chromosome-scale genome assembly and fine QTLs mapping of seed size and weight traits in castor bean.

Castor bean (Ricinus communis L., Euphorbiaceae) is a critical biodiesel crop and its seed derivatives have important industrial applications. Due to lack of a high-density genetic map, the breeding and genetic improvement of castor bean has been largely restricted. In this study, based on a recombinant inbred line (RIL) population consisting of 200 individuals, we generated 8,896 high-quality genomic SNP markers and constructed a high-resolution genetic map with 10 linkage groups (LGs), spanning 1,852.33 centiMorgan (cM). Based on the genetic map, 996 scaffolds from the draft reference genome were anchored onto 10 pseudo-chromosomes, covering 84.43% of the castor bean genome. Furthermore, the quality of the pseudo-chromosome scale assembly genome was confirmed via genome collinearity analysis within the castor bean genome as well as between castor bean and cassava. Our results provide new evidence that the phylogenetic position of castor bean is relatively solitary from other taxa in the Euphorbiaceae family. Based on the genetic map, we identified 16 QTLs that control seed size and weight (covering 851 candidate genes). The findings will be helpful for further research into potential new mechanisms controlling seed size and weight in castor bean. The genetic map and improved pseudo-chromosome scale genome provide crucial foundations for marker-assisted selection (MAS) of QTL governing important agronomic traits, as well as the accelerated molecular breeding of castor bean in a cost-effective pattern.

Global Gene Expression of Seed Coat Tissues Reveals a Potential Mechanism of Regulating Seed Size Formation in Castor Bean.

The physiological and molecular basis of seed size formation is complex, and the development of seed coat (derived from integument cells) might be a critical factor that determines seed size formation for many endospermic seeds. Castor bean (Ricinus communis L.), a model system of studying seed biology, has large and persistent endosperm with a hard seed coat at maturity. Here, we investigated the potential molecular mechanisms underlying seed size formation in castor bean by comparing the difference between global gene expression within developing seed coat tissues between the large-seed ZB107 and small-seed ZB306. First, we observed the cell size of seed coat and concluded that the large seed coat area of ZB107 resulted from more cell numbers (rather than cell size). Furthermore, we found that the lignin proportion of seed coat was higher in ZB306. An investigation into global gene expression of developing seed coat tissues revealed that 815 genes were up-regulated and 813 were down-regulated in ZB306 relative to ZB107. Interestingly, we found that many genes involved in regulating cell division were up-regulated in ZB107, whereas many genes involved in regulating lignin biosynthesis (including several NAC members, as well as MYB46/83 and MYB58/63) and in mediating programmed cell death (such as CysEP1 and ßVPE) were up-regulated in ZB306. Furthermore, the expression patterns of the genes mentioned above indicated that the lignification of seed coat tissues was enhanced and occurred earlier in the developing seeds of ZB306. Taken together, we tentatively proposed a potential scenario for explaining the molecular mechanisms of seed coat governing seed size formation in castor bean by increasing the cell number and delaying the onset of lignification in seed coat tissues in large-seed ZB107. This study not only presents new information for possible modulation of seed coat related genes to improve castor seed yield, but also provides new insights into understanding the molecular basis of seed size formation in endospermic seeds with hard seed coat.

Multi-gene co-expression can improve comprehensive resistance to multiple abiotic stresses in Brassica napus L.

Rapeseed (Brassica napus L.) is an important oil crop worldwide. For current B. napus production, it is urgent to develop new varieties with higher seed productivity and increased stress tolerance for better adaptation to the abiotic stresses as a result of global climate change. Genetic engineering, to some extent, can overcome the limitations of genetic exchange in conventional breeding. Consequently, it considered as an effective method for improving modern crop breeding for B. napus. Since crop stress resistance is a polygenic complex trait, only by multi-gene synergistic effects can effectively achieve the comprehensive stress resistance of crops. Hence, in the present study, five stress resistance genes, NCED3, ABAR, CBF3, LOS5, and ICE1 were transferred into B. napus. Compared with wildtype (WT) plants, the multi-gene transformants K15 exhibited pronounced growth advantage under both normal growth and stress conditions. Additionally, K15 plants also showed significantly higher resistance response to multiple stresses at seed germination and seedling stages than WT plants. Furthermore, K15 plants had significantly higher leaf temperature and significantly lower stomatal aperture and water loss rate than WT plants, which indicated that the water-holding capacity of K15 plants was significantly superior to that of WT plants after stress treatment. In addition, K15 plants had significantly higher abscisic acid (ABA) content and significantly lower malondialdehyde (MDA) content than WT plants. In conclusion, the above results suggested that multi-gene co-expression could rapidly trigger plant stress resistance, reduce the stress injury on plants and synergistically improve the comprehensive resistance of B. napus.