Hop (Humulus lupulus L.) is an emblematic industrial crop in the French North East region that developed at the same time as the brewing activity. Presently, this sector, especially microbreweries, are interested in endemic wild hops, which give beer production a local signature. In this study, we investigated the genetic and metabolic diversity of thirty-six wild hops sampled in various ecological environments. These wild accessions were propagated aeroponically and cultivated under uniform conditions (the same soil and the same environmental factors). Our phytochemical approach based on UHPLC-ESI-MS/MS analysis led to the identification of three metabolic clusters based on leaf content and characterized by variations in the contents of twelve specialized metabolites that were identified (including xanthohumol, bitter acids, and their oxidized derivatives). Furthermore, molecular characterization was carried out using sixteen EST-SSR microsatellites, allowing a genetic affiliation of our wild hops with hop varieties cultivated worldwide and wild hops genotyped to date using this method. Genetic proximity was observed for both European wild and hop varieties, especially for Strisselspalt, the historical variety of our region. Finally, our findings collectively assessed the impact of the hop genotype on the chemical phenotype through multivariate regression tree (MRT) analysis. Our results highlighted the 'WRKY 224' allele as a key discriminator between high- and low-metabolite producers. Moreover, the model based on genetic information explained 40% of the variance in the metabolic data. However, despite this strong association, the model lacked predictive power, suggesting that its applicability may be confined to the datasets analyzed.

• MeSH
• fenotyp MeSH
• genetická variace * MeSH
• genotyp MeSH
• Humulus * genetika   metabolismus   klasifikace   chemie   MeSH
• listy rostlin genetika   metabolismus   chemie   MeSH
• mikrosatelitní repetice genetika   MeSH
• tandemová hmotnostní spektrometrie MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Hop (Humulus lupulus L.) bitter acids are valuable metabolites for the brewing industry. They are biosynthesized and accumulate in glandular trichomes of the female inflorescence (hop cone). The content of alpha bitter acids, such as humulones, in hop cones can differentiate aromatic from bitter hop cultivars. These contents are subject to genetic and environmental control but significantly correlate with the number and size of glandular trichomes (lupulin glands). RESULTS: We evaluated the expression levels of 37 genes involved in bitter acid biosynthesis and morphological and developmental differentiation of glandular trichomes to identify key regulatory factors involved in bitter acid content differences. For bitter acid biosynthesis genes, upregulation of humulone synthase genes, which are important for the biosynthesis of alpha bitter acids in lupulin glands, could explain the higher accumulation of alpha bitter acids in bitter hops. Several transcription factors, including HlETC1, HlMYB61 and HlMYB5 from the MYB family, as well as HlGLABRA2, HlCYCB2-4, HlZFP8 and HlYABBY1, were also more highly expressed in the bitter hop cultivars; therefore, these factors may be important for the higher density of lupulin glands also seen in the bitter hop cultivars. CONCLUSIONS: Gene expression analyses enabled us to investigate the differences between aromatic and bitter hops. This study confirmed that the bitter acid content in glandular trichomes (lupulin glands) is dependent on the last step of alpha bitter acid biosynthesis and glandular trichome density.

• Klíčová slova
• Bitter acids, Differential gene expression, Glandular trichome development, Hop, Humulus lupulus, Lupulin gland,
• MeSH
• Humulus metabolismus   MeSH
• transkripční faktory metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• transkripční faktory MeSH

Viroids are small infectious pathogens, composed of a short single-stranded circular RNA. Hop (Humulus lupulus L.) plants are hosts to four viroids from the family Pospiviroidae. Hop latent viroid (HLVd) is spread worldwide in all hop-growing regions without any visible symptoms on infected hop plants. In this study, we evaluated the influence of HLVd infection on the content and the composition of secondary metabolites in maturated hop cones, together with gene expression analyses of involved biosynthesis and regulation genes for Saaz, Sládek, Premiant and Agnus cultivars. We confirmed that the contents of alpha bitter acids were significantly reduced in the range from 8.8% to 34% by viroid infection. New, we found that viroid infection significantly reduced the contents of xanthohumol in the range from 3.9% to 23.5%. In essential oils of Saaz cultivar, the contents of monoterpenes, terpene epoxides and terpene alcohols were increased, but the contents of sesquiterpenes and terpene ketones were decreased. Secondary metabolites changes were supported by gene expression analyses, except essential oils. Last-step biosynthesis enzyme genes, namely humulone synthase 1 (HS1) and 2 (HS2) for alpha bitter acids and O-methytransferase 1 (OMT1) for xanthohumol, were down-regulated by viroid infection. We found that the expression of ribosomal protein L5 (RPL5) RPL5 and the splicing of transcription factor IIIA-7ZF were affected by viroid infection and a disbalance in proteosynthesis can influence transcriptions of biosynthesis and regulatory genes involved in of secondary metabolites biosynthesis. We suppose that RPL5/TFIIIA-7ZF regulatory cascade can be involved in HLVd replication as for other viroids of the family Pospiviroidae.

• Klíčová slova
• HLVd, Humulus lupulus, bitter acids content, differential gene expression, essential oils, hop, hop latent viroid, xanthohumol,
• Publikační typ
• časopisecké články MeSH

The hop plant (Humulus lupulus L.) produces several valuable secondary metabolites, such as prenylflavonoid, bitter acids, and essential oils. These compounds are biosynthesized in glandular trichomes (lupulin glands) endowed with pharmacological properties and widely implicated in the beer brewing industry. The present study is an attempt to generate exhaustive information of transcriptome dynamics and gene regulatory mechanisms involved in biosynthesis and regulation of these compounds, developmental changes including trichome development at three development stages, namely leaf, bract, and mature lupulin glands. Using high-throughput RNA-Seq technology, a total of 61.13, 50.01, and 20.18 Mb clean reads in the leaf, bract, and lupulin gland libraries, respectively, were obtained and assembled into 43,550 unigenes. The putative functions were assigned to 30,996 transcripts (71.17%) based on basic local alignment search tool similarity searches against public sequence databases, including GO, KEGG, NR, and COG families, which indicated that genes are principally involved in fundamental cellular and molecular functions, and biosynthesis of secondary metabolites. The expression levels of all unigenes were analyzed in leaf, bract, and lupulin glands tissues of hop. The expression profile of transcript encoding enzymes of BCAA metabolism, MEP, and shikimate pathway was most up-regulated in lupulin glands compared with leaves and bracts. Similarly, the expression levels of the transcription factors and structural genes that directly encode enzymes involved in xanthohumol, bitter acids, and terpenoids biosynthesis pathway were found to be significantly enhanced in lupulin glands, suggesting that production of these metabolites increases after the leaf development. In addition, numerous genes involved in primary metabolism, lipid metabolism, photosynthesis, generation of precursor metabolites/energy, protein modification, transporter activity, and cell wall component biogenesis were differentially regulated in three developmental stages, suggesting their involvement in the dynamics of the lupulin gland development. The identification of differentially regulated trichome-related genes provided a new foundation for molecular research on trichome development and differentiation in hop. In conclusion, the reported results provide directions for future functional genomics studies for genetic engineering or molecular breeding for augmentation of secondary metabolite content in hop.

• Klíčová slova
• Humulus lupulus, RNA sequencing, bitter acids, lupulin glands, prenylflavonoids, terpenoids, trichome,
• MeSH
• flavonoidy biosyntéza   chemie   metabolismus   MeSH
• genová ontologie MeSH
• Humulus chemie   metabolismus   MeSH
• listy rostlin genetika   metabolismus   MeSH
• propiofenony chemie   metabolismus   MeSH
• regulace genové exprese u rostlin MeSH
• rostlinné proteiny genetika   metabolismus   MeSH
• sekvenování transkriptomu MeSH
• terpeny chemie   metabolismus   MeSH
• transkripční faktory metabolismus   MeSH
• transkriptom genetika   MeSH
• trichomy genetika   metabolismus   ultrastruktura   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• flavonoidy MeSH
• propiofenony MeSH
• rostlinné proteiny MeSH
• terpeny MeSH
• transkripční faktory MeSH
• xanthohumol MeSH Prohlížeč

Viroids are small non-capsidated, single-stranded, covalently-closed circular noncoding RNA replicons of 239-401 nucleotides that exploit host factors for their replication, and some cause disease in several economically important crop plants, while others appear to be benign. The proposed mechanisms of viroid pathogenesis include direct interaction of the genomic viroid RNA with host factors and post-transcriptional or transcriptional gene silencing via viroid-derived small RNAs (vd-sRNAs) generated by the host defensive machinery. Humulus lupulus (hop) plants are hosts to several viroids among which Hop latent viroid (HLVd) and Citrus bark cracking viroid (CBCVd) are attractive model systems for the study of viroid-host interactions due to the symptomless infection of the former and severe symptoms induced by the latter in this indicator host. To better understand their interactions with hop plant, a comparative transcriptomic analysis based on RNA sequencing (RNA-seq) was performed to reveal the transcriptional alterations induced as a result of single HLVd and CBCVd infection in hop. Additionally, the effect of HLVd on the aggressiveness of CBCVd that underlies severe stunting in hop in a mixed infection was studied by transcriptomic analysis. Our analysis revealed that CBCVd infection resulted in dynamic changes in the activity of genes as compared to single HLVd infection and their mixed infection. The differentially expressed genes that are involved in defense, phytohormone signaling, photosynthesis and chloroplasts, RNA regulation, processing and binding; protein metabolism and modification; and other mechanisms were more modulated in the CBCVd infection of hop. Nevertheless, Gene Ontology (GO) classification and pathway enrichment analysis showed that the expression of genes involved in the proteolysis mechanism is more active in a mixed infection as compared to a single one, suggesting co-infecting viroids may result in interference with host factors more prominently. Collectively, our results provide a deep transcriptome of hop and insight into complex single HLVd, CBCVd, and their coinfection in hop-plant interactions.

• Klíčová slova
• Citrus bark cracking viroid, Hop latent viroid, Humulus lupulus, co-infection, differentially expressed genes, transcriptome profiling,
• MeSH
• Humulus genetika   virologie   MeSH
• nemoci rostlin genetika   virologie   MeSH
• transkriptom * MeSH
• viroidy patogenita   MeSH
• Publikační typ
• časopisecké články MeSH

Viroids are smallest known pathogen that consist of non-capsidated, single-stranded non-coding RNA replicons and they exploits host factors for their replication and propagation. The severe stunting disease caused by Citrus bark cracking viroid (CBCVd) is a serious threat, which spreads rapidly within hop gardens. In this study, we employed comprehensive transcriptome analyses to dissect host-viroid interactions and identify gene expression changes that are associated with disease development in hop. Our analysis revealed that CBCVd-infection resulted in the massive modulation of activity of over 2000 genes. Expression of genes associated with plant immune responses (protein kinase and mitogen-activated protein kinase), hypersensitive responses, phytohormone signaling pathways, photosynthesis, pigment metabolism, protein metabolism, sugar metabolism, and modification, and others were altered, which could be attributed to systemic symptom development upon CBCVd-infection in hop. In addition, genes encoding RNA-dependent RNA polymerase, pathogenesis-related protein, chitinase, as well as those related to basal defense responses were up-regulated. The expression levels of several genes identified from RNA sequencing analysis were confirmed by qRT-PCR. Our systematic comprehensive CBCVd-responsive transcriptome analysis provides a better understanding and insights into complex viroid-hop plant interaction. This information will assist further in the development of future measures for the prevention of CBCVd spread in hop fields.

• Klíčová slova
• Citrus bark cracking viroid, differentially expressed genes, hop, pathogen, transcriptome analysis, viroids,
• MeSH
• Humulus genetika   metabolismus   virologie   MeSH
• listy rostlin genetika   metabolismus   virologie   MeSH
• nemoci rostlin genetika   virologie   MeSH
• regulace genové exprese u rostlin MeSH
• rostlinné proteiny genetika   metabolismus   MeSH
• rostlinné viry genetika   izolace a purifikace   fyziologie   MeSH
• stanovení celkové genové exprese MeSH
• viroidy klasifikace   genetika   izolace a purifikace   fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• rostlinné proteiny MeSH