Massively parallel reporter assays combined with cell-type specific eQTL identified a functional melanoma risk variant in HIV-1 restriction gene, MX2 [MPRA NGS]
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ABSTRACT:
PROVIDER: PRJNA530577 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR8838560_1.fastq.gz | Fastqsanger.gz | |||
| SRR8838560_2.fastq.gz | Fastqsanger.gz | |||
| SRR8838570_1.fastq.gz | Fastqsanger.gz | |||
| SRR8838570_2.fastq.gz | Fastqsanger.gz | |||
| SRR8838561_1.fastq.gz | Fastqsanger.gz |
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Project description:Genome-wide association studies (GWAS) have identified twenty melanoma susceptibility loci. To identify susceptibility genes and variants simultaneously from multiple GWAS loci, we integrated massively-parallel reporter assays (MPRA) with melanocyte-specific expression quantitative trait locus (eQTL) profiling. We identified thirty-nine candidate functional variants displaying allelic transcriptional activity, of which nine from four loci were also correlated with local gene expression in melanocytes (CTSS, CASP8, MX2, and MAFF). Among these, we further characterized the locus in MX2 gene on chromosome band Chr21q22.3 and validated a functional variant, rs398206. The functional variant mediates allelic transcriptional activity via binding of the transcription factor, YY1. This allelic transcriptional regulation largely accounts for a significant cis-eQTL of the HIV-1 restriction gene, MX2, in primary melanocytes, where the melanoma risk-associated A allele is correlated with higher MX2 levels. Melanocyte-specific transgenic expression of human MX2 in a zebrafish model demonstrated an accelerated melanoma formation in a BRAFV600E background. Thus, using an efficient scalable approach to streamline GWAS follow-up functional studies, we uncovered a pleiotropic function of MX2 in melanoma susceptibility.
2020-04-20 | GSE129242 | GEO
Project description:Genome-wide association studies (GWAS) have identified twenty melanoma susceptibility loci. To identify susceptibility genes and variants simultaneously from multiple GWAS loci, we integrated massively-parallel reporter assays (MPRA) with melanocyte-specific expression quantitative trait locus (eQTL) profiling. We identified thirty-nine candidate functional variants displaying allelic transcriptional activity, of which nine from four loci were also correlated with local gene expression in melanocytes (CTSS, CASP8, MX2, and MAFF). Among these, we further characterized the locus in MX2 gene on chromosome band Chr21q22.3 and validated a functional variant, rs398206. The functional variant mediates allelic transcriptional activity via binding of the transcription factor, YY1. This allelic transcriptional regulation largely accounts for a significant cis-eQTL of the HIV-1 restriction gene, MX2, in primary melanocytes, where the melanoma risk-associated A allele is correlated with higher MX2 levels. Melanocyte-specific transgenic expression of human MX2 in a zebrafish model demonstrated an accelerated melanoma formation in a BRAFV600E background. Thus, using an efficient scalable approach to streamline GWAS follow-up functional studies, we uncovered a pleiotropic function of MX2 in melanoma susceptibility.
2020-04-23 | GSE149161 | GEO
Project description:Genome-wide association studies (GWAS) have identified twenty melanoma susceptibility loci. To identify susceptibility genes and variants simultaneously from multiple GWAS loci, we integrated massively-parallel reporter assays (MPRA) with melanocyte-specific expression quantitative trait locus (eQTL) profiling. We identified thirty-nine candidate functional variants displaying allelic transcriptional activity, of which nine from four loci were also correlated with local gene expression in melanocytes (CTSS, CASP8, MX2, and MAFF). Among these, we further characterized the locus in MX2 gene on chromosome band Chr21q22.3 and validated a functional variant, rs398206. The functional variant mediates allelic transcriptional activity via binding of the transcription factor, YY1. This allelic transcriptional regulation largely accounts for a significant cis-eQTL of the HIV-1 restriction gene, MX2, in primary melanocytes, where the melanoma risk-associated A allele is correlated with higher MX2 levels. Melanocyte-specific transgenic expression of human MX2 in a zebrafish model demonstrated an accelerated melanoma formation in a BRAFV600E background. Thus, using an efficient scalable approach to streamline GWAS follow-up functional studies, we uncovered a pleiotropic function of MX2 in melanoma susceptibility.
2020-04-20 | GSE129237 | GEO
Project description:The most recent genome-wide association study (GWAS) of cutaneous melanoma identified 54 risk-associated loci, but functional variants and their target genes for most have not been established. Here, we performed massively parallel reporter assays (MPRA) using malignant melanoma and normal melanocyte cells and further integrated multi-layer annotation to systematically prioritize functional variants and susceptibility genes from these GWAS loci. Of 1,992 risk-associated variants tested in MPRA, we identified 285 from 42 loci (78% of the known loci) displaying significant allelic transcriptional activities in either cell type (FDR < 1%). We further characterized MPRA-significant variants by motif prediction, epigenomic annotation, and statistical/functional fine-mapping to create integrative variant scores, which prioritized one to six plausible candidate variants per locus for the 42 loci and nominated a single variant for 43% of these loci. Overlaying the MPRA-significant variants with genome-wide significant expression or methylation quantitative trait loci (QTLs) from melanocytes or melanomas identified candidate susceptibility genes for 60% of variants (172 of 285 variants). CRISPRi of top-scoring variants validated their cis-regulatory effect on the eQTL target genes, MAFF (22q13.1) and GPRC5A (12p13.1). Finally, we identified 36 melanoma-specific and 45 melanocyte-specific MPRA-significant variants, a subset of which are linked to cell-type-specific target genes. Analyses of transcription factor availability in MPRA datasets and variant-transcription factor interaction in eQTL datasets highlighted the roles of transcription factors in cell-type-specific variant functionality. In conclusion, MPRA along with variant scoring effectively prioritized plausible candidates for most melanoma GWAS loci and highlighted cellular contexts where the susceptibility variants are functional.
2022-11-28 | GSE210354 | GEO
Project description:Motivation:The majority of the human genome is composed of non-coding regions containing regulatory elements such as enhancers, which are crucial for controlling gene expression. Many variants associated with complex traits are in these regions, and may disrupt gene regulatory sequences. Consequently, it is important to not only identify true enhancers but also to test if a variant within an enhancer affects gene regulation. Recently, allele-specific analysis in high-throughput reporter assays, such as massively parallel reporter assays (MPRAs), have been used to functionally validate non-coding variants. However, we are still missing high-quality and robust data analysis tools for these datasets. Results:We have further developed our method for allele-specific analysis QuASAR (quantitative allele-specific analysis of reads) to analyze allele-specific signals in barcoded read counts data from MPRA. Using this approach, we can take into account the uncertainty on the original plasmid proportions, over-dispersion, and sequencing errors. The provided allelic skew estimate and its standard error also simplifies meta-analysis of replicate experiments. Additionally, we show that a beta-binomial distribution better models the variability present in the allelic imbalance of these synthetic reporters and results in a test that is statistically well calibrated under the null. Applying this approach to the MPRA data, we found 602 SNPs with significant (false discovery rate 10%) allele-specific regulatory function in LCLs. We also show that we can combine MPRA with QuASAR estimates to validate existing experimental and computational annotations of regulatory variants. Our study shows that with appropriate data analysis tools, we can improve the power to detect allelic effects in high-throughput reporter assays. Availability and implementation:http://github.com/piquelab/QuASAR/tree/master/mpra. Contact:fluca@wayne.edu or rpique@wayne.edu. Supplementary information:Supplementary data are available online at Bioinformatics.
| S-EPMC6049023 | biostudies-literature
Project description:We performed a massively parallel reporter assay on 2,396 genomic regions containing single nucleotide polymorphisms that are in high linkage disequilibrium with 97 lead variants from an obesity GWAS (PMID: 25673413). Regions were transfected into human SGBS preadipocytes, SGBS mature adipocytes, 3T3-L1 preadipocytes, HT22 hippocampal cells, and GT1-7 cells and assessed for enhancer activity. The processed file contains the MPRA barcodes.
2021-05-22 | E-MTAB-10463 | biostudies-arrayexpress
Project description:We employ a massively parallel reporter assay (MPRA) to measure the ex vivo activities of hundreds of K562 and HepG2 enhancers with known transcription factor motif instances. For seven selected motifs that correspond to known or predicted activators and repressors in the two cell types, we make directed modifications of the bases corresponding to these motifs and observe the changes in enhancer activity. Reporter mRNA-seq from HepG2 and K562 cells transfected with a ~55,000-plex MPRA plasmid pool containing 5,418 mutated human enhancer sequences, each linked to 10 distinct 10-nt tags. The reporter mRNA tags facilitate quantitation of their abundances. The same tags were also sequenced from the transfected MPRA plasmid pool to facilitate normalization to plasmid copy numbers.
2013-03-15 | E-GEOD-33367 | biostudies-arrayexpress
Project description:The most recent genome-wide association study (GWAS) of cutaneous melanoma identified 54 risk-associated loci, but functional variants and their target genes for most have not been established. Here, we performed massively parallel reporter assays (MPRA) using malignant melanoma and normal melanocyte cells and further integrated multi-layer annotation to systematically prioritize functional variants and susceptibility genes from these GWAS loci. Of 1,992 risk-associated variants tested in MPRA, we identified 285 from 42 loci (78% of the known loci) displaying significant allelic transcriptional activities in either cell type (FDR < 1%). We further characterized MPRA-significant variants by motif prediction, epigenomic annotation, and statistical/functional fine-mapping to create integrative variant scores, which prioritized one to six plausible candidate variants per locus for the 42 loci and nominated a single variant for 43% of these loci. Overlaying the MPRA-significant variants with genome-wide significant expression or methylation quantitative trait loci (QTLs) from melanocytes or melanomas identified candidate susceptibility genes for 60% of variants (172 of 285 variants). CRISPRi of top-scoring variants validated their cis-regulatory effect on the eQTL target genes, MAFF (22q13.1) and GPRC5A (12p13.1). Finally, we identified 36 melanoma-specific and 45 melanocyte-specific MPRA-significant variants, a subset of which are linked to cell-type-specific target genes. Analyses of transcription factor availability in MPRA datasets and variant-transcription factor interaction in eQTL datasets highlighted the roles of transcription factors in cell-type-specific variant functionality. In conclusion, MPRA along with variant scoring effectively prioritized plausible candidates for most melanoma GWAS loci and highlighted cellular contexts where the susceptibility variants are functional.
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Project description:A massively parallel reporter assay, MPRA, was conducted in mouse embryonic stem cells (mESC). Synthetic cis-regulatory elements comprised of binding sites for pluripotency transcription factors and genomic sequences with comparable binding sites configurations were used in the assay. Transcripts of dsRed were amplified via PCR from the end of the transcript to sequence 3' UTR barcodes.
2019-09-18 | GSE120240 | GEO
Project description:Genome-wide association studies (GWAS) have identified ~20 melanoma susceptibility loci, most of which are not functionally characterized. Here we report an approach integrating massively-parallel reporter assays (MPRA) with cell-type-specific epigenome and expression quantitative trait loci (eQTL) to identify susceptibility genes/variants from multiple GWAS loci. From 832 high-LD variants, we identify 39 candidate functional variants from 14 loci displaying allelic transcriptional activity, a subset of which corroborates four colocalizing melanocyte cis-eQTL genes. Among these, we further characterize the locus encompassing the HIV-1 restriction gene, MX2 (Chr21q22.3), and validate a functional intronic variant, rs398206. rs398206 mediates the binding of the transcription factor, YY1, to increase MX2 levels, consistent with the cis-eQTL of MX2 in primary human melanocytes. Melanocyte-specific expression of human MX2 in a zebrafish model demonstrates accelerated melanoma formation in a BRAFV600E background. Our integrative approach streamlines GWAS follow-up studies and highlights a pleiotropic function of MX2 in melanoma susceptibility.
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