Siqi Zhao, Clarice K. Y. Hong, Connie A. Myers, David M. Granas, Michael A. White et al.
Massively parallel reporter gene assays are key tools in regulatory genomics, but cannot be used to identify cell-type specific regulatory elements without performing assays serially across different cell types. To address this problem, we developed a single-cell massively parallel reporter assay (scMPRA) to measure the activity of libraries of cis-regulatory sequences (CRSs) across multiple cell-types simultaneously. We assayed a library of core promoters in a mixture of HEK293 and K562 cells and showed that scMPRA is a reproducible, highly parallel, single-cell reporter gene assay that detects cell-type specific cis-regulatory activity. We then measured a library of promoter variants across multiple cell types in live mouse retinas and showed that subtle genetic variants can produce cell-type specific effects on cis-regulatory activity. We anticipate that scMPRA will be widely applicable for studying the role of CRSs across diverse cell types.
Siqi Zhao, Clarice K. Y. Hong, Connie A. Myers, David M. Granas, Michael A. White, Joseph C. Corbo, Barak A. Cohen. A single-cell massively parallel reporter assay detects cell-type-specific gene regulation. 2023-01-12. https://doi.org/10.1038/s41588-022-01278-7
The 676-member core-promoter library was assayed by scMPRA in K562 cells alone across two biological replicates. Author summaries report mean promoter activity across G1, S, and G2/M cell-cycle phases and across CD34+/CD38−, differentiated, and CD24+ substates; these related analyses are combined into one promoter-level table.
The same 676-member core-promoter library was transfected separately into K562 and HEK293 cells for bulk MPRA validation. The table contains author GEO counts for reporter RNA and input plasmid DNA plus the author-provided log2 RNA/DNA activity score for each cell line.
A 676-member library of 133-bp core promoters plus four synthetic controls was transfected into HEK293 and K562 cells, mixed 1:1, and read out by 10x single-cell RNA sequencing with cBC/rBC reporter barcodes. The table contains author mean activities for both cell types across two biological replicates and joined DESeq2 statistics where the promoter gene name was unambiguous.
A 115-member library containing two independently synthesized wild-type Gnb3 promoter controls and 113 targeted promoter variants was introduced into newborn mouse retinas by electroporation and measured by scMPRA in Rod, Bipolar, Glia, and Neuron cells. The table joins full promoter sequences to author mean activities across two biological replicates and derives effects relative to the two-copy wild-type reference in each cell type.
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 18 files (ZIP)GSE188639_k562_bulk_exp.csv.gzGSE188639_mixed_cell_bulk_exp.csv.gzGSE188639_retina_pBC_exp_rep1.csv.gzGSE188639_retina_pBC_exp_rep2.csv.gzk562_cell_cycle_phase.csvk562_cell_state_clustering.csvmixed_cell_identity.csvprom_lib_info_new.tsvREADME.txtscMPRA_readme.mdsupplementary_tables/supplementary_table_1.csvsupplementary_tables/supplementary_table_2.csvsupplementary_tables/supplementary_table_3.csvsupplementary_tables/supplementary_table_4.csvsupplementary_tables/supplementary_table_5.csvsupplementary_tables/supplementary_table_6.csvsupplementary_tables/supplementary_table_7.csvsupplementary_tables.xlsx