Genome 3 T A Brown Pdf.zip
Adrian Rocher
Complex physiological adaptations often involve the coordination of molecular responses across multiple tissues. Establishing transcriptomic resources for non-traditional model organisms with phenotypes of interest can provide a foundation for understanding the genomic basis of these phenotypes, and the degree to which these resemble, or contrast, those of traditional model organisms. Here, we present a one-of-a-kind gene expression dataset generated from multiple tissues of two hibernating brown bears (Ursus arctos).
This dataset is comprised of 26 samples collected from 13 tissues of two hibernating brown bears. These samples were collected opportunistically and are typically not possible to attain, resulting in a highly unique and valuable gene expression dataset. In combination with previously published datasets, this new transcriptomic resource will facilitate detailed investigation of hibernation physiology in bears, and the potential to translate aspects of this biology to treat human disease.
The brown planthopper, Nilaparvata lugens, the most destructive pest of rice, is a typical monophagous herbivore that feeds exclusively on rice sap, which migrates over long distances. Outbreaks of it have re-occurred approximately every three years in Asia. It has also been used as a model system for ecological studies and for developing effective pest management. To better understand how a monophagous sap-sucking arthropod herbivore has adapted to its exclusive host selection and to provide insights to improve pest control, we analyzed the genomes of the brown planthopper and its two endosymbionts.
Our study reveals a series of complex adaptations of the brown planthopper involving a variety of biological processes, that result in its highly destructive impact on the exclusive host rice. All these findings highlight potential directions for effective pest control of the planthopper.
The brown planthopper. (A) Short-winged (brachypterous) and long-winged (macropterous) female adults and nymph. (B) Schematic diagram of BPH distribution worldwide and possible northward migratory routes (arrows) in East China.
The BPH genome size is estimated to be approximately 1.2 Gbp using a k-mer approach, which is consistent with our previous estimates using flow cytometry. However, analysis of 17-mers suggest higher than expected heterozygosity of the BPH genome despite 13 generations of inbreeding, which presents a challenge for de novo genome assembly when using only whole-genome shotgun (WGS) sequence data. To overcome the challenge caused by high heterozygosity and repeat sequence content, we employed a hybrid method that integrates WGS sequences with pooled fosmid sequencing. The TrimDup module in Rabbit was used to remove redundant and heterozygous sequences [13]. Finally we assembled a draft BPH genome of 1.14 Gbp, with a scaffold N50 of 356.6 kbp and a contig N50 of 24.2 kbp (Table 1). We evaluated the completeness of the draft genome assembly by mapping expressed sequence tags (ESTs) to the genome and by calculating coverage for a set of 248 core eukaryotic genes using CEGMA [14], which show genome coverage rates of 97.1% and >96%, respectively (Tables S1 to S8 and Figures S1 to S5 in Additional file 1).
A significant proportion of the BPH genome contains a high level of repetitive sequences (48.6%, including tandem repeats and transposable elements), which is a larger fraction than that measured in the pea aphid (33.3%) [15]; tandem repeats account for 6.4% of the whole genome. Transposable elements (TEs) were identified at both the DNA and inferred protein level. The TEs account for approximately 38.90% of the BPH genome, including DNA repeats (14.2%), long interspersed nuclear elements (LINEs; 16.0%), long terminal repeats (LTRs; 14.8%), short interspersed nuclear elements (SINEs; 0.7%), and unknown repeat types (1.9%). Comparison of TEs identified through homology-based and de novo prediction approaches against those from Repbase revealed a shift of the peak sequence divergence ratio. This finding suggests that the BPH-specific TEs, especially DNA transposons, have evolved relatively recently, and likely contribute to the large genome size of BPH (Tables S9 and S10 and Figure S8 in Additional file 1).
We predicted protein-coding genes using GENEWISE [17], an homology-based method referring to protein sequences from four representative insects and from human. We also used the programs GENSCAN [18] and AUGUSTUS [19] for additional ab initio gene predictions. These results were then combined using GLEAN to generate a consensus gene set [20]. A 2.47 Gbp RNA-seq data set was additionally used to complement the combined gene set. Finally, we created a reference gene set containing 27,571 protein-coding genes for BPH. Among the 15 arthropod genomes compared in this study, the numbers of predicted genes and species-specific genes in BPH were lower than in the pea aphid (Table 1), but higher than those of most other insects. The lack of accumulated knowledge on arthropod genomes in general might have attributed to the elevated species-specific gene components in BPH because sequenced arthropod genomes are limited and highly biased in phylogenetic coverage. For instance, the first sequenced crustacean, the waterflea (Daphnia pulex), has an unexpected proportion of unknown genes (36% of over 30,000 genes) when compared with other genomes [21]. Similarly, as most published insect genomes are from holometabolans, 37% of the predicted genes in the pea aphid (the first basal hemimetabolous species with sequenced genome) could not be matched to any known species [15]. Although the likelihood of overestimation during gene predictions cannot be completely ruled out, the pattern of elevated proportions of species-specific genes is shared among all hemipteran genomes (A. pisum, Rhodnius prolixus, and N. lugens (Figure 2). We expect that a higher level of homology can be discovered when additional genomes are sequenced for more hemipteran insects.
Gene family expansions and contractions in the brown planthopper compared with other arthropod genomes. Numbers for expanded (green) and contracted (red) gene families are shown below branches or taxon names with percentages indicated by pie charts.
Phylogenetic relationships and gene orthology based on the genomes of 15 arthropod species. (A) phylogenetic relations of BPH to insects and other arthropods based on single-copy orthologous genes obtained from full genomes. Thirteen insect species were used for the analysis, including Bombyx mori, Danaus plexippus, Anopheles gambiae, Aedes aegypti, Drosophila melanogaster, Tribolium castaneum, Apis mellifera, Camponotus floridanus, Nasonia vitripennis, Pediculus humanus, Rhodnius prolixus, Nilaparvata lugens, and Acyrthosiphon pisum. Two Arthropoda animals (Daphnia pulex, Tetranychus urticae) were used as outgroup taxa. Branch lengths represents divergence times estimated by second codon positions of 318 single-copy genes (Table S17 in Additional file 1) using PhyML [80],[81], with a gamma distribution across sites and an HKY85 substitution model. The branch supports were inferred based on approximate likelihood ratio test (aLRT) (B) Gene orthology comparison among the genomes of 15 arthropod species. Note: the order of the 15 species follows that in Figure 3A. 1:1:1 refers to single-copy gene orthologs found across all 15 lineages. N:N:N refers to multi-copy gene paralogs found across all 15 lineages. Diptera, Lepidoptera, Hymenoptera, Hemiptera, and Insect refer to taxon-specific genes that are present only in the relevant lineage. SD indicates species-specific genes in multiple copies. ND indicates species-specific genes in single copies.
Both the percentage of species-specific genes and the total gene number of all three hemipteran genomes are higher than for other insect genomes, indicating an overall expansion of the gene repertoire in species of the hemipteran lineage (Figure 2; Tables S18 and S19 in Additional file 1). The expanded genes are over-represented in Gene Ontology terms including sodium channel activity, zinc ion binding, helicase activity, and so on, whereas gene losses are most pronounced in detoxification and digestion genes, including P450, GST, and chymotrypsin gene families (Tables S20 and S21 in Additional file 1). These lineage-specific gene expansions and contractions (or losses) likely reflect the evolutionary adaptation of hemipteran insects to plant phloem sap as their food sources and their symbiotic dependencies.
Chemoreception is essential for herbivorous insects in host plant selection [25]. Four multi-gene families are involved in the chemoreceptor system: odorant binding proteins (OBPs), chemosensory proteins (CSPs), odorant receptors (ORs), and gustatory receptors (GRs). In the BPH genome, we identified 11 genes coding for OBPs, 17 for CSPs, 50 for ORs, and 10 for GRs.
The genes encoding cytochrome P450 monooxygenases (P450s) and glutathione S-transferases (GSTs) are members of the major multigene enzyme families that are primarily responsible for detoxification of xenobiotic compounds, for example, insecticides and plant secondary metabolites [31],[32]. The expansion and contraction of these gene families in insects seem to be correlated with the levels of exposure to external stressors, for example, host defense systems and environmental toxins. A comparison of insect species with available genomes (Table 3) reveals that N. lugens has much fewer P450 genes (67) compared with other insects; it is ranked second to A. mellifera (which has 46) in having the fewest genes. The genes encoding cytochrome P450 monooxygenases, including CYP6a2, CYP6a13, CYP6a14, CYP6k1 and CYP3A18, which belong to the CYP4 subfamily, have all been lost by N. lugens. It has been suggested that the relatively low number of P450 genes in honeybee may be a consequence of the social organization of the beehive, which probably shields the queen and larvae from environmental exposure to toxins, permitting a subsequent loss of P450 genes in this species [33]. The comparison of the genome data also shows that N. lugens and A. mellifera have the least number of GSTs among all sequenced insects, only half that in A. pisum (Table 3). Interestingly, the plant sap-sucking but polyphagous A. pisum possesses more P450 and GST genes, which is congruent with its need to detoxify compounds from hundreds of host species.
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