Dataset Information

Genome-wide gene expression analysis of amphioxus (Branchiostoma belcheri) following lipopolysaccharide challenge using strand-specific RNA-seq.

ABSTRACT: Amphioxus is the closest living proxy for exploring the evolutionary origin of the immune system in vertebrates. To understand the immune responses of amphioxus to lipopolysaccharide (LPS), 5 ribosomal RNA (rRNA)-depleted libraries of amphioxus were constructed, including one control (0 h) library and 4 treatment libraries at 6, 12, 24, and 48 h post-injection (hpi) with LPS. The transcriptome of Branchiostoma belcheri was analyzed using strand-specific RNA sequencing technology (RNA-seq). A total of 6161, 6665, 7969, and 6447 differentially expressed genes (DEGs) were detected at 6, 12, 24, and 48 hpi, respectively, compared with expression levels at 0 h. We identified amphioxus genes active during the acute-phase response to LPS at different time points after stimulation. Moreover, to better visualize the resolution phase of the immune process during immune response, we identified 6057 and 5235 DEGs at 48 hpi by comparing with 6 and 24 hpi, respectively. Through real-time quantitative PCR (qRT-PCR) analysis of 12 selected DEGs, we demonstrated the accuracy of the RNA-seq data in this study. Functional enrichment analysis of DEGs demonstrated that most terms were related to defense and immune responses, disease and infection, cell apoptosis, and metabolism and catalysis. Subsequently, we identified 1330, 485, 670, 911, and 1624 time-specific genes (TSGs) at 0, 6, 12, 24, and 48 hpi. Time-specific terms at each of 5 time points were primarily involved in development, immune signaling, signal transduction, DNA repair and stability, and metabolism and catalysis, respectively. As this is the first study to report the transcriptome of an organism with primitive immunity following LPS challenge at multiple time points, it provides gene expression information for further research into the evolution of immunity in vertebrates.

SUBMITTER: Zhang QL

PROVIDER: S-EPMC5731807 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Genome-wide gene expression analysis of amphioxus (Branchiostoma belcheri) following lipopolysaccharide challenge using strand-specific RNA-seq.

Zhang Qi-Lin QL Zhu Qian-Hua QH Xie Zheng-Qing ZQ Xu Bin B Wang Xiu-Qiang XQ Chen Jun-Yuan JY

RNA biology 20170921 12

Amphioxus is the closest living proxy for exploring the evolutionary origin of the immune system in vertebrates. To understand the immune responses of amphioxus to lipopolysaccharide (LPS), 5 ribosomal RNA (rRNA)-depleted libraries of amphioxus were constructed, including one control (0 h) library and 4 treatment libraries at 6, 12, 24, and 48 h post-injection (hpi) with LPS. The transcriptome of Branchiostoma belcheri was analyzed using strand-specific RNA sequencing technology (RNA-seq). A tot ...[more]

PMID: 28837390

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Project description:Global climatic fluctuations governed the ancestral demographic histories of species and contributed to place the current population status into a more extensive ecological and evolutionary context. Genetic variations will leave unambiguous signatures in the patterns of intraspecific genetic variation in extant species since the genome of each individual is an imperfect mosaic of the ancestral genomes. Here, we report the genome sequences of 20 Branchiostoma individuals by whole-genome resequencing strategy. We detected over 140 million genomic variations for each Branchiostoma individual. In particular, we applied the pairwise sequentially Markovian coalescent (PSMC) method to estimate the trajectories of changes in the effective population size (N e) of Branchiostoma population during the Pleistocene. We evaluated the threshold of sequencing depth for proper inference of demographic histories using PSMC was ≥25×. The PSMC results highlight the role of historical global climatic fluctuations in the long-term population dynamics of Branchiostoma. The inferred ancestral N e of the Branchiostoma belcheri populations from Zhanjiang and Xiamen (China) seawaters was different in amplitude before the first (mutation rate = 3 × 10-9) or third glaciation (mutation rate = 9 × 10-9) of the Pleistocene, indicating that the two populations most probably started to evolve in isolation in their respective seas after the first or third glaciation of the Pleistocene. A pronounced population bottleneck coinciding with the last glacial maximum was observed in all Branchiostoma individuals, followed by a population expansion occurred during the late Pleistocene. Species that have experienced long-term declines may be especially vulnerable to recent anthropogenic activities. Recently, the industrial pollution and the exploitation of sea sand have destroyed the harmonious living environment of amphioxus species. In the future, we need to protect the habitat of Branchiostoma and make full use of these detected genetic variations to facilitate the functional study of Branchiostoma for adaptation to local environments.

Project description:As the extant representatives of the basal chordate lineage, amphioxi (including the genera Branchiostoma, Asymmetron and Epigonichthys) play important roles in tracing the state of chordate ancestry. Previous studies have reported that members of the Branchiostoma species have similar morphological phenotypic characteristics, but in contrast, there are high levels of genetic polymorphisms in the populations. Here, we resequenced 20 Branchiostomabelcheri genomes to an average depth of approximately 12.5X using the Illumina HiSeq 2000 platform. In this study, over 52 million variations (~12% of the total genome) were detected in the B. belcheri population, and an average of 12.8 million variations (~3% of the total genome) were detected in each individual, confirming that Branchiostoma is one of the most genetically diverse species sequenced to date. Demographic inference analysis highlighted the role of historical global temperature in the long-term population dynamics of Branchiostoma, and revealed a population expansion at the Greenlandian stage of the current geological epoch. We detected 594 Single nucleotide polymorphism and 148 Indels in the Branchiostoma mitochondrial genome, and further analyzed their genetic mutations. A recent study found that the epithelial cells of the digestive tract in Branchiostoma can directly phagocytize food particles and convert them into absorbable nontoxic nutrients using powerful digestive and immune gene groups. In this study, we predicted all potential mutations in intracellular digestion-associated genes. The results showed that most "probably damaging" mutations were related to rare variants (MAF<0.05) involved in strengthening or weakening the intracellular digestive capacity of Branchiostoma. Due to the extremely high number of polymorphisms in the Branchiostoma genome, our analysis with a depth of approximately 12.5X can only be considered a preliminary analysis. However, the novel variant dataset provided here is a valuable resource for further investigation of phagocytic intracellular digestion in Branchiostoma and determination of the phenotypic and genotypic features of Branchiostoma.

Project description:Among chordate taxa, the cephalochordates diverged earlier than urochordates and vertebrates; thus, they retain unique, primitive developmental features. In particular, the amphioxus notochord has muscle-like properties, a feature not seen in urochordates or vertebrates. Amphioxus contains two Brachyury genes, Bra1 and Bra2. Bra2 is reportedly expressed in the blastopore, notochord, somites, and tail bud, in contrast to a low level of Bra1 expression only in notochord. To distinguish the expression profiles of the two Brachyury genes at the single-cell level, we carried out single-cell RNA-seq (scRNA-seq) analysis using the amphioxus, Branchiostoma japonicum. This scRNA-seq analysis classified B. japonicum embryonic cells into 15 clusters at developmental stages from midgastrula to early swimming larva. Brachyury was expressed in cells of clusters 4, 5, 8, and 9. We first confirmed that cluster 8 comprises cells that form somites since this cluster specifically expresses four myogenic factor genes. Cluster 9 contains a larger number of cells with high levels of Bra2 expression and a smaller number of cells with Bra1 expression. Simultaneous expression in cluster 9 of tool-kit genes, including FoxA, Goosecoid, and hedgehog, showed that this cluster comprises cells that form the notochord. Expression of Bra2, but not Bra1, in cells of clusters 4 and 5 at the gastrula stage together with expression of Wnt1 and Caudal indicates that clusters 4 and 5 comprise cells of the blastopore, which contiguously form the tail bud. In addition, Hox1, Hox3, and Hox4 were highly expressed in Bra2-expressing clusters 4, 5, 8, and 9 in a temporally coordinated manner, suggesting roles of anterior Hox genes in specification of mesodermal organs, including somites, notochord, and tail bud. This scRNA-seq analysis therefore highlights differences between the two Brachyury genes in relation to embryonic regions in which they are expressed and their levels of expression. Bra2 is the ancestral Brachyury in amphioxus, since expression in the blastopore is shared with other deuterostomes. On the other hand, Bra1 is a duplicate copy and likely evolved a supplementary function in notochord and somite formation in the Branchiostoma lineage.

Dataset Information

Genome-wide gene expression analysis of amphioxus (Branchiostoma belcheri) following lipopolysaccharide challenge using strand-specific RNA-seq.

Publications

Genome-wide gene expression analysis of amphioxus (Branchiostoma belcheri) following lipopolysaccharide challenge using strand-specific RNA-seq.

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