Genome-wide analysis of the H3K27me3 epigenome and transcriptome in brassica rapa

Background Genome-wide maps of histone modifications have been obtained for several plant species. However, most studies focus on model systems and do not enforce FAIR data management principles. Here we study the H3K27me3 epigenome and associated transcriptome of Brassica rapa, an important vegetable cultivated worldwide. Findings We performed H3K27me3 chromatin immunoprecipitation followed by high-throughput sequencing and transcriptomic analysis by 3′-end RNA sequencing from B. rapa leaves and inflorescences. To analyze these data we developed a Reproducible Epigenomic Analysis pipeline using Galaxy and Jupyter, packaged into Docker images to facilitate transparency and reuse. We found that H3K27me3 covers roughly one-third of all B. rapa protein-coding genes and its presence correlates with low transcript levels. The comparative analysis between leaves and inflorescences suggested that the expression of various floral regulatory genes during development depends on H3K27me3. To demonstrate the importance of H3K27me3 for B. rapa development, we characterized a mutant line deficient in the H3K27 methyltransferase activity. We found that braA.clf mutant plants presented pleiotropic alterations, e.g., curly leaves due to increased expression and reduced H3K27me3 levels at AGAMOUS-like loci. Conclusions We characterized the epigenetic mark H3K27me3 at genome-wide levels and provide genetic evidence for its relevance in B. rapa development. Our work reveals the epigenomic landscape of H3K27me3 in B. rapa and provides novel genomics datasets and bioinformatics analytical resources. We anticipate that this work will lead the way to further epigenomic studies in the complex genome of Brassica crops

Cloning, heterologous expression and biochemical characterization of plastidial sn-glycerol-3-phosphate acyltransferase from Helianthus annuus

© 2015 Elsevier Ltd. All rights reserved. The acyl-[acyl carrier protein]:sn-1-glycerol-3-phosphate acyltransferase (GPAT; E.C. 2.3.1.15) catalyzes the first step of glycerolipid assembly within the stroma of the chloroplast. In the present study, the sunflower (Helianthus annuus, L.) stromal GPAT was cloned, sequenced and characterized. We identified a single ORF of 1344 base pairs that encoded a GPAT sharing strong sequence homology with the plastidial GPAT from Arabidopsis thaliana (ATS1, At1g32200). Gene expression studies showed that the highest transcript levels occurred in green tissues in which chloroplasts are abundant. The corresponding mature protein was heterologously overexpressed in Escherichia coli for purification and biochemical characterization. In vitro assays using radiolabelled acyl-ACPs and glycerol-3-phosphate as substrates revealed a strong preference for oleic versus palmitic acid, and weak activity towards stearic acid. The positional fatty acid composition of relevant chloroplast phospholipids from sunflower leaves did not reflect the in vitro GPAT specificity, suggesting a more complex scenario with mixed substrates at different concentrations, competition with other acyl-ACP consuming enzymatic reactions, etc. In summary, this study has confirmed the affinity of this enzyme which would partly explain the resistance to cold temperatures observed in sunflower plants.Thanks are due to A. Gonzalez-Callejas and B. Lopez-Cordero for skillful technical assistance. This work was supported by the ‘‘Ministerio de Economia y Competitividad’’ and FEDER, project AGL2011-23187.Peer Reviewe

Regulation of gene expression in roots of the pH-sensitive Vaccinium corymbosum and the pH-tolerant Vaccinium arboreum in response to near neutral pH stress using RNA-Seq

Abstract Background Blueberries are one of the few horticultural crops adapted to grow in acidic soils. Neutral to basic soil pH is detrimental to all commonly cultivated blueberry species, including Vaccinium corymbosum (VC). In contrast, the wild species V. arboreum (VA) is able to tolerate a wider range of soil pH. To assess the molecular mechanisms involved in near neutral pH stress response, plants from pH-sensitive VC (tetraploid) and pH-tolerant VA (diploid) were grown at near neutral pH 6.5 and at the preferred pH of 4.5. Results Transcriptome sequencing of root RNA was performed for 4 biological replications per species x pH level interaction, for a total of 16 samples. Reads were mapped to the reference genome from diploid V. corymbosum, transforming ~55% of the reads to gene counts. A quasi-likelihood F test identified differential expression due to pH stress in 337 and 4867 genes in VA and VC, respectively. Both species shared regulation of genes involved in nutrient homeostasis and cell wall metabolism. VA and VC exhibited differential regulation of signaling pathways related to abiotic/biotic stress, cellulose and lignin biosynthesis, and nutrient uptake. Conclusions The specific responses in VA likely facilitate tolerance to higher soil pH. In contrast, response in VC, despite affecting a greater number of genes, is not effective overcoming the stress induced by pH. Further inspection of those genes with differential expression that are specific in VA may provide insight on the mechanisms towards tolerance

Additional file 5: Table S3. of Regulation of gene expression in roots of the pH-sensitive Vaccinium corymbosum and the pH-tolerant Vaccinium arboreum in response to near neutral pH stress using RNA-Seq

Enrichment analysis of GO terms. Genes showing differential expression in VA (A) and VC (B) were subjected to singular enrichment analysis in AgriGO. The table contains output from the analysis performed using the combination AgriGO and REVIGO tools. GO terms with FDR < 0.5 after Yekutieli multi-test in AgriGO were used on REVIGO and searched for genes with that specific annotation. (XLSX 184 kb

Species diversity and phylogeography of Cornus kousa (Asian dogwood) captured by genomic and genic microsatellites

14 Pág.Cornus kousa (Asian dogwood), an East Asia native tree, is the most economically important species of the dogwood genus, owing to its desirable horticultural traits and ability to hybridize with North America-native dogwoods. To assess the species genetic diversity and to better inform the ongoing and future breeding efforts, we assembled an herbarium and arboretum collection of 131 noncultivated C. kousa specimens. Genotyping and capillary electrophoresis analyses of our C. kousa collection with the newly developed genic and published nuclear genomic microsatellites permitted assessment of genetic diversity and evolutionary history of the species. Regardless of the microsatellite type used, the study yielded generally similar insights into the C. kousa diversity with subtle differences deriving from and underlining the marker used. The accrued evidence pointed to the species distinct genetic pools related to the plant country of origin. This can be helpful in the development of the commercial cultivars for this important ornamental crop with increased pyramided utility traits. Analyses of the C. kousa evolutionary history using the accrued genotyping datasets pointed to an unsampled ancestor population, possibly now extinct, as per the phylogeography of the region. To our knowledge, there are few studies utilizing the same gDNA collection to compare performance of genomic and genic microsatellites. This is the first detailed report on C. kousa species diversity and evolutionary history inference.This research was funded fully by the United States Department of Agriculture—Agricultural Research Service grant (NACA 58-6062-6) to the Trigiano Lab to investigate woody ornamental genomics. The funding agency had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Funding for open access to this research was provided by University of Tennessee’s Open Publishing Support Fund. The authors thank the following herbaria for samples for destructive testing used in this study: Carnegie of Natural History Herbarium, Pittsburgh, PA, USA, and Ray J. Davis Herbarium, Idaho State University, Pocatello, ID, USA. We are also grateful to the following arboreta for samples of their materials: Morton Arboretum, Lisle, IL, USA; U.S. National Arboretum, Beltsville, MD, USA; and Morris Arboretum of the University of Pennsylvania, Philadelphia, PA, USA. The use of any trade names is for identification purposes only and does not imply their endorsement by the authors or the study funding entities.Peer reviewe

Genome-wide analysis of the H3K27me3 epigenome and transcriptome in brassica rapa

Background Genome-wide maps of histone modifications have been obtained for several plant species. However, most studies focus on model systems and do not enforce FAIR data management principles. Here we study the H3K27me3 epigenome and associated transcriptome of Brassica rapa, an important vegetable cultivated worldwide. Findings We performed H3K27me3 chromatin immunoprecipitation followed by high-throughput sequencing and transcriptomic analysis by 3′-end RNA sequencing from B. rapa leaves and inflorescences. To analyze these data we developed a Reproducible Epigenomic Analysis pipeline using Galaxy and Jupyter, packaged into Docker images to facilitate transparency and reuse. We found that H3K27me3 covers roughly one-third of all B. rapa protein-coding genes and its presence correlates with low transcript levels. The comparative analysis between leaves and inflorescences suggested that the expression of various floral regulatory genes during development depends on H3K27me3. To demonstrate the importance of H3K27me3 for B. rapa development, we characterized a mutant line deficient in the H3K27 methyltransferase activity. We found that braA.clf mutant plants presented pleiotropic alterations, e.g., curly leaves due to increased expression and reduced H3K27me3 levels at AGAMOUS-like loci. Conclusions We characterized the epigenetic mark H3K27me3 at genome-wide levels and provide genetic evidence for its relevance in B. rapa development. Our work reveals the epigenomic landscape of H3K27me3 in B. rapa and provides novel genomics datasets and bioinformatics analytical resources. We anticipate that this work will lead the way to further epigenomic studies in the complex genome of Brassica crops

Conserved and distinct roles of H3K27me3 demethylases regulating flowering time in Brassica rapa

14 Pág.Epigenetic regulation is necessary for optimal organism development and preservation of gene expression profiles in the cell. In plants, the trimethylation of histone H3 lysine 27 (H3K27me3) is a silencing epigenetic mark relevant for developmental transitions like flowering. The floral transition is a key agronomic trait; however, the epigenetic mechanisms of flowering time regulation in crops remain poorly understood. Here we study the Jumonji H3K27me3 demethylases BraA.REF6 and BraA.ELF6 in Brassica rapa. Phenotypic characterization of novel mutant lines and genome-wide H3K27me3 chromatin immunoprecipitation and transcriptomic analyses indicated that BraA.REF6 plays a greater role than BraA.ELF6 in fine-tuning H3K27me3 levels. In addition, we found that braA.elf6 mutants were early flowering due to high H3K27me3 levels at B. rapa homologs of the floral repressor FLC. Unlike mutations in Arabidopsis thaliana, braA.ref6 mutants were late flowering without altering the expression of B. rapa FLC genes. Remarkably, we found that BraA.REF6 regulated a number of gibberellic acid (GA) biosynthetic genes, including a homolog of GA1, and that GA-treatment complemented the late flowering mutant phenotype. This study increases our understanding of the epigenetic regulation of flowering time in B. rapa, highlighting conserved and distinct regulatory mechanisms between model and crop species.This study was supported by grants BIO2015-68031-R and RTI2018-097749-B-I00 to PC, and PID2019-104899GB-I00 to JAJ and MP from MCIN/AEI/10.13039/501100011033 and FEDER; and fellowships BES-2016-078939 to LP and RYC-2013-14689 to PC from MCIN/AEI/10.13039/501100011033 and FSE. MPM was supported by a Postdoctoral contract associated to the “Severo Ochoa Program for Centres of Excellence in R&D” grant SEV-2016-0672 (2017–2021) to CBGP from MCIN/AEI/10.13039/501100011033.Peer reviewe

Triacylglycerols Biosynthesis in Sunflower Seeds.

The amount of oil in seeds of different species varies from 1% to as much as 60% of seed dry weight. The oilseed storage reserve provides the necessary energy for the germination and establishment of the subsequent seedling. In sunflower, triacylglycerols (TAGs) represent the main component of seed oil. Sunflower storage lipid synthesis occurs in developing seeds over a period of about two weeks. The biosynthesis of triacylglycerols (TAGs) is a complex process consisting of three well-differentiated phases: (1) de novo synthesis of fatty acids in the plastid, (2) formation of the acyl-CoA pool in the cytosol, and (3) TAG assembling in the endoplasmic reticulum of developing seed tissues. Synthesized TAGs are then stored into densely packed oil bodies (oleosomes) until their utilization. TAG assembly in the ER is closely associated with membrane metabolism and involves the action of several membrane-bound acyltransferase enzymes. Until recently TAG biosynthesis was viewed as a sequential linear process where acyl-CoAs were sequentially assembled onto the sn-1, sn-2, and sn-3 positions of the glycerol backbone, through the so-called Kennedy or glycerol phosphate pathway. However, in the last decade, new activities involved in alternative pathways also operating in the TAG biosynthesis have been discovered and characterized. Therefore, multiple interconnected pathways for TAG synthesis have actually been elucidated in plants. Overall, the sunflower TAG synthesis model highlights a complex network of enzymatic reactions mediating the movement of acyl groups between PC, acyl-CoA, DAG, PA, and TAG and contributes to the final stereospecific composition of storage TAG.Peer Reviewe