Most fruits in our daily diet are the products of domestication and breeding. Here we report a map of genome variation for a major fruit that encompasses ~3.6 million variants, generated by deep resequencing of 115 cucumber lines sampled from 3,342 accessions worldwide. Comparative analysis suggests that fruit crops underwent narrower bottlenecks during domestication than grain crops. We identified 112 putative domestication sweeps; 1 of these regions contains a gene involved in the loss of bitterness in fruits, an essential domestication trait of cucumber. We also investigated the genomic basis of divergence among the cultivated populations and discovered a natural genetic variant in a β-carotene hydroxylase gene that could be used to breed cucumbers with enhanced nutritional value. The genomic history of cucumber evolution uncovered here provides the basis for future genomics-enabled breeding.
Targeted genome engineering (also known as genome editing) has emerged as an alternative to classical plant breeding and transgenic (GMO) methods to improve crop plants. Until recently, available tools for introducing site-specific double strand DNA breaks were restricted to zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENs). However, these technologies have not been widely adopted by the plant research community due to complicated design and laborious assembly of specific DNA binding proteins for each target gene. Recently, an easier method has emerged based on the bacterial type II CRISPR (clustered regularly interspaced short palindromic repeats)/Cas (CRISPR-associated) immune system. The CRISPR/Cas system allows targeted cleavage of genomic DNA guided by a customizable small noncoding RNA, resulting in gene modifications by both non-homologous end joining (NHEJ) and homology-directed repair (HDR) mechanisms. In this review we summarize and discuss recent applications of the CRISPR/Cas technology in plants.
Sustainable intensification of crop production is essential to ensure food demand is matched by supply as the human population continues to increase. This will require high-yielding crop varieties that can be grown sustainably with fewer inputs on less land. Both plant breeding and genetic modification (GM) methods make valuable contributions to varietal improvement, but targeted genome engineering promises to be critical to elevating future yields. Most such methods require targeting DNA breaks to defined locations followed by either nonhomologous end joining (NHEJ) or homologous recombination. Zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) can be engineered to create such breaks, but these systems require two different DNA binding proteins flanking a sequence of interest, each with a C-terminal FokI nuclease module. We report here that the bacterial clustered, regularly interspaced, short palindromic repeats (CRISPR) system, comprising a CRISPR-associated (Cas)9 protein and an engineered single guide RNA (sgRNA) that specifies a targeted nucleic acid sequence, is applicable to plants to induce mutations at defined loci.
Recent pathogenomic research on plant parasitic oomycete effector function and plant host responses has resulted in major conceptual advances in plant pathology, which has been possible thanks to the availability of genome sequences.
Few microorganisms match the impact that the oomycetes have had on mankind. This distinct lineage of eukaryotes is well-known for its most notorious member, Phytophthora infestans, the agent of the nineteenth century Irish potato famine, and several other devastating pathogens of cultivated and wild plants . Indeed, more than 60% of oomycete species infect plants . Less known is the fact that many oomycetes are parasitic on animals, from freshwater fish and crustaceans to mammals, such as livestock, pets, and humans . Animal parasitic oomycetes have received much less attention than their plant pathogenic kin, and our understanding of their virulence mechanisms is rudimentary. However, research momentum is poised to accelerate with the first report of the genome of an animal parasitic oomycete. In this issue of PLOS Genetics, Jiang et al.  describe the 63 Mbp genome sequence of the fish pathogen Saprolegnia parasitica and highlight a distinct repertoire of candidate virulence genes.
We now know what caused the Irish potato famine. Scientists have pinpointed the pathogen by using plant samples collected in the mid-19th century. Weekends on All Things Considered host Jacki Lyden talks about it with the study's co-author, Sophien Kamoun of the Sainsbury Lab in the United Kingdom.
Other interviews about the Yoshida et al. eLife paper:
Background - Wheat yellow (stripe) rust caused by Puccinia striiformis f. sp. tritici (PST) is one of the most devastating diseases of wheat worldwide. To design effective breeding strategies that maximize the potential for durable disease resistance it is important to understand the molecular basis of PST pathogenicity. In particular, the characterisation of the structure, function and evolutionary dynamics of secreted effector proteins that are detected by host immune receptors can help guide and prioritize breeding efforts. However, to date, our knowledge of the effector repertoire of cereal rust pathogens is limited.
Results - We re-sequenced genomes of four PST isolates from the US and UK to identify effector candidates and relate them to their distinct virulence profiles. First, we assessed SNP frequencies between all isolates, with heterokaryotic SNPs being over tenfold more frequent (5.29 +/- 2.23 SNPs/kb) than homokaryotic SNPs (0.41 +/- 0.28 SNPs/kb). Next, we implemented a bioinformatics pipeline to integrate genomics, transcriptomics, and effector-focused annotations to identify and classify effector candidates in PST. RNAseq analysis highlighted transcripts encoding secreted proteins that were significantly enriched in haustoria compared to infected tissue. The expression of 22 candidate effector genes was characterised using qRT-PCR, revealing distinct temporal expression patterns during infection in wheat. Lastly, we identified proteins that displayed non-synonymous substitutions specifically between the two UK isolates PST-87/7 and PST-08/21, which differ in virulence to two wheat varieties. By focusing on polymorphic variants enriched in haustoria, we identified five polymorphic effector candidates between PST-87/7 and PST-08/21 among 2,999 secreted proteins. These allelic variants are now a priority for functional validation as virulence/avirulence effectors in the corresponding wheat varieties.
Conclusions - Integration of genomics, transcriptomics, and effector-directed annotation of PST isolates has enabled us to move beyond the single isolate-directed catalogues of effector proteins and develop a framework for mining effector proteins in closely related isolates and relate these back to their defined virulence profiles. This should ultimately lead to more comprehensive understanding of the PST pathogenesis system, an important first step towards developing more effective surveillance and management strategies for one of the most devastating pathogens of wheat.
Genome sequences of plant fungal pathogens have enabled the identification of effectors that cooperatively modulate the cellular environment for successful fungal growth and suppress host defense. Identification and characterization of novel effector proteins are crucial to understand pathogen virulence and host plant defense mechanisms. Previous reports indicate that the Pseudomonas syringae pv. tomato DC3000 type III secretion system (T3SS) can be used to study how non-bacterial effectors manipulate dicot plant cell function using the Effector Detector Vector (pEDV) system. Here we report a pEDV-based effector delivery system in which the T3SS of Burkholderia glumae, an emerging rice pathogen, is used to translocate the AVR-Pik and AVR-Pii effectors of fungal pathogen Magnaporthe oryzae to rice cytoplasm. The translocated AVR-Pik and AVR-Pii showed avirulence activity when tested in rice cultivars containing the cognate R genes. AVR-Pik reduced and delayed the hypersensitive response triggered by B. glumae in the non-host plant Nicotiana benthamiana indicative of an immunosuppressive virulence activity. AVR proteins fused with fluorescent protein and nuclear localization signal were delivered by B. glumae T3SS and observed in the nuclei of infected cells in rice, wheat, barley and N. benthamiana. Our bacterial T3SS-enabled eukaryotic effector delivery and subcellular localization assays provide a useful method to identify and study effector functions in monocot plants.
The majority of agronomically important crop traits are quantitative, meaning that they are controlled by multiple genes each with a small effect (quantitative trait loci: QTL). QTL mapping and isolation is important for efficient crop breeding by marker-assisted selection (MAS) and for a better understanding of the molecular mechanisms underlying the traits. Since it requires the development and selection of DNA markers for linkage analysis, QTL analysis has been however time consuming and labor intensive. Here we report a rapid identification of plant QTL by whole genome resequencing of DNAs from two populations each composed of 20-50 individuals showing extreme opposite trait values for a given phenotype in a segregating progeny. We propose to name this approach QTL-seq as applied to plant species. We applied QTL-seq to rice recombinant inbred lines (RILs) and F2 populations and successfully identified QTL for important agronomic traits, such as partial resistance to the fungal rice blast disease and seedling vigor. Simulation study showed that QTL-seq is able to detect QTL over wide ranges of experimental variables, and the method can be generally applied in population genomics studies to rapidly identify genomic regions that underwent artificial or natural selective sweeps.
Welcome to Open Access Data and Crowdsourced analyses!
On this website you'll be able to get data to do your own analyses on ash and ash dieback. You can see the results of other peoples work as soon as it is available and share your own discoveries in the same way.
You will always get full credit for your work and in doing so contribute to a real community effort.
The symbiotic association between plants and arbuscular mycorrhizal fungi is almost ubiquitous within the plant kingdom , and the early stages of the association are controlled by plant-derived strigolactone acting as a signal to the fungus in the rhizosphere [2–4] and lipochito-oligosaccharides acting as fungal signals to the plant . Hyphopodia form at the root surface, allowing the initial invasion, and this is analogous to appressoria, infection structures of pathogenic fungi and oomycetes. Here, we characterize RAM2, a gene of Medicago truncatula required for colonization of the root by mycorrhizal fungi, which is necessary for appropriate hyphopodia and arbuscule formation. RAM2 encodes a glycerol-3-phosphate acyl transferase (GPAT) and is involved in the production of cutin monomers. Plants defective in RAM2 are unable to be colonized by arbuscular mycorrhizal fungi but also show defects in colonization by an oomycete pathogen, with the absence of appressoria formation. RAM2 defines a direct signaling function, because exogenous addition of the C16 aliphatic fatty acids associated with cutin are sufficient to promote hyphopodia/ appressoria formation. Thus, cutin monomers act as plant signals that promote colonization by arbuscular mycorrhizal fungi, and this signaling function has been recruited by pathogenic oomycetes to facilitate their own invasion.
Phytopathogenic oomycetes, such as Phytophthora infestans, secrete an arsenal of effector proteins that modulate plant innate immunity to enable infection. We describe CRN8, a host-translocated effector of P. infestans that has kinase activity in planta. CRN8 is a modular protein of the CRN effector family. The C-terminus of CRN8 localizes to the host nucleus and triggers cell death when the protein is expressed in planta. Cell death induction by CRN8 is dependent on its localization to the plant nucleus, which requires a functional nuclear localization signal (NLS). The C-terminal sequence of CRN8 has similarity to a serine/threonine RD kinase domain. We demonstrated that CRN8 is a functional RD kinase and that its auto-phosphorylation is dependent on an intact catalytic site. Co-immunoprecipitation experiments revealed that CRN8 forms a dimer or multimer. Heterologous expression of CRN8 in planta resulted in enhanced virulence by P. infestans. In contrast, in planta expression of the dominant-negative CRN8R469A;D470A resulted in reduced P. infestans infection, further implicating CRN8 in virulence. Overall, our results indicate that similar to animal parasites, plant pathogens also translocate biochemically active kinase effectors inside host cells.
Every four years, the Olympic Games plays host to competitors who have built on their natural talent by training for many years to become the best in their chosen discipline. Similar spirit and endeavour can be found throughout the microbial world, in which every day is a competition to survive and thrive. Microorganisms are trained through evolution to become the fittest and the best adapted to a particular environmental niche or lifestyle, and to innovate when the 'rules of the game' are changed by alterations to their natural habitats. In this Essay, we honour the best competitors in the microbial world by inviting them to take part in the inaugural Microbial Olympics.
Plant pathogens among the Microbial Olympics medalists!
Accurate identification of DNA polymorphisms using next-generation sequencing technology is challenging because of a high rate of sequencing error and incorrect mapping of reads to reference genomes. Currently available short read aligners and DNA variant callers suffer from these problems. We developed the Coval software to improve the quality of short read alignments. Coval is designed to minimize the incidence of spurious alignment of short reads, by filtering mismatched reads that remained in alignments after local realignment and error correction of mismatched reads. The error correction is executed based on the base quality and allele frequency at the non-reference positions for an individual or pooled sample. We demonstrated the utility of Coval by applying it to simulated genomes and experimentally obtained short-read data of rice, nematode, and mouse. Moreover, we found an unexpectedly large number of incorrectly mapped reads in ‘targeted’ alignments, where the whole genome sequencing reads had been aligned to a local genomic segment, and showed that Coval effectively eliminated such spurious alignments. We conclude that Coval significantly improves the quality of short-read sequence alignments, thereby increasing the calling accuracy of currently available tools for SNP and indel identification. Coval is available at http://sourceforge.net/projects/coval105/.
Puccinia monoica is a spectacular plant parasitic rust fungus that triggers the formation of flower-like structures (pseudoflowers) in its Brassicaceae host plant Boechera stricta. Pseudoflowers mimic in shape, color, nectar and scent co-occurring and unrelated flowers such as buttercups. They act to attract insects thereby aiding spore dispersal and sexual reproduction of the rust fungus. Although much ecological research has been performed on P.monoica-induced pseudoflowers, this system has yet to be investigated at the molecular or genomic level. To date, the molecular alterations underlying the development of pseudoflowers and the genes involved have not been described. To address this, we performed gene expression profiling to reveal 256 plant biological processes that are significantly altered in pseudoflowers. Among these biological processes, plant genes involved in cell fate specification, regulation of transcription, reproduction, floral organ development, anthocyanin (major floral pigments) and terpenoid biosynthesis (major floral volatile compounds) were down-regulated in pseudoflowers. In contrast, plant genes involved in shoot, cotyledon and leaf development, carbohydrate transport, wax biosynthesis, cutin transport and L-phenylalanine metabolism (pathway that results in phenylethanol and phenylacetaldehyde volatile production) were up-regulated. These findings point to an extensive reprogramming of host genes by the rust pathogen to induce floral mimicry. We also highlight 31 differentially regulated plant genes that are enriched in the biological processes mentioned above, and are potentially involved in the formation of pseudoflowers. This work illustrates the complex perturbations induced by rust pathogens in their host plants, and provides a starting point for understanding the molecular mechanisms of pathogen-induced floral mimicry.
Advances in genome sequencing technologies have enabled researchers and breeders to rapidly associate phenotypic variation to genome sequence differences. We recently took advantage of next-generation sequencing technology to develop MutMap, a method that allows rapid identification of causal nucleotide changes of rice mutants by whole genome resequencing of pooled DNA of mutant F2 progeny derived from crosses made between candidate mutants and the parental line. Here we describe MutMap+, a versatile extension of MutMap, that identifies causal mutations by comparing SNP frequencies of bulked DNA of mutant and wild-type progeny of M3 generation derived from selfing of an M2 heterozygous individual. Notably, MutMap+ does not necessitate artificial crossing between mutants and the wild-type parental line. This method is therefore suitable for identifying mutations that cause early development lethality, sterility, or generally hamper crossing. Furthermore, MutMap+ is potentially useful for gene isolation in crops that are recalcitrant to artificial crosses.
Next-generation sequencing allows the identification of mutations responsible for mutant phenotypes by whole-genome resequencing and alignment to a reference genome. However, when the resequenced cultivar/line displays significant structural variation from the reference genome, mutations in the genome regions missing from the reference (gaps) cannot be identified by simple alignment.Here we report on a method called ‘MutMap-Gap’, which involves delineating a candidate region harboring a mutation of interest using the recently reported MutMap method, followed by de novo assembly, alignment, and identification of the mutation within genome gaps.We applied MutMap-Gap to isolate the blast resistant gene Pii from the rice cv Hitomebore using mutant lines that have lost Pii function.MutMap-Gap should prove useful for cloning genes that exhibit significant structural variations such as disease resistance genes of the nucleotide-binding site-leucine rich repeat (NBS-LRR) class.
Plant nucleotide-binding leucine-rich repeat (NB-LRR) proteins serve as intracellular sensors to detect pathogen effectors and trigger immune responses. Transcription of the NB-LRR-encoding Resistance (R) genes needs to be tightly controlled to avoid inappropriate defense activation. How the expression of the NB-LRR R genes is regulated is poorly understood. The Arabidopsis snc1 mutant carries a gain-of-function mutation in a TIR-NB-LRR-encoding gene, resulting in the constitutive activation of plant defence responses. A snc1 suppressor screen identified modifier of snc1, 9 (mos9), which partially suppresses the autoimmune phenotypes of snc1. Positional cloning revealed that MOS9 encodes a plant-specific protein of unknown function. Expression analysis showed that MOS9 is required for the full expression of TIR-NB-LRR protein-encoding RPP4 and SNC1, both of which reside in the RPP4 cluster. Co-immunoprecipitation and mass spectrometry analyses revealed that MOS9 associates with the Set1 class H3K4 methyl transferase ATXR7. Like MOS9, ATXR7 is also required for the full expression of SNC1 and the autoimmune phenotypes in the snc1 mutant. In atxr7 mutant plants, the expression of RPP4 is similarly reduced and resistance against Hyaloperonospora arabidopsidis Emwa1 is compromised. Consistent with the attenuated expression of SNC1 and RPP4, H3K4me3 marks are reduced around the promoters of SNC1 and RPP4 in mos9 plants. Our data suggest that MOS9 functions together with ATXR7 to regulate the expression of SNC1 and RPP4 through H3K4 methylation, which plays an important role in fine-tuning their transcription levels and functions in plant defence.
Phytophthora infestans, the cause of potato late blight, is infamous for having triggered the Irish Great Famine in the 1840s. Until the late 1970s, P. infestans diversity outside of its Mexican center of origin was low, and one scenario held that a single strain, US-1, had dominated the global population for 150 years; this was later challenged based on DNA analysis of historical herbarium specimens. We have compared the genomes of 11 herbarium and 15 modern strains. We conclude that the nineteenth century epidemic was caused by a unique genotype, HERB-1, that persisted for over 50 years. HERB-1 is distinct from all examined modern strains, but it is a close relative of US-1, which replaced it outside of Mexico in the twentieth century. We propose that HERB-1 and US-1 emerged from a metapopulation that was established in the early 1800s outside of the species' center of diversity.
Ash dieback is a devastating fungal disease of ash trees that has swept across Europe and recently reached the UK. This emergent pathogen has received little study in the past and its effect threatens to overwhelm the ash populations. In response to this we have produced some initial genomics datasets and taken the unusual step of releasing them to the scientific for analysis without first performing our own. In this manner we hope to 'crowdsource' analyses and bring the expertise of the community to bear on this problem as quickly as possible. Our data has been released through our website at oadb.tsl.ac.uk and a public GitHub repository.
Chalara ash dieback was first confirmed in the natural environment in the UK in late autumn based on samples from Ashwellthorpe Wood near Norwich. We decided early on in this project that speed would be a critical driver given the emergency nature of the problem. We decided that we should generate genetic sequences as rapidly as possible, release them to the community, and prompt the crowdsourcing exercise we have been publicizing since Friday.
The normal procedure would be to culture the pathogen and sequence the genome and transcriptome from cultured material and controlled laboratory infections. Here, we decided to take the unusual step of directly sequencing the “interaction transcriptome” of a lesion dissected from an infected ash twig. This was the most rapid way to proceed to generate useful information without proceeding through standard laboratory culturing. This is the shortest route from the wood to the sequencer to the computer. The question that many of you must be asking is how useful is this data? This post addresses this question and summarizes the preliminary analyses that the TSL team has produced.
Every plant is closely associated with a variety of living organisms. Therefore, deciphering howplants interact with mutualistic and parasitic organisms is essential for a comprehensive understanding of the biology of plants. The field of plant–biotic interactions has recently coalesced around an integrated model. Major classes of molecular players both from plants and their associated organisms have been revealed. These include cell surface and intracellular immune receptors of plants, as well as apoplastic and host-cell translocated (cytoplasmic) effectors of the invading organism. This article focuses on effectors, molecules secreted by plant-associated organisms that alter plant processes. Effectors have emerged as a central class of molecules in our integrated view of plant–microbe interactions. Their study has significantly contributed to advancing our knowledge of plant hormones, plant development, plant receptors, and epigenetics. Many pathogen effectors are extraordinary examples of biological innovation; they include some of themost remarkable proteins knownto function inside plant cells. Here, we review some of the key concepts that have emerged from the study of the effectors of plant-associated organisms. In particular, we focus on how effectors function in plant tissues and discuss future perspectives in the field of effector biology.
Pest and pathogen losses jeopardise global food security and ever since the 19th century Irish famine, potato late blight has exemplified this threat. The causal oomycete pathogen, Phytophthora infestans, undergoes major population shifts in agricultural systems via the successive emergence and migration of asexual lineages. The phenotypic and genotypic bases of these selective sweeps are largely unknown but management strategies need to adapt to reflect the changing pathogen population. Here, we used molecular markers to document the emergence of a lineage, termed 13_A2, in the European P. infestans population, and its rapid displacement of other lineages to exceed 75% of the pathogen population across Great Britain in less than three years. We show that isolates of the 13_A2 lineage are among the most aggressive on cultivated potatoes, outcompete other aggressive lineages in the field, and overcome previously effective forms of plant host resistance. Genome analyses of a 13_A2 isolate revealed extensive genetic and expression polymorphisms particularly in effector genes. Copy number variations, gene gains and losses, amino-acid replacements and changes in expression patterns of disease effector genes within the 13_A2 isolate likely contribute to enhanced virulence and aggressiveness to drive this population displacement. Importantly, 13_A2 isolates carry intact and in planta induced Avrblb1, Avrblb2 and Avrvnt1 effector genes that trigger resistance in potato lines carrying the corresponding R immune receptor genes Rpi-blb1, Rpi-blb2, and Rpi-vnt1.1. These findings point towards a strategy for deploying genetic resistance to mitigate the impact of the 13_A2 lineage and illustrate how pathogen population monitoring, combined with genome analysis, informs the management of devastating disease epidemics.
Plant pathogens secrete effector proteins to modulate plant immunity and promote host colonization. Plant nucleotide binding leucine-rich repeat (NB-LRR) immunoreceptors recognize specific pathogen effectors directly or indirectly. Little is known about how NB-LRR proteins recognize effectors of filamentous plant pathogens, such as Phytophthora infestans. AVR2 belongs to a family of 13 sequence-divergent P. infestans RXLR effectors that are differentially recognized by members of the R2 NB-LRR family in Solanum demissum. We report that the putative plant phosphatase BSU-LIKE PROTEIN1 (BSL1) is required for R2-mediated perception of AVR2 and resistance to P. infestans. AVR2 associates with BSL1 and mediates the interaction of BSL1 with R2 in planta, possibly through the formation of a ternary complex. Strains of P. infestans that are virulent on R2 potatoes express an unrecognized form, Avr2-like (referred to as A2l). A2L can still interact with BSL1 but does not promote the association of BSL1 with R2. Our findings show that recognition of the P. infestans AVR2 effector by the NB-LRR protein R2 requires the putative phosphatase BSL1. This reveals that, similar to effectors of phytopathogenic bacteria, recognition of filamentous pathogen effectors can be mediated via a host protein that interacts with both the effector and the NB-LRR immunoreceptor.