Fig. 4: Key regulators and transcriptional network of aestivation in A. japonicus.

a Identification of differentially expressed genes (DEGs) and differentially expressed transcriptional factors (DE-TFs) in four organs (body wall, muscle, respiratory tree and intestine) during different states of aestivation. Venn diagrams and histograms show the shared gene numbers between organs and the absolute gene numbers in each organ. Among the four organs, body wall shows the most DEGs and DE-TFs, representing the most responsive organ during sea cucumber aestivation. b Expression profiles of nine TFs showing differential expression during aestivation in all four organs. Compared with other TFs, Klf2 and Egr1 are the most significant TFs, especially in body wall, likely playing important roles in the regulation of aestivation. Aestivation states: non-aestivation (Non_aes); early aestivation (Early_aes); deep aestivation (aes); and arousal from aestivation (Aro). c Expression heatmap of Klf2, Egr1 and clock-related genes during different aestivation states (according to quantitative PCR results), and the inferred clock gene-controlled regulation model. Klf2 and Egr1 may trigger the upregulation of Cry1 (either directly or indirectly through Clock and Bmal1) during sea cucumber aestivation, which propels the animal into an extended sleep phase, and decreased Cry1 expression makes the animal awaken from aestivation. Aestivation states are the same as depicted in (b) except Pre_aro representing initial arousal from aestivation. d Co-expression TF network of the aestivation-responsive model AM7. Klf2 and Egr1 are recognized as hub transcription factors in the network. The TFs showing differential expression in all organs are labeled in red, whereas for the remaining DE-TFs showing differential expression in at least one organ are labeled in yellow. e KEGG enrichment analysis of the AM7 module. The AM7 module governs diverse gene pathways, including those participating in cell proliferation and differentiation, seasonal rhythmicity and immune responses, suggesting the complex mechanism of molecular regulation during sea cucumber aestivation. The circle size and filled portion represent the gene numbers (from the AM7 module) and percentage of differentially expressed genes (DGEs) in a given pathway, respectively. The statistical significance is colored according to Q values