Massively parallel reporter assays combined with cell-type specific eQTL informed multiple melanoma loci and identified a pleiotropic function of HIV-1 restriction gene,MX2, in melanoma promotion

Abstract

Genome-wide association studies (GWAS) have identified ∼20 melanoma susceptibility loci. To identify susceptibility genes and variants simultaneously from multiple GWAS loci, we integrated massively-parallel reporter assays (MPRA) with cell type-specific epigenomic data as well as melanocyte-specific expression quantitative trait loci (eQTL) profiling. Starting from 16 melanoma loci, we selected 832 variants overlapping active regions of chromatin in cells of melanocytic lineage and identified 39 candidate functional variants displaying allelic transcriptional activity by MPRA. For four of these loci, we further identified four colocalizing melanocyte cis-eQTL genes (CTSS, CASP8, MX2, and MAFF) matching the allelic activity of MPRA functional variants. Among these, we further characterized the locus encompassing the HIV-1 restriction gene, MX2, on chromosome band Chr21q22.3 and validated a functional variant, rs398206, among multiple high LD variants. rs398206 mediates allelic transcriptional activity via binding of the transcription factor, YY1. This allelic transcriptional regulation is consistent with a significant cis-eQTL of MX2 in primary human melanocytes, where the melanoma risk-associated A allele of rs398206 is correlated with higher MX2 levels. Melanocyte-specific transgenic expression of human MX2 in a zebrafish model demonstrated accelerated melanoma formation in a BRAFV600E background. Thus, using an efficient scalable approach to streamline GWAS follow-up functional studies, we identified multiple candidate melanoma susceptibility genes and variants, and uncovered a pleiotropic function of MX2 in melanoma susceptibility.

Systematic identification of cis-regulatory variants that cause gene expression differences in a yeast cross

“…Other applications have assayed sequences that promote splicing ( Cheung et al, 2019 ; Rosenberg et al, 2015 ), translation ( Goodman et al, 2013 ; Weingarten-Gabbay et al, 2016 ), DNA methylation ( Krebs et al, 2014 ) and RNA editing ( Safra et al, 2017 ). More recently, MPRAs have identified individual human DNA variants that alter gene expression ( Tewhey et al, 2016 ; Ulirsch et al, 2016 ) in studies ranging in scale from variants in specific regions implicated by genome-wide association studies for a given disease ( Choi et al, 2019 ; Liu et al, 2017 ; Pashos et al, 2017 ; Vockley et al, 2015 ) to a genome-wide survey of nearly six million common single-nucleotide polymorphisms ( van Arensbergen et al, 2019 ). In spite of these successes, the size of the human genome, which harbors tens of millions of rare as well as common variants ( 1000 Genomes Project Consortium et al, 2015 ), combined with a high degree of tissue-specificity in gene expression and the activity of regulatory DNA GTEx Consortium et al, 2017 ; Inoue et al, 2019 ; Maricque et al, 2019 have complicated dissection of causal variants in local eQTLs.…”

Massively parallel identification of causal variants underlying gene expression differences in a yeast cross

“…Other applications have assayed sequences that promote splicing (Cheung et al, 2019;Rosenberg et al, 2015), translation (Goodman et al, 2013;Weingarten-Gabbay et al, 2016), DNA methylation (Krebs et al, 2014) and RNA editing (Safra et al, 2017). More recently, MPRAs have identified individual human DNA variants that alter gene expression (Tewhey et al, 2016;Ulirsch et al, 2016) in studies ranging in scale from variants in specific regions implicated by genome-wide association studies for a given disease (Choi et al, 2019;Liu et al, 2017;Pashos et al, 2017;Vockley et al, 2015) to a genome-wide survey of nearly six million common single-nucleotide polymorphisms (Arensbergen et al, 2019). In spite of these successes, the size of the human genome, which harbors tens of millions of rare as well as common variants (Consortium, 2015), combined with a high degree of tissue-specificity in gene expression and the activity of regulatory DNA (GTEx Consortium et al, 2017;Inoue et al, 2019;Maricque et al, 2019) have complicated dissection of causal variants in local eQTLs.…”