Alan Y. Du, Xiaoyu Zhuo, Vasavi Sundaram, Nicholas O. Jensen, Hemangi G. Chaudhari et al.
Many transposable elements (TEs) contain transcription factor binding sites and are implicated as potential regulatory elements. However, TEs are rarely functionally tested for regulatory activity, which in turn limits our understanding of how TE regulatory activity has evolved. We systematically tested the human LTR18A subfamily for regulatory activity using massively parallel reporter assay (MPRA) and found AP-1- and CEBP-related binding motifs as drivers of enhancer activity. Functional analysis of evolutionarily reconstructed ancestral sequences revealed that LTR18A elements have generally lost regulatory activity over time through sequence changes, with the largest effects occurring owing to mutations in the AP-1 and CEBP motifs. We observed that the two motifs are conserved at higher rates than expected based on neutral evolution. Finally, we identified LTR18A elements as potential enhancers in the human genome, primarily in epithelial cells. Together, our results provide a model for the origin, evolution, and co-option of TE-derived regulatory elements.
Alan Y. Du, Xiaoyu Zhuo, Vasavi Sundaram, Nicholas O. Jensen, Hemangi G. Chaudhari, Nancy L. Saccone, Barak A. Cohen, Ting Wang. Functional characterization of enhancer activity during a long terminal repeat's evolution. 2022-10-03. https://doi.org/10.1101/gr.276863.122
The shared 59,470-barcode library represented motif-focused and 160-bp tiled LTR18A fragments, ancestral reconstructions, reverse complements, motif mutants, and controls; it was transfected into human K562 chronic myelogenous leukemia cells. Three biological transfections were measured by RNA/DNA barcode enrichment normalized to the basal hsp68 promoter.
The shared 59,470-barcode library represented motif-focused and 160-bp tiled LTR18A fragments, ancestral reconstructions, reverse complements, motif mutants, and controls; it was transfected into human HepG2 liver carcinoma cells. Three biological transfections were measured by RNA/DNA barcode enrichment normalized to the basal hsp68 promoter.
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 20 files (ZIP)geo_barcode_counts/GSM6050128_HepG2_run1_BC_counts.txt.gzgeo_barcode_counts/GSM6050129_HepG2_run2_BC_counts.txt.gzgeo_barcode_counts/GSM6050130_HepG2_run3_BC_counts.txt.gzgeo_barcode_counts/GSM6050131_HepG2_run4_BC_counts.txt.gzgeo_barcode_counts/GSM6050132_K562_run1_BC_counts.txt.gzgeo_barcode_counts/GSM6050133_K562_run2_BC_counts.txt.gzgeo_barcode_counts/GSM6050134_K562_run3_BC_counts.txt.gzgeo_barcode_counts/GSM6050135_K562_run4_BC_counts.txt.gzgeo_barcode_counts/GSM6050136_input_DNA_run1_BC_counts.txt.gzgeo_barcode_counts/GSM6050137_input_DNA_run2_BC_counts.txt.gzgeo_barcode_counts/GSM6050138_input_DNA_run3_BC_counts.txt.gzGSE201068_family.soft.gzGSE201068_HepG2_enrichment_scores.txt.gzGSE201068_K562_enrichment_scores.txt.gzsource_notes.txtsupp_gr.276863.122_Supplemental_Data_S1.zipsupp_gr.276863.122_Supplemental_Methods.pdfsupplemental_data_s1/HepG2_enrichment_scores.tsvsupplemental_data_s1/K562_enrichment_scores.tsvsupplemental_data_s1/MPRA_library_design.tsv