Calmodulin-like protein 3 is an estrogen receptor alpha coregulator for gene expression and drug response in a SNP, estrogen, and SERM-dependent fashion

Abstract

BackgroundWe previously performed a case–control genome-wide association study in women treated with selective estrogen receptor modulators (SERMs) for breast cancer prevention and identified single nucleotide polymorphisms (SNPs) in ZNF423 as potential biomarkers for response to SERM therapy. The ZNF423rs9940645 SNP, which is approximately 200 bp away from the estrogen response elements, resulted in the SNP, estrogen, and SERM-dependent regulation of ZNF423 expression and, “downstream”, that of BRCA1.MethodsElectrophoretic mobility shift assay–mass spectrometry was performed to identify proteins binding to the ZNF423 SNP and coordinating with estrogen receptor alpha (ERα). Clustered, regularly interspaced short palindromic repeats (CRISPR)/Cas9 genome editing was applied to generate ZR75-1 breast cancer cells with different ZNF423 SNP genotypes. Both cultured cells and mouse xenograft models with different ZNF423 SNP genotypes were used to study the cellular responses to SERMs and poly(ADP-ribose) polymerase (PARP) inhibitors.ResultsWe identified calmodulin-like protein 3 (CALML3) as a key sensor of this SNP and a coregulator of ERα, which contributes to differential gene transcription regulation in an estrogen and SERM-dependent fashion. Furthermore, using CRISPR/Cas9-engineered ZR75-1 breast cancer cells with different ZNF423 SNP genotypes, striking differences in cellular responses to SERMs and PARP inhibitors, alone or in combination, were observed not only in cells but also in a mouse xenograft model.ConclusionsOur results have demonstrated the mechanism by which the ZNF423 rs9940645 SNP might regulate gene expression and drug response as well as its potential role in achieving more highly individualized breast cancer therapy.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-017-0890-x) contains supplementary material, which is available to authorized users.

Single Nucleotide Polymorphisms (SNPs) Distant from Xenobiotic Response Elements Can Modulate Aryl Hydrocarbon Receptor Function: SNP-Dependent CYP1A1 Induction

“…Based on our previous observations with respect to the modulation of ERa-ERE binding and downstream gene transcription (Ingle et al, 2013(Ingle et al, , 2016Ho et al, 2016;Liu et al, 2017;Qin et al, 2017), in the present study, we set out to test the hypothesis that SNPs hundreds of bp distant from XREs might influence AHR-XRE binding and downstream gene expression. We chose to study CYP1A1, a prototypical AHR-regulated gene, by analyzing DNA sequences across and both up and downstream of the open reading frame for putative XREs and common SNPs that mapped 6 500 bp of those XREs.…”

“…We have previously reported that proteins encoded by members of the testis specific Y-like (TSPYL) gene family were coregulators for the transcription of cytochrome P450s, including CYP3A4, CYP2C9, CYP2C19, CYP17A1, and CYP19A1 (Liu et al, 2013;Qin et al, 2018). Coregulators like members of the TSPYL gene family may, among other mechanisms, bind to specific DNA sequences, alter the nucleosome structure or recruit RNA polymerase, thus facilitating gene transcription (Qin et al, 2017). Therefore, changes in the binding of coregulators to DNA sequences as a result of SNPs that map hundreds of base pairs from response elements might well affect downstream gene transcription.…”

“…Functional genomic studies demonstrated that the ZNF423 rs9940645 SNP, which maps 200 bp away from an ERE cluster, could influence ERa-ERE binding and ZNF423 transcription (Ingle et al, 2013). Furthermore, subsequent studies identified calmodulin-like protein 3 as a co-regulator of ERa that "sensed" genotypes for the ZNF423 intron two SNP (Qin et al, 2017).…”

Single Nucleotide Polymorphisms at a Distance from Aryl Hydrocarbon Receptor (AHR) Binding Sites Influence AHR Ligand–Dependent Gene Expression

“…Obviously, this study has not resulted in an exhaustive list of AHR ligand-dependent PGx-eQTLs, but this series of experiments has provided evidence that SNPs across the genome that are at a distance from AHREs can influence AHR ligand-dependent expression. Based on the results of our previous studies, these SNPs may interact with coregulator(s) that binds to AHR to form a larger complex and may alter complex stability binding at that locus (Liu et al, 2012(Liu et al, , 2018aZhou, 2016;Ho et al, 2017;Qin et al, 2017). Future studies aimed at understanding AHR PGx-eQTLs should focus on identifying mechanisms underlying the function of these SNPs and mechanisms responsible for their cell-specific effects.…”

“…Obviously, SNPs can create/disrupt canonical TF-binding motifs (Knight et al, 1999;Bream et al, 2002;Ingle et al, 2010), but our previous studies identified several examples of SNPs that map hundreds of base pairs (bp) distant from ligand-activated TF binding sites that could influence TF binding in a SNP genotype-by-drug exposuredependent manner (Liu et al, 2012(Liu et al, , 2018aZhou, 2016;Ho et al, 2017;Qin et al, 2017). One of those studies identified a PGx-eQTL for the aryl hydrocarbon receptor (AHR), a ligand-activated TF that is an important environmental modulator for several critical pathways and processes, including chemical toxicology, immune stimulation, cancer, and the kynurenine pathway (Stockinger et al, 2014;Liu et al, 2018b;Neavin et al, 2018).…”