Experiment / E5HHP61KIIntegrated lentiMPRA

Integrated lentiMPRA of duplicated CRE tiles in SH-SY5Y

Influence of cis-regulatory elements on expression divergence in human segmental duplications

A shared library of 200-bp tiles from 26 human-specific segmental-duplication gene families, including human ancestral/derived paralog and panTro6 chimpanzee homolog sequences plus scrambled negative controls, was delivered by lentivirus to the SH-SY5Y neuroblastoma cell line. Three technical replicates were quantified from integrated DNA and expressed RNA barcodes with MPRAnalyze; the processed table retains 7,760 construct-level test measurements that pass the available construct and result QC.

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Organism: Human
Taxonomy ID: NCBITaxon:9606
Biosample: CVCL:0019
Reference genome: GRCh38
Design focus: Region-focused
Region of interest: Not reported / not applicable

Perturbation & assay details

Basal / Untreated

Candidate CREs were identified from remapped H3K27ac, ATAC-seq, and DNase-seq peaks in ancestral HSD loci and tiled at 200 bp with 100-bp overlap. Homologous human GRCh38 and chimpanzee panTro6 sequences were synthesized with universal pLS-SceI priming sequences, cloned into a lentiviral reporter, and barcode-associated with MPRAflow. SH-SY5Y cells were infected at approximately MOI 40 in DMEM/F12 GlutaMAX with 10% FBS and 2.5 micrograms/mL protamine sulfate; nucleic acids were extracted 48 hours after infection. Barcode association used a PE150 NextSeq run and DNA/RNA barcode counting used three PE15 NextSeq runs. Activity is the MPRAnalyze estimated transcription rate alpha from integrated DNA and expressed RNA counts.

Processed data

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Visible columns (28 of 28)

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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 · 28 definitions

element_id: Unique published construct/sequence name.
ancestral_coord_grch38: GRCh38 coordinate of the ancestral HSD tile used as the sequence-family anchor.
homolog_coord: One or more homologous coordinates represented by the construct, from Supplemental Table S6; coordinates may be human GRCh38 or chimpanzee panTro6 and are pipe-delimited when shared.
homolog_species: Species represented by the homologous coordinate(s): human, chimpanzee, or human;chimpanzee for a construct shared across species.
homolog_class: Coordinate class relative to the ancestral anchor: human_ancestral, human_derived, chimpanzee_ortholog, or combinations joined with +.
ancestral_coord_strand: Strand of the ancestral GRCh38 coordinate as supplied in Supplemental Table S6.
nearest_feature_strand: Strand assignment from the nearest annotated feature as supplied in Supplemental Table S6.
alignment_size_difference_bp: Difference in original homologous alignment size from the 200-bp target, before trimming or padding.
sequence_200bp: The synthesized 200-bp regulatory sequence for the construct, without the universal priming flanks.
grch38_t2t_chm13_comparison: Supplemental Table S6 comparison of the sequence between GRCh38 and T2T-CHM13, such as identical, different, mixed, or NA.
pantro6_mpantro3_comparison: Supplemental Table S6 comparison of the chimpanzee sequence between panTro6 and mPanTro3 haplotype 1.
construct_type: Published MPRA quantification type; retained rows are test constructs.
target_coordinates: Target coordinate field from the authors' MPRA quantification output.
ancestor_locus_from_quant: Ancestral tile/locus field from the authors' MPRA quantification output.
alpha_activity: MPRAnalyze estimated transcription rate alpha, the primary construct activity measure modeled from DNA and RNA barcode counts.
zscore: MPRAnalyze z-score activity statistic from the published quantification output.
pval_zscore: P-value associated with the z-score activity statistic.
mad_score: Median-absolute-deviation activity score used for testing against scrambled negative controls.
pval_mad: P-value for the median-absolute-deviation activity test against scrambled negative controls.
activity_qvalue: Author-supplied qval field from the MPRA quantification export, retained as provided.
active_vs_scramble: Derived Boolean activity call: true when pval_mad < 0.05, matching the paper's active-sequence threshold.
log2fc_vs_chimpanzee: Published pairwise log2 fold change for this construct relative to the chimpanzee baseline; blank when no exact Supplemental Table S9 row is available.
pval_vs_chimpanzee: Published pairwise p-value for the construct-versus-chimpanzee homolog test.
fdr_vs_chimpanzee: Published FDR for the construct-versus-chimpanzee homolog test.
log2fc_vs_human_ancestral: Published pairwise log2 fold change for this construct relative to the human ancestral homolog; blank when no exact Supplemental Table S11 row is available.
pval_vs_human_ancestral: Published pairwise p-value for the construct-versus-human-ancestral homolog test.
fdr_vs_human_ancestral: Published FDR for the construct-versus-human-ancestral homolog test.
qc_pass: Boolean indicating that the row passed the package-level construct/result QC filter and was retained.

Quality control

The authors used three technical replicates, MPRAflow barcode-insert association with map quality 1 and minimum barcode coverage 10, manual review of promiscuous barcode assignments, total-sum depth normalization, and MPRAnalyze models. Within-cell-type replicate reproducibility was reported as mean pairwise r = 0.93 for both DNA and RNA libraries. The 500 scrambled ancestral tiles served as negative controls; activity was called by the authors at pval.mad < 0.05, yielding 1,056 active sequences in SH-SY5Y. For this package, only published quantification rows with type = test, complete numeric activity fields, an exact 200-bp sequence, and a matching Supplemental Table S6 annotation were retained. The source quantification contains 7,760 test constructs after the authors' barcode/count filtering; inactive but technically valid constructs remain in the table and are marked by active_vs_scramble = false.

Curation notes

This child experiment represents the SH-SY5Y condition of the shared integrated lentiMPRA library. The table was assembled from the published Supplemental Table S6 sequence annotations, Supplemental Table S7 SH-SY5Y activity results, the authors' sh_quant.txt export, and Supplemental Table S9/S11 pairwise result sheets. It is construct-level rather than barcode-level and does not include raw FASTQ. Multi-target names and coordinates are retained verbatim; combined homolog_class values indicate one construct mapped to multiple homologs. The paper notes that both host cell lines are cancer-derived or have abnormal karyotypes. The parent raw_data folder preserves the full supplement, MPRA code/data archive, and ENA run metadata.

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