A reference genome of Commelinales provides insights into the commelinids evolution and global spread of water hyacinth (Pontederia crassipes).

Commelinales belongs to the commelinids clade, which also comprises Poales that includes the most important monocot species, such as rice, wheat, and maize. No reference genome of Commelinales is currently available. Water hyacinth (Pontederia crassipes or Eichhornia crassipes), a member of Commelinales, is one of the devastating aquatic weeds, although it is also grown as an ornamental and medical plant. Here, we present a chromosome-scale reference genome of the tetraploid water hyacinth with a total length of 1.22 Gb (over 95% of the estimated size) across 8 pseudochromosome pairs. With the representative genomes, we reconstructed a phylogeny of the commelinids, which supported Zingiberales and Commelinales being sister lineages of Arecales and shed lights on the controversial relationship of the orders. We also reconstructed ancestral karyotypes of the commelinids clade and confirmed the ancient commelinids genome having 8 chromosomes but not 5 as previously reported. Gene family analysis revealed contraction of disease-resistance genes during polyploidization of water hyacinth, likely a result of fitness requirement for its role as a weed. Genetic diversity analysis using 9 water hyacinth lines from 3 continents (South America, Asia, and Europe) revealed very closely related nuclear genomes and almost identical chloroplast genomes of the materials, as well as provided clues about the global dispersal of water hyacinth. The genomic resources of P. crassipes reported here contribute a crucial missing link of the commelinids species and offer novel insights into their phylogeny.

MicroRNAs constitute an additional layer in plant response to simultaneous bio- and abiotic stresses as exemplified by UV-B radiation and flg22-treatment on Arabidopsis thaliana.

Plants are confronted with various environmental stresses and develop sophisticated adaptive mechanisms. Our previous work demonstrated that the crosstalk of flg22 and ultraviolet (UV)-B-induced signalling cascades reprograms the expression of flavonol pathway genes (FPGs), benefiting plant defence responses. Although several transcription factors have been identified to be involved in this crosstalk, the underlying mechanism is largely unclear. Here, we analyzed microRNAs (miRNAs) and identified 126, 129 and 113 miRNAs with altered abundances compared to untreated control in flg22-, UV-B- and flg22/UV-B-treated seedlings, respectively. Two distinct modules were identified: The first consists of 10 miRNAs repressed by UV-B but up-regulated by flg22, and the second with five miRNAs repressed by flg22 but up-regulated by UV-B. In Arabidopsis, the knockdown of miR858a, a representative of module I, increased the abundance of CHS (a marker gene for FPGs), whereas its overexpression reduced CHS. Conversely, knockout of miR164b from module II decreased CHS and its overexpression increased CHS transcript levels. These data suggest a decisive role of miRNAs in the crosstalk. In the next, we described the interaction between miR858a and its target MYB111 (a positive regulator of FPGs) from module I in detail. We showed that MYB111 was profoundly post-transcriptionally regulated by miR858a during the crosstalk, whose expression was specifically but antagonistically controlled by UVR8- and FLS2-mediated signallings. Moreover, transcriptional monitoring using the GUS reporter gene demonstrates that miRNA-mediated posttranscriptional regulation is the main driving force in reprogramming the expression of FPGs and regulates plant adaptation to multiple concurrent environmental stresses.

QTL mapping and transcriptome analysis identify novel QTLs and candidate genes in Brassica villosa for quantitative resistance against Sclerotinia sclerotiorum.

KEY MESSAGE: Novel QTLs and candidate genes for Sclerotinia-resistance were identified in B. villosa, a wild Brassica species, which represents a new genetic source for improving oilseed rape resistance to SSR. Sclerotinia stem rot (SSR), caused by Sclerotinia sclerotiorum, is one of the most destructive diseases in oilseed rape growing regions. To date, there is no effective genetic resistance against S. sclerotiorum in the B. napus germplasm and knowledge of the molecular plant-fungal interaction is also limited. To identify new resistance resources, we screened a set of wild Brassica species and identified B. villosa (BRA1896) with a high level of Sclerotinia-resistance. Two segregating F2 populations for Sclerotinia-resistance, generated by interspecific crosses between the resistant B. villosa (BRA1896) and the wild susceptible B. oleracea (BRA1909) were assessed for Sclerotinia-resistance. Genetic mapping using a 15-k Illumina Infinium SNP-array resulted in a high-density genetic map containing 1,118 SNP markers and spanning a total genetic length of 792.2 cM. QTL analysis revealed seven QTLs explaining 3.8% to 16.5% of phenotypic variance. Intriguingly, RNAseq-based transcriptome analysis identified genes and pathways specific to B. villosa, of which a cluster of five genes encoding putative receptor-like kinases (RLKs) and two pathogenesis-related (PR) proteins are co-localized within a QTL on chromosome C07. Furthermore, transcriptomic analysis revealed enhanced ethylene (ET)-activated signaling in the resistant B. villosa, which is associated with a stronger plant immune response, depressed cell death, and enhanced phytoalexin biosynthesis compared to the susceptible B. oleracea. Our data demonstrates that B. villosa represents a novel and unique genetic source for improving oilseed rape resistance against SSR.

RNAseq-Based Working Model for Transcriptional Regulation of Crosstalk between Simultaneous Abiotic UV-B and Biotic Stresses in Plants.

Plants adjust their secondary metabolism by altering the expression of corresponding genes to cope with both abiotic and biotic stresses. In the case of UV-B radiation, plants produce protective flavonoids; however, this reaction is impeded during pattern-triggered immunity (PTI) induced by pathogens. Pathogen attack can be mimicked by the application of microbial associated molecular patterns (e.g., flg22) to study crosstalk between PTI and UV-B-induced signaling pathways. Switching from Arabidopsis cell cultures to in planta studies, we analyzed whole transcriptome changes to gain a deeper insight into crosstalk regulation. We performed a comparative transcriptomic analysis by RNAseq with four distinct mRNA libraries and identified 10778, 13620, and 11294 genes, which were differentially expressed after flg22, UV-B, and stress co-treatment, respectively. Focusing on genes being either co-regulated with the UV-B inducible marker gene chalcone synthase CHS or the flg22 inducible marker gene FRK1 identified a large set of transcription factors from diverse families, such as MYB, WRKY, or NAC. These data provide a global view of transcriptomic reprogramming during this crosstalk and constitute a valuable dataset for further deciphering the underlying regulatory mechanism(s), which appear to be much more complex than previously anticipated. The possible involvement of MBW complexes in this context is discussed.

Population genomic analysis reveals domestication of cultivated rye from weedy rye.

Rye (Secale cereale) is an important crop with multiple uses and a valuable genetic resource for wheat breeding. However, due to its complex genome and outcrossing nature, the origin of cultivated rye remains elusive. The geneticist N.I. Vavilov proposed that cultivated rye had been domesticated from weedy rye, rather than directly from wild species like other crops. Unraveling the domestication history of rye will extend our understanding of crop evolution and upend our inherent understanding of agricultural weeds. To this end, in this study we generated the 8.5 Tb of whole-genome resequencing data from 116 worldwide accessions of wild, weedy, and cultivated rye, and demonstrated that cultivated rye was domesticated directly from weedy relatives with a similar but enhanced genomic selection by humans. We found that a repertoire of genes that experienced artificial selection is associated with important agronomic traits, including shattering, grain yield, and disease resistance. Furthermore, we identified a composite introgression in cultivated rye from the wild perennial Secale strictum and detected a 2-Mb introgressed fragment containing a candidate ammonium transporter gene with potential effect on the grain yield and plant growth of rye. Taken together, our findings unravel the domestication history of cultivated rye, suggest that interspecific introgression serves as one of the likely causes of obscure species taxonomy of the genus Secale, and provide an important resource for future rye and wheat breeding.

MicroRNA840 (MIR840) accelerates leaf senescence by targeting the overlapping 3'UTRs of PPR and WHIRLY3 in Arabidopsis thaliana.

MicroRNAs negatively regulate gene expression by promoting target mRNA cleavage and/or impairing its translation, thereby playing a crucial role in plant development and environmental stress responses. In Arabidopsis, the MIR840 gene is located within the overlapping 3'UTR of the PPR and WHIRLY3 (WHY3) genes, both being predicted targets of miR840* and miR840, the short maturation products of MIR840. Gain- and loss-of-function of MIR840 in Arabidopsis resulted in opposite senescence phenotypes. The highest expression levels of the MIR840 precursor transcript pre-miR840 were observed at senescence initiation, and pre-miR840 expression is significantly correlated with a reduction in PPR, but not WHY3, transcript levels. Although a reduction of transcript level of PPR, but not WHY3 transcript levels were not significantly affected by MIR840 overexpression, its protein levels were strongly reduced. Mutating the cleavage sites or replacing the target sequences abolishes the miR840*/miR840-mediated degradation of PPR transcripts and accumulation of WHY3 protein. In support for this, concurrent knockdown of both PPR and WHY3 in wild-type plants resulted in a senescence phenotype resembling that of the MIR840-overexpressing plant. This indicates that both PRR and WHY3 are targets in the MIR840-mediated senescence pathway. Moreover, single knockout mutants of PPR and WHY3 show a convergent upregulated subset of senescence-associated genes, which are also found among those induced by MIR840 overexpression. Our data provide evidence for a regulatory role of MIR840 in plant senescence.

Dual-Localized WHIRLY1 Affects Salicylic Acid Biosynthesis via Coordination of ISOCHORISMATE SYNTHASE1, PHENYLALANINE AMMONIA LYASE1, and S-ADENOSYL-L-METHIONINE-DEPENDENT METHYLTRANSFERASE1.

Salicylic acid (SA) influences developmental senescence and is spatiotemporally controlled by various mechanisms, including biosynthesis, transport, and conjugate formation. Altered localization of Arabidopsis WHIRLY1 (WHY1), a repressor of leaf natural senescence, in the nucleus or chloroplast causes a perturbation in SA homeostasis, resulting in adverse plant senescence phenotypes. WHY1 loss-of-function mutation resulted in SA peaking 5 d earlier compared to wild-type plants, which accumulated SA at 42 d after germination. SA accumulation coincided with an early leaf-senescence phenotype, which could be prevented by ectopic expression of the nuclear WHY1 isoform (nWHY1). However, expressing the plastid WHY1 isoform (pWHY1) greatly enhanced cellular SA levels. Transcriptome analysis in the WHY1 loss-of-function mutant background following expression of either pWHY1 or nWHY1 indicated that hormone metabolism-related genes were most significantly altered. The pWHY1 isoform predominantly affected stress-related gene expression, whereas nWHY1 primarily controlled developmental gene expression. Chromatin immunoprecipitation-quantitative PCR assays indicated that nWHY1 directly binds to the promoter region of isochorismate synthase1 (ICS1), thus activating its expression at later developmental stages, but that it indirectly activates S-adenosyl- l -Met-dependent methyltransferase1 (BSMT1) expression via ethylene response factor 109 (ERF109). Moreover, nWHY1 repressed expression of Phe ammonia lyase-encoding gene (PAL1) via R2R3-MYB member 15 (MYB15) during the early stages of development. Interestingly, rising SA levels exerted a feedback effect by inducing nWHY1 modification and pWHY1 accumulation. Thus, the alteration of WHY1 organelle isoforms and the feedback of SA are involved in a circularly integrated regulatory network during developmental or stress-induced senescence in Arabidopsis.

Applications of CRISPR/Cas to Improve Crop Disease Resistance: Beyond Inactivation of Susceptibility Factors.

Current crop production systems are prone to increasing pathogen pressure. Fundamental understanding of molecular plant-pathogen interactions, the availability of crop and pathogen genomic information, as well as emerging genome editing permits a novel approach for breeding of crop disease resistance. We describe here strategies to identify new targets for resistance breeding with focus on interruption of the compatible plant-pathogen interaction by CRISPR/Cas-mediated genome editing. Basically, crop genome editing can be applied in several ways to achieve this goal. The most common approach focuses on the "simple" knockout by non-homologous end joining repair of plant susceptibility factors required for efficient host colonization. However, genome re-writing via homology-directed repair or base editing can also prevent host manipulation by changing the targets of pathogen-derived effectors or molecules beyond recognition, which also decreases plant susceptibility. We conclude that genome editing by CRISPR/Cas will become increasingly indispensable to generate in relatively short time beneficial resistance traits in crops to meet upcoming challenges.

Phytohormone crosstalk in the host-Verticillium interaction.

Functional genomics can be applied to shed light on the Brassica napus - Verticillium interaction. RNAseq data indicated already that abscisic acid (ABA) is apparently involved in the early oilseed rape response to fungal infection with Verticillium longisporum isolate 43 (Vl43). A calreticulin (CRT1a) was identified as novel susceptibility factor for Vl43 infecting both Arabidopsis and oilseed rape. CRT1a is involved in calcium homeostasis and contributes in the endoplasmatic reticulum to the unfolded protein response. The latter function could either affect the correct folding of other susceptibility factors or of negative regulators in ethylene (ET) signaling. Which CRT1a function is affected in the mutants is currently unknown, but both hypotheses can explain that crt1a loss-of-function mutants display increased resistance to V. longisporum and enhanced expression of ethylene signaling related genes. This indicates that besides other phytohormones such as ABA or salicylic acid (SA) also ET plays a critical role in the plant-Verticillium interaction, which might be exploited to improve plant resistance.

Loss of function of CRT1a (calreticulin) reduces plant susceptibility to Verticillium longisporum in both Arabidopsis thaliana and oilseed rape (Brassica napus).

Brassica napus is highly susceptible towards Verticillium longisporum (Vl43) with no effective genetic resistance. It is believed that the fungus reprogrammes plant physiological processes by up-regulation of so-called susceptibility factors to establish a compatible interaction. By transcriptome analysis, we identified genes, which were activated/up-regulated in rapeseed after Vl43 infection. To test whether one of these genes is functionally involved in the infection process and loss of function would lead to decreased susceptibility, we firstly challenged KO lines of corresponding Arabidopsis orthologs with Vl43 and compared them with wild-type plants. Here, we report that the KO of AtCRT1a results in drastically reduced susceptibility of plants to Vl43. To prove crt1a mutation also decreases susceptibility in B. napus, we identified 10 mutations in a TILLING population. Three T3 mutants displayed increased resistance as compared to the wild type. To validate the results, we generated CRISPR/Cas-induced BnCRT1a mutants, challenged T2 plants with Vl43 and observed an overall reduced susceptibility in 3 out of 4 independent lines. Genotyping by allele-specific sequencing suggests a major effect of mutations in the CRT1a A-genome copy, while the C-genome copy appears to have no significant impact on plant susceptibility when challenged with Vl43. As revealed by transcript analysis, the loss of function of CRT1a results in activation of the ethylene signalling pathway, which may contribute to reduced susceptibility. Furthermore, this study demonstrates a novel strategy with great potential to improve plant disease resistance.

Suppression of abscisic acid biosynthesis at the early infection stage of Verticillium longisporum in oilseed rape (Brassica napus).

Verticillium longisporum infects oilseed rape (Brassica napus) and Arabidopsis thaliana. To investigate the early response of oilseed rape to the fungal infection, we determined transcriptomic changes in oilseed rape roots at 6 days post-inoculation (dpi) by RNA-Seq analysis, in which non-infected roots served as a control. Strikingly, a subset of genes involved in abscisic acid (ABA) biosynthesis was found to be down-regulated and the ABA level was accordingly attenuated in 6 dpi oilseed rape as compared with the control. Gene expression analysis revealed that this was mainly attributed to the suppression of BnNCED3-mediated ABA biosynthesis, involving, for example, BnWRKY57. However, this down-regulation of ABA biosynthesis could not be observed in infected Arabidopsis roots. Arabidopsis ABA- defective mutants nced3 and aao3 displayed pronounced tolerance to the fungal infection with delayed and impeded symptom development, even though fungal colonization was not affected in both mutants. These data suggest that ABA appears to be required for full susceptibility of Arabidopsis to the fungal infection. Furthermore, we found that in both 6 dpi oilseed rape and the Arabidopsis nced3 mutant, the salicylic acid (SA) signalling pathway was induced while the jasmonic acid (JA)/ethylene (ET) signalling pathway was concomitantly mitigated. Following these data, we conclude that in oilseed rape the V. longisporum infection triggers a host-specific suppression of the NCED3-mediated ABA biosynthesis, consequently increasing plant tolerance to the fungal infection. We believe that this might be part of the virulence strategy of V. longisporum to initiate/establish a long-lasting compatible interaction with oilseed rape (coexistence), which appears to be different from the infection process in Arabidopsis.

BvcZR3 and BvHs1pro-1 Genes Pyramiding Enhanced Beet Cyst Nematode (Heterodera schachtii Schm.) Resistance in Oilseed Rape (Brassica napus L.).

Beet cyst nematode (Heterodera schachtii Schm.) is one of the most damaging pests in sugar beet growing areas around the world. The Hs1pro-1 and cZR3 genes confer resistance to the beet cyst nematode, and both were cloned from sugar beet translocation line (A906001). The translocation line carried the locus from B. procumbens chromosome 1 including Hs1pro-1 gene and resistance gene analogs (RGA), which confer resistance to Heterodera schachtii. In this research, BvHs1pro-1 and BvcZR3 genes were transferred into oilseed rape to obtain different transgenic lines by A. tumefaciens mediated transformation method. The cZR3Hs1pro-1 gene was pyramided into the same plants by crossing homozygous cZR3 and Hs1pro-1 plants to identify the function and interaction of cZR3 and Hs1pro-1 genes. In vitro and in vivo cyst nematode resistance tests showed that cZR3 and Hs1pro-1 plants could be infested by beet cyst nematode (BCN) juveniles, however a large fraction of penetrated nematode juveniles was not able to develop normally and stagnated in roots of transgenic plants, consequently resulting in a significant reduction in the number of developed nematode females. A higher efficiency in inhibition of nematode females was observed in plants expressing pyramiding genes than in those only expressing a single gene. Molecular analysis demonstrated that BvHs1pro-1 and BvcZR3 gene expressions in oilseed rape constitutively activated transcription of plant-defense related genes such as NPR1 (non-expresser of PR1), SGT1b (enhanced disease resistance 1) and RAR1 (suppressor of the G2 allele of skp1). Transcript of NPR1 gene in transgenic cZR3 and Hs1pro-1 plants were slightly up-regulated, while its expression was considerably enhanced in cZR3Hs1pro-1 gene pyramiding plants. The expression of EDS1 gene did not change significantly among transgenic cZR3, Hs1pro-1 and cZR3Hs1pro-1 gene pyramiding plants and wild type. The expression of SGT1b gene was slightly up-regulated in transgenic cZR3 and Hs1pro-1 plants compared with the wild type, however, its expression was not changed in cZR3Hs1pro-1 gene pyramiding plant and had no interaction effect. RAR1 gene expression was significantly up-regulated in transgenic cZR3 and cZR3Hs1pro-1 genes pyramiding plants, but almost no expression was found in Hs1pro-1 transgenic plants. These results show that nematode resistance genes from sugar beet were functional in oilseed rape and conferred BCN resistance by activation of a CC-NBS-LRR R gene mediated resistance response. The gene pyramiding had enhanced resistance, thus offering a novel approach for the BCN control by preventing the propagation of BCN in oilseed rape. The transgenic oilseed rape could be used as a trap crop to offer an alternative method for beet cyst nematode control.

Suppression of UV-B stress induced flavonoids by biotic stress: Is there reciprocal crosstalk?

Plants respond to abiotic UV-B stress with enhanced expression of genes for flavonoid production, especially the key-enzyme chalcone synthase (CHS). Some flavonoids are antioxidative, antimicrobial and/or UV-B protective secondary metabolites. However, when plants are challenged with concomitant biotic stress (simulated e.g. by the bacterial peptide flg22, which induces MAMP triggered immunity, MTI), the production of flavonoids is strongly suppressed in both Arabidopsis thaliana cell cultures and plants. On the other hand, flg22 induces the production of defense related compounds, such as the phytoalexin scopoletin, as well as lignin, a structural barrier thought to restrict pathogen spread within the host tissue. Since all these metabolites require the precursor phenylalanine for their production, suppression of the flavonoid production appears to allow the plant to focus its secondary metabolism on the production of pathogen defense related compounds during MTI. Interestingly, several flavonoids have been reported to display anti-microbial activities. For example, the plant flavonoid phloretin targets the Pseudomonas syringae virulence factors flagella and type 3 secretion system. That is, suppression of flavonoid synthesis during MTI might have also negative side-effects on the pathogen defense. To clarify this issue, we deployed an Arabidopsis flavonoid mutant and obtained genetic evidence that flavonoids indeed contribute to ward off the virulent bacterial pathogen Pseudomonas syringae pv. tomato (Pst) DC3000. Finally, we show that UV-B attenuates expression of the flg22 receptor FLS2, indicating that there is negative and reciprocal interaction between this abiotic stress and the plant-pathogen defense responses.

Echinochloa crus-galli genome analysis provides insight into its adaptation and invasiveness as a weed.

Barnyardgrass (Echinochloa crus-galli) is a pernicious weed in agricultural fields worldwide. The molecular mechanisms underlying its success in the absence of human intervention are presently unknown. Here we report a draft genome sequence of the hexaploid species E. crus-galli, i.e., a 1.27 Gb assembly representing 90.7% of the predicted genome size. An extremely large repertoire of genes encoding cytochrome P450 monooxygenases and glutathione S-transferases associated with detoxification are found. Two gene clusters involved in the biosynthesis of an allelochemical 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) and a phytoalexin momilactone A are found in the E. crus-galli genome, respectively. The allelochemical DIMBOA gene cluster is activated in response to co-cultivation with rice, while the phytoalexin momilactone A gene cluster specifically to infection by pathogenic Pyricularia oryzae. Our results provide a new understanding of the molecular mechanisms underlying the extreme adaptation of the weed.

Analysis of transcriptional and epigenetic changes in hybrid vigor of allopolyploid Brassica napus uncovers key roles for small RNAs.

Heterosis is a fundamental biological phenomenon characterized by the superior performance of a hybrid compared with its parents. The underlying molecular basis for heterosis, particularly for allopolyploids, remains elusive. In this study we analyzed the transcriptomes of Brassica napus parental lines and their F1 hybrids at three stages of early flower development. Phenotypically, the F1 hybrids show remarkable heterosis in silique number and grain yield. Transcriptome analysis revealed that various phytohormone (auxin and salicylic acid) response genes are significantly altered in the F1 hybrids relative to the parental lines. We also found evidence for decreased expression divergence of the homoeologous gene pairs in the allopolyploid F1 hybrids and suggest that high-parental expression-level dominance plays an important role in heterosis. Small RNA and methylation studies aimed at examining the epigenetic effect of the changes in gene expression level in the F1 hybrids showed that the majority of the small interfering RNA (siRNA) clusters had a higher expression level in the F1 hybrids than in the parents, and that there was an increase in genome-wide DNA methylation in the F1 hybrid. Transposable elements associated with siRNA clusters had a higher level of methylation and a lower expression level in the F1 hybrid, implying that the non-additively expressed siRNA clusters resulted in lower activity of the transposable elements through DNA methylation in the hybrid. Our data provide insights into the role that changes in gene expression pattern and epigenetic mechanisms contribute to heterosis during early flower development in allopolyploid B. napus.

Investigation of the crosstalk between the flg22 and the UV-B-induced flavonol pathway in Arabidopsis thaliana seedlings.

In Arabidopsis cell culture, we have shown that flavonol metabolite accumulation depends on expression of the flavonol pathway genes (FPGs), which are up-regulated by UV-B irradiation but repressed during microbe-associated molecular pattern (MAMP) -triggered immunity (MTI) induced by the bacterial elicitor flg22. The suppression of flavonoid production during MTI is believed to allow the plant focusing its metabolism on the pathogen defense by directing phenylalanine resources from UV-B protective flavonol production towards production of phytoalexins and cell wall fortification by lignin incorporation. Here, we show that UV-B-induced FPGs are also suppressed by flg22 in Arabidopsis seedlings, demonstrating that this kind of 'crosstalk' is fully functional in planta. We believe that this system based on seedlings of the model plant Arabidopsis thaliana constitutes a valuable tool for further dissection of the underlying molecular mechanism, for example, by deploying gain/loss-of-function mutants. Furthermore, we observed some differences in the expression patterns of MYB transcription factors (TFs) as compared to data from the cell culture system. The new data suggest that in planta the TF MYB111 might play a more dominant role than the TF MYB12, which was strongly regulated in cell cultures. Thus, we can present an updated working model how this crosstalk might function.

Identification, evolution, and expression partitioning of miRNAs in allopolyploid Brassica napus.

The recently published genome of Brassica napus offers for the first time the opportunity to gain insights into the genomic organization and the evolution of miRNAs in oilseed rape. In this study, 12 small RNA libraries from two B. napus cultivars (Tapidor and Ningyou7) and their four double-haploid lines were sequenced, employing the newly sequenced B. napus genome, together with genomes of its progenitors Brassica rapa and Brassica oleracea. A total of 645 miRNAs including 280 conserved and 365 novel miRNAs were identified. Comparative analysis revealed a high level of genomic conservation of MIRNAs (75.9%) between the subgenomes of B. napus and its two progenitors' genomes, and MIRNA lost/gain events (133) occurred in B. napus after its speciation. Furthermore, significant partitioning of miRNA expressions between the two subgenomes in B. napus was detected. The data of degradome sequencing, miRNA-mediated cleavage, and expression analyses support specific interactions between miRNAs and their targets in the modulation of diverse physiological processes in roots and leaves, as well as in biosynthesis of, for example, glucosinolates and lipids in oilseed rape. These data provide a first genome-wide view on the origin, evolution, and genomic organization of B. napus MIRNAs.

A host plant genome (Zizania latifolia) after a century-long endophyte infection.

Despite the importance of host-microbe interactions in natural ecosystems, agriculture and medicine, the impact of long-term (especially decades or longer) microbial colonization on the dynamics of host genomes is not well understood. The vegetable crop 'Jiaobai' with enlarged edible stems was domesticated from wild Zizania latifolia (Oryzeae) approximately 2000 years ago as a result of persistent infection by a fungal endophyte, Ustilago esculenta. Asexual propagation via infected rhizomes is the only means of Jiaobai production, and the Z. latifolia-endophyte complex has been maintained continuously for two centuries. Here, genomic analysis revealed that cultivated Z. latifolia has a significantly smaller repertoire of immune receptors compared with wild Z. latifolia. There are widespread gene losses/mutations and expression changes in the plant-pathogen interaction pathway in Jiaobai. These results show that continuous long-standing endophyte association can have a major effect on the evolution of the structural and transcriptomic components of the host genome.

Identification and characterization of microRNAs in oilseed rape (Brassica napus) responsive to infection with the pathogenic fungus Verticillium longisporum using Brassica AA (Brassica rapa) and CC (Brassica oleracea) as reference genomes.

Verticillium longisporum, a soil-borne pathogenic fungus, causes vascular disease in oilseed rape (Brassica napus). We proposed that plant microRNAs (miRNAs) are involved in the plant-V. longisporum interaction. To identify oilseed rape miRNAs, we deep-sequenced two small RNA libraries made from V. longisporum infected/noninfected roots and employed Brassica rapa and Brassica oleracea genomes as references for miRNA prediction and characterization. We identified 893 B. napus miRNAs representing 360 conserved and 533 novel miRNAs, and mapped 429 and 464 miRNAs to the AA and CC genomes, respectively. Microsynteny analysis with the conserved miRNAs and their flanking protein coding sequences revealed 137 AA-CC genome syntenic miRNA pairs and 61 AA and 42 CC genome-unique miRNAs. Sixty-two miRNAs were responsive to the V. longisporum infection. We present data for specific interactions and simultaneously reciprocal changes in the expression levels of the miRNAs and their targets in the infected roots. We demonstrate that miRNAs are involved in the plant-fungus interaction and that miRNA168-Argonaute 1 (AGO1) expression modulation might act as a key regulatory module in a compatible plant-V. longisporum interaction. Our results suggest that V. longisporum may have evolved a virulence mechanism by interference with plant miRNAs to reprogram plant gene expression and achieve infection.

The interplay of transcription factors in suppression of UV-B induced flavonol accumulation by flg22.

Biotic stress can be mimicked by application of elicitors, which comprise of microbe-associated molecular patterns (MAMPs). Treatment of plant cell cultures with MAMPs such as flg22 suppressed the expression of UV-B-induced flavonol pathway genes (FPGs) in parsley, carrot and Arabidopsis. This is thought to allow the plant focusing its secondary metabolism on the pathogen defense during MAMP-triggered immunity (MTI). Recently we reported that this suppression also depends on prevention of histone 3 acetylation at lysine 9 (H3K9ac), a hallmark for gene activation. Here we describe a possible regulation between UV-B and flg22 signaling cascades, and the interplay of MYB and WRKY transcription factors in regulating the expression of the FPGs.