A chromosome-scale epigenetic map of theHydragenome reveals conserved regulators of cell state

Abstract

The epithelial and interstitial stem cells of the freshwater polypHydraare the best characterized stem cell systems in any cnidarian, providing valuable insight into cell type evolution and the origin of stemness in animals. However, little is known about the transcriptional regulatory mechanisms that determine how these stem cells are maintained and how they give rise to their diverse differentiated progeny. To address such questions, a thorough understanding of transcriptional regulation inHydrais needed. To this end, we generated extensive new resources for characterizing transcriptional regulation inHydra, including new genome assemblies forHydra oligactisand the AEP strain ofHydra vulgaris, an updated whole-animal single-cell RNA-seq atlas, and genome-wide maps of chromatin interactions, chromatin accessibility, sequence conservation, and histone modifications. These data revealed the existence of large kilobase-scale chromatin interaction domains in theHydragenome that contain transcriptionally co-regulated genes. We also uncovered the transcriptomic profiles of two previously molecularly uncharacterized cell types, isorhiza-containing nematocytes and somatic gonad ectoderm. Finally, we identified novel candidate regulators of cell-type-specific transcription, several of which have likely been conserved at least since the divergence ofHydraand the jellyfishClytia hemisphaericaover 400 million years ago.

Differentiation trajectories of theHydranervous system reveal transcriptional regulators of neuronal fate

“…Peak accessibility near some neuronal subtypespecific genes varied between samples generated from different transgenic lines but reflected the cell types collected from those transgenic lines (Fig S8). Notably, these data identified accessible regions near neuronal genes that were not detected in our previously collected whole-animal data (Cazet et al, 2023), which demonstrates the utility of these data for identifying the regulatory regions that drive neuronal expression (Figure 3C-H).…”

“…We combined our new data with the neuronal single-cell transcriptomes we previously collected (Siebert et al, 2019) for downstream processing (Table S2). For all libraries, the sequencing reads were mapped to the Hydra vulgaris strain AEP gene models (Cazet et al, 2023) and processed following standard procedures (Fig S3) (Macosko et al, 2015). After filtering, we recovered 35,071 single-cell neural transcriptomes with a detected median of 1371.5 genes and 2887.5 UMIs per cell (Table S2).…”

“…10x Genomics Single Cell RNA Sequencing and Processing Sorted transgenic Hydra cells were processed for sequencing using the Chromium Next GEM Single Cell 3' kit v3.1 (10x Genomics) according to manufacturer's instructions at the University of California, Davis Sequencing Core Facility. Data were aligned to the AEP genome assembly (Cazet et al, 2023) and were analyzed using previously described pipelines (Satija et al, 2015;Siebert et al, 2019). See Supplemental Material for details.…”

“…Gene expression values are depicted by average expression across all cells (dot color) and percent expression within each cluster (dot size). Putative TFs were identified using PFAM annotations (data from (Cazet et al, 2023;Siebert et al, 2019). A total of six neuron-enriched ATAC-seq libraries were generated for this study: two from the Tg(tba1c:mNeonGreen) cj1-gt transgenic line and four from the Tg(actin1:GFP) rs3-in transgenic line (Keramidioti et al, 2023).…”

“…In both comparisons, there is significant enrichment (p < 0.001, t-test) of peaks within 10,000 base pairs of the transcription start site of neuronal genes as compared to non-neuronal genes. data set (Cazet et al, 2023;Siebert et al, 2019). (D,E) Alpha tubulin has enriched expression in all neurons as compared to non-neuronal cell types.…”