Paleopolyploidy in the Brassicales: Analyses of the Cleome Transcriptome Elucidate the History of Genome Duplications in Arabidopsis and Other Brassicales

The analysis of the Arabidopsis genome revealed evidence of three ancient polyploidy events in the evolution of the Brassicaceae, but the exact phylogenetic placement of these events is still not resolved. The most recent event is called the At-α (alpha) or 3R, the intermediate event is referred to as the At-β (beta) or 2R, and the oldest is the At-γ (gamma) or 1R. It has recently been established that At-γ is shared with other Rosids, including papaya (Carica), poplar (Populus), and grape (Vitis), whereas data to date suggest that At-α is Brassicaceae specific. To address more precisely when the At-α and At-β events occurred and which plant lineages share these paleopolyploidizations, we sequenced and analyzed over 4,700 normalized expressed sequence tag sequences from the Cleomaceae, the sister family to the Brassicaceae. Analysis of these Cleome data with homologous sequences from other Rosid genomes (Arabidopsis, Carica, Gossypium, Populus, and Vitis) yielded three major findings: 1) confirmation of a Cleome-specific paleopolyploidization (Cs-α) that is independent of the Brassicaceae At-α paleopolyploidization; 2) Cleome and Arabidopsis share the At-β duplication, which is lacking from papaya within the Brassicales; and 3) rates of molecular evolution are faster for the herbaceous annual taxa Arabidopsis and Cleome than the other predominantly woody perennial Rosid lineages. These findings contribute to our understanding of the dynamics of genome duplication and evolution within one of the most comprehensively surveyed clades of plants, the Rosids, and clarify the complex history of the At-α, At-β, and At-γ duplications of Arabidopsis

Insight into the evolution of the Solanaceae from the parental genomes of Petunia hybrida

Petunia hybrida is a popular bedding plant that has a long history as a genetic model system. We report the whole-genome sequencing and assembly of inbred derivatives of its two wild parents, P. axillaris N and P. inflata S6. The current assemblies include 91.3% and 90.2% coverage of their diploid genomes (1.4 Gb; 2n=14) containing 32,928 and 36,697 protein-coding genes, respectively. The Petunia lineage has experienced at least two rounds of paleohexaploidization, the older gamma hexaploidy event, which is shared with other Eudicots, and the more recent Solanaceae paleohexaploidy event that is shared with tomato and other Solanaceae species. Transcription factors that were targets of selection during the shift from bee- to moth pollination reside in particularly dynamic regions of the genome, which may have been key to the remarkable diversity of floral color patterns and pollination systems. The high quality genome sequences will enhance the value of Petunia as a model system for basic and applied research on a variety of unique biological phenomena

Genomic investigations of unexplained acute hepatitis in children

Since its first identification in Scotland, over 1,000 cases of unexplained paediatric hepatitis in children have been reported worldwide, including 278 cases in the UK1. Here we report an investigation of 38 cases, 66 age-matched immunocompetent controls and 21 immunocompromised comparator participants, using a combination of genomic, transcriptomic, proteomic and immunohistochemical methods. We detected high levels of adeno-associated virus 2 (AAV2) DNA in the liver, blood, plasma or stool from 27 of 28 cases. We found low levels of adenovirus (HAdV) and human herpesvirus 6B (HHV-6B) in 23 of 31 and 16 of 23, respectively, of the cases tested. By contrast, AAV2 was infrequently detected and at low titre in the blood or the liver from control children with HAdV, even when profoundly immunosuppressed. AAV2, HAdV and HHV-6 phylogeny excluded the emergence of novel strains in cases. Histological analyses of explanted livers showed enrichment for T cells and B lineage cells. Proteomic comparison of liver tissue from cases and healthy controls identified increased expression of HLA class 2, immunoglobulin variable regions and complement proteins. HAdV and AAV2 proteins were not detected in the livers. Instead, we identified AAV2 DNA complexes reflecting both HAdV-mediated and HHV-6B-mediated replication. We hypothesize that high levels of abnormal AAV2 replication products aided by HAdV and, in severe cases, HHV-6B may have triggered immune-mediated hepatic disease in genetically and immunologically predisposed children

Synteny Identifies Reliable Orthologs for Phylogenomics and Comparative Genomics of the Brassicaceae

Large genomic data sets are becoming the new normal in phylogenetic research, but the identification of true orthologous genes and the exclusion of problematic paralogs is still challenging when applying commonly used sequencing methods such as target enrichment. Here, we compared conventional ortholog detection using OrthoFinder with ortholog detection through genomic synteny in a data set of 11 representative diploid Brassicaceae whole-genome sequences spanning the entire phylogenetic space. Then, we evaluated the resulting gene sets regarding gene number, functional annotation, and gene and species tree resolution. Finally, we used the syntenic gene sets for comparative genomics and ancestral genome analysis. The use of synteny resulted in considerably more orthologs and also allowed us to reliably identify paralogs. Surprisingly, we did not detect notable differences between species trees reconstructed from syntenic orthologs when compared with other gene sets, including the Angiosperms353 set and a Brassicaceae-specific target enrichment gene set. However, the synteny data set comprised a multitude of gene functions, strongly suggesting that this method of marker selection for phylogenomics is suitable for studies that value downstream gene function analysis, gene interaction, and network studies. Finally, we present the first ancestral genome reconstruction for the Core Brassicaceae which predating the Brassicaceae lineage diversification ∼25 million years ago

Highly differentiated genomic properties underpin the different cell walls of Poaceae and eudicots

Plant cell walls of Poaceae and eudicots differ substantially, both in the content and composition of their components. However, the genomic and genetic basis underlying these differences is not fully resolved. In this research, we analyzed multiple genomic properties of 150 cell wall gene families across 169 angiosperm genomes. The properties analyzed include gene presence/absence, copy number, synteny, occurrence of tandem gene clusters, and phylogenetic gene diversity. Results revealed a profound genomic differentiation of cell wall genes between Poaceae and eudicots, often associated with the cell wall diversity between these plant groups. For example, overall patterns of gene copy number variation and synteny were clearly divergent between Poaceae and eudicot species. Moreover, differential Poaceae–eudicot copy number and genomic contexts were observed for all the genes within the BEL1-like HOMEODOMAIN 6 regulatory pathway, which respectively induces and represses secondary cell wall synthesis in Poaceae and eudicots. Similarly, divergent synteny, copy number, and phylogenetic gene diversification were observed for the major biosynthetic genes of xyloglucans, mannans, and xylans, potentially contributing to the differences in content and types of hemicellulosic polysaccharides differences in Poaceae and eudicot cell walls. Additionally, the Poaceae-specific tandem clusters and/or higher copy number of PHENYLALANINE AMMONIA-LYASE, CAFFEIC ACID O-METHYLTRANSFERASE, or PEROXIDASE genes may underly the higher content and larger variety of phenylpropanoid compounds observed in Poaceae cell walls. All these patterns are discussed in detail in this study, along with their evolutionary and biological relevance for cell wall (genomic) diversification between Poaceae and eudicots

Flowering locus C (FLC) is a potential major regulator of glucosinolate content across developmental stages of Aethionema arabicum (brassicaceae)

The biochemical defense of plants can change during their life-cycle and impact herbivore feeding and plant fitness. The annual species Aethionema arabicum is part of the sister clade to all other Brassicaceae. Hence, it holds a phylogenetically important position for studying crucifer trait evolution. Glucosinolates (GS) are essentially Brassicales-specific metabolites involved in plant defense. Using two Ae. arabicum accessions (TUR and CYP) we identify substantial differences in glucosinolate profiles and quantities between lines, tissues and developmental stages. We find tissue specific side-chain modifications in aliphatic GS: methylthioalkyl in leaves, methylsulfinylalkyl in fruits, and methylsulfonylalkyl in seeds. We also find large differences in absolute glucosinolate content between the two accessions (up to 10-fold in fruits) that suggest a regulatory factor is involved that is not part of the quintessential glucosinolate biosynthetic pathway. Consistent with this hypothesis, we identified a single major multi-trait quantitative trait locus controlling total GS concentration across tissues in a recombinant inbred line population derived from TUR and CYP. With fine-mapping, we narrowed the interval to a 58 kb region containing 15 genes, but lacking any known GS biosynthetic genes. The interval contains homologs of both the sulfate transporter SULTR2;1 and FLOWERING LOCUS C. Both loci have diverse functions controlling plant physiological and developmental processes and thus are potential candidates regulating glucosinolate variation across the life-cycle of Aethionema. Future work will investigate changes in gene expression of the candidates genes, the effects of GS variation on insect herbivores and the trade-offs between defense and reproduction.</p

Genomic Blocks in Aethionema arabicum Support Arabideae as Next Diverging Clade in Brassicaceae

The tribe Aethionemeae is sister to all other crucifers, making it a crucial group for unraveling genome evolution and phylogenetic relationships within the crown group Brassicaceae. In this study, we extend the analysis of Brassicaceae genomic blocks (GBs) to Aethionema whereby we identified unique block boundaries shared only with the tribe Arabideae. This was achieved using bioinformatic methods to analyze synteny between the recently updated genome sequence of Aethionema arabicum and other high-quality Brassicaceae genome sequences. We show that compared to the largely conserved genomic structure of most non-polyploid Brassicaceae lineages, GBs are highly rearranged in Aethionema. Furthermore, we detected similarities between the genomes of Aethionema and Arabis alpina, in which also a high number of genomic rearrangements compared to those of other Brassicaceae was found. These similarities suggest that tribe Arabideae, a clade showing conflicting phylogenetic position between studies, may have diverged before diversification of the other major lineages, and highlight the potential of synteny information for phylogenetic inference.</p

Flowering locus C (FLC) is a potential major regulator of glucosinolate content across developmental stages of Aethionema arabicum (brassicaceae)

The biochemical defense of plants can change during their life-cycle and impact herbivore feeding and plant fitness. The annual species Aethionema arabicum is part of the sister clade to all other Brassicaceae. Hence, it holds a phylogenetically important position for studying crucifer trait evolution. Glucosinolates (GS) are essentially Brassicales-specific metabolites involved in plant defense. Using two Ae. arabicum accessions (TUR and CYP) we identify substantial differences in glucosinolate profiles and quantities between lines, tissues and developmental stages. We find tissue specific side-chain modifications in aliphatic GS: methylthioalkyl in leaves, methylsulfinylalkyl in fruits, and methylsulfonylalkyl in seeds. We also find large differences in absolute glucosinolate content between the two accessions (up to 10-fold in fruits) that suggest a regulatory factor is involved that is not part of the quintessential glucosinolate biosynthetic pathway. Consistent with this hypothesis, we identified a single major multi-trait quantitative trait locus controlling total GS concentration across tissues in a recombinant inbred line population derived from TUR and CYP. With fine-mapping, we narrowed the interval to a 58 kb region containing 15 genes, but lacking any known GS biosynthetic genes. The interval contains homologs of both the sulfate transporter SULTR2;1 and FLOWERING LOCUS C. Both loci have diverse functions controlling plant physiological and developmental processes and thus are potential candidates regulating glucosinolate variation across the life-cycle of Aethionema. Future work will investigate changes in gene expression of the candidates genes, the effects of GS variation on insect herbivores and the trade-offs between defense and reproduction.</p

Drivers of metabolic diversification: how dynamic genomic neighbourhoods generate new biosynthetic pathways in the Brassicaceae

Plants produce an array of specialized metabolites with important ecological functions. The mechanisms underpinning the evolution of new biosynthetic pathways are not well‐understood. Here, we exploit available genome sequence resources to investigate triterpene biosynthesis across the Brassicaceae. Oxidosqualene cyclases (OSCs) catalyze the first committed step in triterpene biosynthesis. Systematic analysis of 13 sequenced Brassicaceae genomes was performed to identify all OSC genes. The genome neighbourhoods (GNs) around a total of 163 OSC genes were investigated to identify Pfam domains significantly enriched in these regions. All‐vs‐all comparisons of OSC neighbourhoods and phylogenomic analysis were used to investigate the sequence similarity and evolutionary relationships of the numerous candidate triterpene biosynthetic gene clusters (BGCs) observed. Functional analysis of three representative BGCs was carried out and their triterpene pathway products were elucidated. Our results indicate that plant genomes are remarkably plastic, and that dynamic GNs generate new biosynthetic pathways in different Brassicaceae lineages by shuffling the genes encoding a core palette of triterpene‐diversifying enzymes, presumably in response to strong environmental selection pressure. These results illuminate a genomic basis for diversification of plant‐specialized metabolism through natural combinatorics of enzyme families, which can be mimicked using synthetic biology to engineer diverse bioactive molecules