Brian S. Roberts, E. Christopher Partridge, Bryan A. Moyers, Vikram Agarwal, Kimberly M. Newberry et al.
Massively parallel reporter assays (MPRAs) are useful tools to characterize regulatory elements in human genomes. An aspect of MPRAs that is not typically the focus of analysis is their intrinsic ability to differentiate activity levels for a given sequence element when placed in both of its possible orientations relative to the reporter construct. Here, we describe pervasive strand asymmetry of MPRA signals in data sets from multiple reporter configurations in both published and newly reported data. These effects are reproducible across different cell types and in different treatments within a cell type and are observed both within and outside of annotated regulatory elements. From elements in gene bodies, MPRA strand asymmetry favors the sense strand, suggesting that function related to endogenous transcription is driving the phenomenon. Similarly, we find that within Alu mobile element insertions, strand asymmetry favors the transcribed strand of the ancestral retrotransposon. The effect is consistent across the multiplicity of Alu elements in human genomes and is more pronounced in less diverged Alu elements. We find sequence features driving MPRA strand asymmetry and show its prediction from sequence alone. We see some evidence for RNA stabilization and transcriptional activation mechanisms and hypothesize that the effect is driven by natural selection favoring efficient transcription. Our results indicate that strand asymmetry is a pervasive and reproducible feature in MPRA data. More importantly, the fact that MPRA asymmetry favors naturally transcribed strands suggests that it stems from preserved biological functions that have a substantial, global impact on gene and genome evolution.
Brian S. Roberts, E. Christopher Partridge, Bryan A. Moyers, Vikram Agarwal, Kimberly M. Newberry, Beth K. Martin, Jay Shendure, Richard M. Myers, Gregory M. Cooper. Genome-wide strand asymmetry in massively parallel reporter activity favors genic strands. 2021-04-20. https://doi.org/10.1101/gr.270751.120
A pooled library of sheared BAC fragments spanning the HTT locus was assayed in human BE(2)-C neuroblastoma cells using an episomal STARR-seq reporter. Plasmid DNA input and reporter-derived RNA were sequenced in three replicate transfections while retaining fragment orientation relative to the GRCh38 reference.
A pooled library of sheared BAC fragments spanning the HTT locus was assayed in human A549 cells using an episomal STARR-seq reporter. Plasmid DNA input and reporter-derived RNA were sequenced in three replicate transfections while retaining fragment orientation relative to the GRCh38 reference.
A pooled library of sheared BAC fragments spanning the HTT locus was assayed in human HepG2 hepatoblastoma cells using an episomal STARR-seq reporter. Plasmid DNA input and reporter-derived RNA were sequenced in three replicate transfections while retaining fragment orientation relative to the GRCh38 reference.
Two BAC-derived fragment libraries spanning the SORT1 locus were assayed in human HepG2 cells with a promoter-less episomal STARR-seq vector. Plasmid DNA input and reporter-derived RNA were sequenced in three replicate transfections while retaining fragment orientation relative to the GRCh38 reference.
A pooled library of sheared BAC fragments spanning the HTT locus was assayed in human K562 myeloid leukemia cells using an episomal STARR-seq reporter. Plasmid DNA input and reporter-derived RNA were sequenced in three replicate transfections while retaining fragment orientation relative to the GRCh38 reference.
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 8 files (ZIP)GSE156857_family.soft.gzGSE156857_filelist.txtGSE156857_HTT_locus_binned_stranded_signal_bedlike.tsv.gzGSE156857_SORT1_locus_binned_stranded_signal_bedlike.tsv.gzREADME.txtsource_manifest.txtsupp_gr.270751.120_Supplemental_FigsTables.pdfsupp_gr.270751.120_Supplemental_Files.zip