Study / S18AXUTON2018-11-19
A massively parallel reporter assay dissects the influence of chromatin structure on cis-regulatory activity
Brett B. Maricque, Hemangi G. Chaudhari, Barak A. Cohen
About this study
A gene’s position in the genome can profoundly affect its expression because regional differences in chromatin modulate the activity of locally acting cis-regulatory sequences (CRSs). Here we study how CRSs and regional chromatin act in concert on a genome-wide scale. We present a massively parallel reporter gene assay that measures the activities of hundreds of different CRSs, each integrated at many specific genomic locations. Although genome location strongly affected CRS activity, the relative strengths of CRSs were maintained at all chromosomal locations. The intrinsic activities of CRSs also correlated with their activities in plasmid-based assays. We explain our data with a quantitative model in which expression levels are set by independent contributions from local CRSs and the regional chromatin environment, rather than by more complex sequence- or protein-specific interactions between these two factors. The methods we present will help investigators determine when regulatory information is integrated in a modular fashion and when regulatory sequences interact in more complex ways.
Full author list & citation
Brett B. Maricque, Hemangi G. Chaudhari, Barak A. Cohen. A massively parallel reporter assay dissects the influence of chromatin structure on cis-regulatory activity. 2018-11-19. https://doi.org/10.1038/nbt.4285
Experiments 2
E324229C1
A barcoded library of 310 regulatory constructs (282 CRSs plus 28 basal controls), with each CRS represented by 25 distinct CRS barcodes, was integrated by Cre-lox cassette exchange into eight mapped K562 landing-pad clones. The table reports the published log2 RNA/DNA expression measurements for each construct across two biological replicates and eight genomic landing pads, together with prior episomal MPRA expression and ENCODE-derived CRS class.
E62EE0D93
A second integrated patchMPRA library tested 24 regulatory sequences, including 13 literature-validated CRSs and 11 CRSs from Library 1, with Hsp68 and MinP promoter variants across 14 K562 landing-pad locations. The table reports published log2(RNA/DNA) expression values for each CRS–landing-pad–promoter combination across three biological replicates, joined to the source library-composition annotations.