BiT-STARR-seq allele-specific regulatory activity in HUVECs with caffeine versus water control
Characterization of caffeine response regulatory variants in vascular endothelial cellsAn episomal BiT-STARR-seq library tested 43,556 SNP-centered 200-nt regulatory targets, with both alleles represented in forward and reverse construct orientations. HUVECs received 1.16 × 10^-3 M caffeine or water vehicle after transfection, with six biological replicates per condition; the table summarizes condition-specific ASE and caffeine-versus-water cASE results.
Processed tables are specific to each experiment. Column names, units, measurements, and table structure are not standardized across the database. Check this experiment’s column definitions and quality-control notes before comparing or combining data.
Perturbation & assay details
1.16 × 10^-3 M caffeine for 24 h versus water vehicle control
The common library contained 43,556 SNP-centered target regions, 87,112 allele-specific constructs, and up to two construct orientations per allele; 1,676 computational negative-control sequences were included. DNA library was transfected into Lonza HUVECs with the Lonza Nucleofector X platform (DS-120, P5 solution), caffeine was added after transfection, cells were incubated 24 h, and RNA was sequenced on an Illumina NextSeq 500. Reads were UMI-filtered for the RDHBVDHBVD pattern, deduplicated, aligned to hg19, and converted to allele/direction counts. QuASAR-MPRA was used for condition-specific ASE and the laboratory's Delta-AST method for cASE.
BiT-STARR-seq (Biallelic Targeted STARR-seq): an episomal, oligo-synthesized self-transcribing reporter assay in which 200-nt SNP-centered inserts were synthesized for reference and alternate alleles and tested in forward and reverse orientations. Reporter RNA and input DNA were sequenced, and allele-specific effects were estimated from allele counts while accounting for DNA-library imbalance.
Processed data
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Filters apply to this table only. The CSV download contains the complete processed table; filtered rows are available through the API.
Column dictionary · 45 definitions
- identifier
- Original study SNP/direction-pair identifier, formatted as rsID_fw or rsID_rv.
- variant_id
- The study rsID extracted from identifier.
- direction
- Reporter construct orientation label from the study: fw or rv.
- chromosome
- Primary hg19 chromosome for the rsID, when resolved.
- position_hg19_0based
- 0-based hg19 genomic position, when resolved.
- position_hg19_1based
- 1-based hg19 genomic position, when resolved.
- coordinate_source
- Source used for the added hg19 coordinate: the older Zenodo S4 coordinate bridge, Ensembl GRCh37 variation mapping, or unresolved.
- geo_count_coordinate_match
- Whether at least one GEO-deposited allele-count row occurs at the resolved chromosome and 1-based position.
- grch37_allele_string
- Ensembl GRCh37 allele string when available; it is not a re-called allele assignment for the MPRA.
- geo_ref_alleles_observed
- Distinct non-dot REF strings observed in the twelve GEO count files at the resolved position.
- geo_alt_alleles_observed
- Distinct non-dot ALT strings observed in the twelve GEO count files at the resolved position; may be NA when no alternate read was observed.
- geo_count_rows_at_coordinate
- Number of GEO count rows across all twelve libraries at the resolved position.
- jaspar_motif_count
- Number of distinct JASPAR PWMScan motif IDs overlapping the rsID in the study's motif annotation.
- jaspar_motif_ids
- Semicolon-separated JASPAR PWMScan motif IDs overlapping the rsID; NA means no motif annotation in supplementary file 2.
- caffeine_effect_estimate
- Published QuASAR-MPRA meta_estimate for the caffeine condition.
- caffeine_effect_se
- Published standard error of the caffeine ASE effect estimate.
- caffeine_replicates_with_nonzero_counts
- Number of the six caffeine replicates containing nonzero allele counts (published n.x).
- caffeine_dna_reference_proportion
- Published reference-allele proportion in the DNA library for caffeine analysis.
- caffeine_z
- Published caffeine ASE meta z-score.
- caffeine_p_value
- Published nominal caffeine ASE p-value.
- caffeine_ase_fdr
- Published Benjamini–Hochberg-adjusted caffeine ASE p-value (meta_padj.x).
- published_new_padj
- Published new_padj field from final supplementary file 4, retained without recomputation.
- caffeine_ase_fdr10_significant
- true when caffeine_ase_fdr < 0.10; otherwise false.
- water_effect_estimate
- Published QuASAR-MPRA meta_estimate for the water vehicle condition.
- water_effect_se
- Published standard error of the water ASE effect estimate.
- water_replicates_with_nonzero_counts
- Number of the six water replicates containing nonzero allele counts (published n.y).
- water_dna_reference_proportion
- Published reference-allele proportion in the DNA library for water analysis.
- water_z
- Published water ASE meta z-score.
- water_p_value
- Published nominal water ASE p-value.
- water_ase_fdr
- Published Benjamini–Hochberg-adjusted water ASE p-value (meta_padj.y).
- water_ase_fdr10_significant
- true when water_ase_fdr < 0.10; otherwise false.
- conditional_ase_z
- Published caffeine-versus-water conditional ASE Delta-Z statistic (case_z).
- conditional_ase_p_value
- Published nominal conditional ASE p-value (case_p).
- conditional_ase_fdr
- Published Benjamini–Hochberg-adjusted conditional ASE p-value (case_padj).
- conditional_ase_fdr5_significant
- true when conditional_ase_fdr < 0.05; otherwise false.
- target_base_mean
- Author-provided DESeq2 normalized mean activity for the coordinate-matched target from supplementary file 1.
- target_activity_log2_fold_change
- Author-provided DESeq2 caffeine-versus-control log2 fold change for the coordinate-matched target.
- target_activity_lfc_se
- Author-provided standard error of the target activity log2 fold change.
- target_activity_stat
- Author-provided DESeq2 test statistic for target activity.
- target_activity_p_value
- Author-provided nominal p-value for target activity.
- target_activity_fdr
- Author-provided Benjamini–Hochberg-adjusted p-value for target activity.
- target_activity_fdr10_significant
- true when the coordinate-matched target activity FDR is <0.10; NA means no unambiguous supplementary-file-1 coordinate join.
- qc_pass_caffeine_4of6
- true when the caffeine identifier has n.x >= 4; this is the paper's replicate-support threshold.
- qc_pass_water_4of6
- true when the water identifier has n.y >= 4; this is the paper's replicate-support threshold.
- qc_pass_any_condition
- Row-level inclusion flag; true when at least one condition passes the 4-of-6 replicate-support threshold.
Quality control
The authors demultiplexed with bcl2fastq, aligned to hg19 with HISAT2, removed reads with short or non-matching RDHBVDHBVD UMIs, deduplicated with UMItools, and generated allele counts with samtools mpileup followed by bcftools query. QuASAR-MPRA modeled allelic imbalance using the DNA-library allele proportion; six replicates were available per condition. The study required each identifier to have nonzero counts in at least 4 of 6 replicates before multiple-testing correction, used Benjamini–Hochberg FDR <10% for ASE and <5% for cASE, and tested 23,814 SNP/direction pairs with both conditions meeting the replicate threshold for cASE. table.csv excludes rows failing the 4-of-6 threshold in both conditions; rows passing only one condition are retained for that valid condition and have NA cASE fields.
Curation notes
This is one combined MPRA experiment because the paper used one shared BiT-STARR-seq library and analyzed six caffeine and six water HUVEC libraries as condition-specific ASE and cASE contrasts; the two library batches are represented by BST5 and BST11 sample prefixes. The final eLife supplementary file 4 is the authoritative source for all effect, p-value, and FDR columns in table.csv. It does not include genomic coordinates, so coordinates are added by rsID using the retained 2022 Zenodo preprint S4 bridge when available and Ensembl's GRCh37 variation mapping for remaining primary-assembly rsIDs; alternate-patch mappings and six unresolved rsIDs are left NA. GEO REF/ALT columns are explicitly labeled observations from deposited count rows and are not re-called variant alleles. Supplementary-file-1 target activity is joined by exact hg19 coordinate and is therefore an annotation rather than a direction-resolved reanalysis. The GEO family metadata and filenames identify the batch-2 libraries as BST11HC1/BST11HW1, whereas the author supplementary QC workbook labels the corresponding rows BST11HC4/BST11HW4; the GEO identifiers are retained as authoritative. The package retains the final eLife supplementary tables and the post-UMI-filter/deduplication GEO allele-count outputs, but not raw sequencing reads.