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With their complex life cycle and highly derived body plan, echinoderms are unique among bilaterians. Although early development has been intensively studied, the molecular mechanisms underlying development of the adult echinoderm and its unusual radial body plan are largely unknown. To investigate the evolution of developmental changes in gene expression underlying radial body plan development and metamorphosis, we assembled a reference transcriptome de novo and used RNA-seq to measure gene expression profiles across larval, metamorphic, and postmetamorphic life cycle phases in the sea urchin Heliocidaris erythrogramma. Our results present a high-resolution view of gene expression dynamics during the complex transition from pre- to postmetamorphic development and suggest that distinct sets of regulatory and effector proteins are used during different life history phases. These analyses provide an important foundation for more detailed analyses of the evolution of the radial adult body of echinoderms.
Fig. 1.â. Developmental transcriptome in Heliocidaris erythrogramma. (A) Our developmental time-course included seven stages spanning premetamorphic (24, 28, 32, 36, and 40 hpf) and postmetamorphic development (96 hpf and 10 dpf). (B) PC analysis of gene expression averaged across biological replicates. PC1 explains 72% of the overall variation and clearly separates pre- from postmetamorphic stages, whereas PC2 explains 18% of the variation. The gray line is superimposed to show that overall variation in gene expression between stages recapitulates our developmental time-course.
Fig. 2.â. Metamorphic and postmetamorphic life cycle stages are characterized by dynamic gene expression patterns. (A) MA plot showing the average expression level (in counts per million) of all 13,017 genes at 40 hpf and the log2 fold-change of each gene from 40 to 96 hpf. Genes with a significant decrease in expression at an FDR of 5% are green, whereas genes with a significant increase in expression are blue. Nonsignificant genes are in gray. Horizontal black bars indicate genes that exhibited at least a 4-fold change in expression (log2FC = 2). (B) Kernel density curves of the expression of genes that significantly decreased (green), increased (blue) or did not change (gray) from 40 to 96 hpf at an FDR of 5%. (C) MA plot showing the average expression level at 96 hpf and the log2FC of each gene from 96 to 10 dpf. Genes with a significant decrease in expression at an FDR of 5% are blue, whereas genes with a significant increase in expression are red. Nonsignificant genes are in gray. (D) Kernel density curves of the expression of genes that significantly decreased (blue), increased (red) or did not change (gray) from 96 hpf to 10 dpf at an FDR of 5%.
Fig. 3.â. Advanced rudiment, early juvenile, and late juveniles exhibit stage-specific enrichment of PANTHER biological process categories. (A) Bubble plot showing significantly enriched PANTHER biological process categories at 40 hpf (green) compared with 96 hpf (blue) at an FDR of 5%. Significant categories were identified by a hypergeometric test for the top 10% of a probability distribution. For each category, significance is represented by bubble color and occupancy by bubble size. For clarity, only categories with at least 50 members are shown (see supplementary tables S1 and S2, Supplementary Material online, for additional categories). (B) Bubble plot showing significantly enriched categories at 96 hpf (blue) compared with 10 dpf (red) at an FDR of 5% (see supplementary tables S3 and S4, Supplementary Material online, for additional categories).
Fig. 4.â. Fuzzy c-means clustering identifies general patterns of gene expression across development. To illustrate major patterns of gene expression, five clusters detected by fuzzy c-means clustering of the average expression value of 4,421 genes (log2FC ⥠2, FDR ⤠0.001) are shown (see supplementary fig. S1, Supplementary Material online, for additional clusters). The ms of a given gene within a cluster is represented by color, with red (ms = 1) indicating high association. For each cluster, significantly enriched PANTHER biological process categories identified by a Wilcoxon signed rank test are shown. For clarity, only the top ten categories are displayed (see supplementary table S5, Supplementary Material online, for additional categories).
Fig. 5.â. Expression of neural transcription factors during juvenile development. (A) Several larval neurogenesis genes are also expressed during juvenile development and generally exhibit a premetamorphic decrease, premetamorphic increase, or a postmetamorphic peak of expression. Each line represents the average expression value (in fragments per kilobase of transcript per million mapped reads) for an individual gene. (B) Juvenile-specific neurogenesis transcription factors exhibit either a premetamorphic increase or a postmetamorphic peak of expression.
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