Dominik Hartl, Arnaud R. Krebs, Josephine Jüttner, Botond Roska, Dirk Schübeler
The retina is composed of ∼50 cell-types with specific functions for the process of vision. Identification of the cis-regulatory elements active in retinal cell-types is key to elucidate the networks controlling this diversity. Here, we combined transcriptome and epigenome profiling to map the regulatory landscape of four cell-types isolated from mouse retinas including rod and cone photoreceptors as well as rare inter-neuron populations such as horizontal and starburst amacrine cells. Integration of this information reveals sequence determinants and candidate transcription factors for controlling cellular specialization. Additionally, we refined parallel reporter assays to enable studying the transcriptional activity of large collection of sequences in individual cell-types isolated from a tissue. We provide proof of concept for this approach and its scalability by characterizing the transcriptional capacity of several hundred putative regulatory sequences within individual retinal cell-types. This generates a catalogue of cis-regulatory regions active in retinal cell types and we further demonstrate their utility as potential resource for cellular tagging and manipulation.
Dominik Hartl, Arnaud R. Krebs, Josephine Jüttner, Botond Roska, Dirk Schübeler. Cis-regulatory landscapes of four cell types of the retina. 2017-10-20. https://doi.org/10.1093/nar/gkx923
The cone/rod library tested candidate regulatory fragments selected from differential methylation patterns in adult mouse cones and rods. The same library was assayed in three cone and three rod biological replicates, and the table retains the published activity ratios and element-level specificity calls for both cell types.
The pilot library tested 384 short candidate cis-regulatory fragments, including highly methylated negative controls, in adult mouse rod photoreceptors in vivo. Three biological replicates of FACS-isolated rods were profiled by barcode RNA sequencing and normalized to barcode abundance in the input AAV pool.
The motif-mutant library tested wild-type and randomized transcription-factor motif versions of two active photoreceptor CREs, Faim-Intra (e107) and Intergenic3 (e483), in adult mouse rods. Four rod biological replicates were measured by barcode RNA sequencing relative to AAV-input barcode abundance, enabling direct mutant-versus-wild-type activity comparisons.
The third library combined five photoreceptor reference fragments with 365 candidate elements selected for hypomethylation in horizontal cells and/or starburst amacrine cells. The library was measured in three biological replicates each of FACS-isolated SBACs and horizontal cells, with three whole-retina RNA readouts retained as a comparator.
Original supplemental and deposited inputs retained for this study. Download files individually or together as a ZIP; nested folders are preserved. Source reuse terms apply, and sequencing reads may be omitted.
Download all 14 files (ZIP)gkx923_supp.zipGSE84589_BCtoPr_Lib1.tab.gzGSE84589_BCtoPr_Lib2.tab.gzGSE84589_BCtoPr_Lib3.tab.gzGSE84589_family.soft.gzGSE84589_Lib1Results.tab.gzGSE84589_Lib2Results.tab.gzGSE84589_Lib3Results.tab.gzGSE84589_Lib4Results.tab.gzGSE84589_LMR_methylation_summary_four_cell_types.tab.gzsupplementary_figures_and_sequences.pdfsupplementary_Lib1_results_with_methylation.tabsupplementary_Lib2_results_with_methylation.tabsupplementary_Lib3_results_with_methylation.tab