Experiment / E82J7WBXUIntegrated lentiMPRA

Chromosomally integrated lentiMPRA of HepG2 candidate liver enhancers (wild-type integrase / WT)

A systematic comparison reveals substantial differences in chromosomal versus episomal encoding of enhancer activity

The 2,440-element 5′ enhancer/3′ barcode library was packaged with wild-type lentiviral integrase and introduced into HepG2 cells for chromosomal integration. Three independent infections were profiled by matched DNA and mRNA barcode sequencing for 2,236 candidate liver enhancers and 204 controls.

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Organism: Human
Taxonomy ID: NCBITaxon:9606
Biosample: CVCL:0027
Reference genome: GRCh37
Design focus: Region-focused
Region of interest: Not reported / not applicable

Perturbation & assay details

Basal / untreated; wild-type-integrase lentiviral delivery

The library used a 5′ 171-bp candidate sequence upstream of a minimal promoter and EGFP, with a 15-bp barcode in the EGFP 3′ UTR; each insert had 100 designed barcodes and the cassette was flanked by antirepressor/SAR elements. HepG2 DNA and mRNA were collected at day 4 after an estimated 50 viral particles per cell; three independent infections were sequenced on Illumina NextSeq.

Processed data

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Column dictionary · 53 definitions

element_id: Published insert/region identifier from GEO ActivityRatios.
design_region_id: Identifier in the oligo design; differs from element_id only for four controls whose activity labels add a bracketed coordinate span.
category: Broad published design category: type1–type4, SRE, or control.
category_detail: Published subtype or control label, such as HNF4A-ChMod, positive, or negative.
element_sequence: The intended 171-bp enhancer or control sequence extracted from the designed 230-bp oligo.
sequence_length_bp: Length of element_sequence in base pairs.
coordinate_assembly: Assembly label parsed from the published identifier; candidate regions use GRCh37 and controls preserve hg18, hg19, or mm9 labels.
coordinate: High-level target interval parsed from the published region identifier.
chromosome: Chromosome parsed from coordinate.
start: 1-based target interval start parsed from coordinate.
end: Target interval end parsed from coordinate.
barcode_count_designed: Number of designed 15-bp reporter barcodes assigned to the insert; the library design used 100.
activity_rep1_rna_dna: Published wild-type-integrase/integrated RNA/DNA activity ratio for replicate 1.
activity_rep2_rna_dna: Published wild-type-integrase/integrated RNA/DNA activity ratio for replicate 2.
activity_rep3_rna_dna: Published wild-type-integrase/integrated RNA/DNA activity ratio for replicate 3.
activity_combined_rna_dna: Published WT activity ratio combined across replicates after replicate-median normalization.
activity_log2_rna_dna: log2 of activity_combined_rna_dna, derived in this package.
replicate_activity_mean_rna_dna: Arithmetic mean of the three published WT replicate ratios; descriptive and not the authors’ combined value.
replicate_activity_sd_rna_dna: Sample standard deviation of the three published WT replicate ratios.
barcodes_common_rep1: Designed barcode IDs observed in both DNA and RNA count files for WT replicate 1.
barcodes_common_rep2: Designed barcode IDs observed in both DNA and RNA count files for WT replicate 2.
barcodes_common_rep3: Designed barcode IDs observed in both DNA and RNA count files for WT replicate 3.
barcodes_common_mean: Arithmetic mean of the three shared RNA/DNA barcode counts.
dna_counts_common_rep1: Sum of assigned DNA counts over barcodes shared by RNA and DNA in WT replicate 1.
dna_counts_common_rep2: Sum of assigned DNA counts over barcodes shared by RNA and DNA in WT replicate 2.
dna_counts_common_rep3: Sum of assigned DNA counts over barcodes shared by RNA and DNA in WT replicate 3.
rna_counts_common_rep1: Sum of assigned RNA counts over barcodes shared by RNA and DNA in WT replicate 1.
rna_counts_common_rep2: Sum of assigned RNA counts over barcodes shared by RNA and DNA in WT replicate 2.
rna_counts_common_rep3: Sum of assigned RNA counts over barcodes shared by RNA and DNA in WT replicate 3.
paired_other_context_activity_combined_rna_dna: Published combined MT/nonintegrating RNA/DNA activity ratio for the paired context.
integrated_minus_episomal_log2_ratio: log2(WT combined activity / MT combined activity); positive values favor the integrated context.
qc_pass: TRUE for rows retained after package-level design, activity, and shared-barcode QC.
OC: Majority HepG2 OpenChromatin state annotation.
SegWay: Majority HepG2 SegWay genome-segmentation annotation.
ChromHMM7: Majority HepG2 7-state ChromHMM annotation.
ChromHMM15: Majority Roadmap HepG2 15-state ChromHMM annotation.
ERBFeature: Majority Ensembl Regulatory Build regulatory-feature annotation.
DHS: Number of element bases overlapping HepG2 DNase hypersensitivity calls.
H3K27ac: Number of element bases covered by HepG2 H3K27ac ChIP-seq peaks.
H3K27acAve: Average HepG2 H3K27ac peak signal over the element.
H3K27acMax: Maximum HepG2 H3K27ac peak signal over the element.
TFcount: Number of distinct assayed transcription factors with ChIP-seq peak overlap.
EP300: Number of element bases covered by HepG2 EP300 ChIP-seq peaks.
FOXA1: Number of element bases covered by HepG2 FOXA1 ChIP-seq peaks.
FOXA2: Number of element bases covered by HepG2 FOXA2 ChIP-seq peaks.
HNF4A: Number of element bases covered by HepG2 HNF4A ChIP-seq peaks.
RAD21: Number of element bases covered by HepG2 RAD21 ChIP-seq peaks.
CHD2: Number of element bases covered by HepG2 CHD2 ChIP-seq peaks.
SMC3: Number of element bases covered by HepG2 SMC3 ChIP-seq peaks.
GC: CADD v1.3 average local GC percentage annotation.
CpG: CADD v1.3 average local CpG percentage annotation.
minDistTSS: CADD v1.3 distance to the closest transcribed-sequence start site.
CADD: CADD v1.3 PHRED-scaled sequence annotation.

Quality control

Author QC was retained: 15-bp barcode consensus sequences were matched to the designed barcode set without clustering; only barcodes observed in both RNA and DNA in the same sample were used; counts were normalized to counts per million; per-insert RNA/DNA ratios were calculated from summed barcode counts; replicate ratios were divided by their replicate median before averaging. Package-level integrity QC additionally required an exact design match, 100 designed barcodes, positive numeric activity in all three replicates, and at least 3 shared RNA/DNA barcodes per replicate. All 2,440/2,440 inserts passed; the minimum shared-barcode count was 9, 9, and 8 across the three WT replicates.

Curation notes

This is the wild-type-integrase arm of a paired WT/MT lentiMPRA and represents random chromosomal integration of the reporter library. The study is region-focused rather than variant-focused: it tests annotated 171-bp candidate liver enhancers and controls, with no designed allele contrasts. ActivityRatios contains all 2,440 inserts; source values marked NA in selected annotation fields are emitted as blank cells, while the primary activity fields are complete. The four endogenous/SRE controls include hg19, hg18, or mm9 labels in their identifiers and are retained as controls.

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