Genetic and environmental risk factors for atherosclerosis regulate transcription of phosphatase and actin regulating gene PHACTR1. (July 2016)

• Record Type: Journal Article
• Authors: Reschen, Michael E.; Lin, Da; Chalisey, Anil; Soilleux, Elizabeth J.; O'Callaghan, Christopher A.
• Is Part Of: Atherosclerosis. Volume 250(2016)
• Page Start: 95
• Page End: 105
• Publication Date: 2016-07
• Subjects: Myocardial infarction -- Atherosclerosis -- Functional genomics -- Genetic polymorphism -- Genomics -- Low-density lipoprotein (LDL) -- Genetic disease -- PHACTR1 -- Expression quantitative trait locus (eQTL) -- Oxidized low density lipoprotein (oxLDL); CAD coronary artery disease -- GWAS genome wide association studies -- SNP single nucleotide polymorphism -- PHACTR1 phosphatase and actin regulator 1 -- oxLDL oxidized low density lipoprotein -- PP1 protein phosphatase 1 -- PBMC peripheral blood mononuclear cells -- TSS transcription start site -- RACE rapid amplification of cDNA ends -- HAEC human aortic endothelial cells -- LPS lipopolysaccharide -- NLS nuclear localization signal -- DAB diaminobenzidine; Arteriosclerosis -- Periodicals; Electronic journals; 616.136
• DOI: 10.1016/j.atherosclerosis.2016.04.025
• Languages: English
• ISSNs: 0021-9150

Abstract

Abstract: Background and aims: Coronary artery disease (CAD) risk is associated with non-coding genetic variants at the phosphatase and actin regulating protein 1( PHACTR1 ) gene locus. The PHACTR1 gene encodes an actin-binding protein with phosphatase regulating activity. The mechanism whereby PHACTR1 influences CAD risk is unknown. We hypothesized that PHACTR1 would be expressed in human cell types relevant to CAD and regulated by atherogenic or genetic factors. Methods and results: Using immunohistochemistry, we demonstrate that PHACTR1 protein is expressed strongly in human atherosclerotic plaque macrophages, lipid-laden foam cells, adventitial lymphocytes and endothelial cells. Using a combination of genomic analysis and molecular techniques, we demonstrate that PHACTR1 is expressed as multiple previously uncharacterized transcripts in macrophages, foam cells, lymphocytes and endothelial cells. Immunoblotting confirmed a total absence of PHACTR1 in vascular smooth muscle cells. Real-time quantitative PCR showed that PHACTR1 is regulated by atherogenic and inflammatory stimuli. In aortic endothelial cells, oxLDL and TNF-alpha both upregulated an intermediate length transcript. A short transcript expressed only in immune cells was upregulated in macrophages by oxidized low-density lipoprotein, and oxidized phospholipids but suppressed by lipopolysaccharide or TNF-alpha. In primary human macrophages, we identified a novel expression quantitative trait locus (eQTL) specific for this short transcript, whereby the risk allele at CAD risk SNP rs9349379 is associated with reduced PHACTR1 expression, similar to the effect of an inflammatory stimulus. Conclusions: Our data demonstrate that PHACTR1 is a key atherosclerosis candidate gene since it is regulated by atherogenic stimuli in macrophages and endothelial cells and we identify an effect of the genetic risk variant on PHACTR1 expression in macrophages that is similar to that of an inflammatory stimulus. Graphical abstract: Highlights: PHACTR1 is expressed as two transcripts in both immune and endothelial cells in human atherosclerotic plaque. Oxidized-LDL upregulates a short PHACTR1 transcript, but suppresses an intermediate length transcript in macrophages. Lipopolysaccharide and TNF-alpha cause the opposite effect with strong suppression of the short transcript in macrophages. The coronary artery disease risk SNP, rs9349379, is associated with expression of the short transcript in macrophages. The effect of the coronary artery disease risk allele on PHACTR1 mirrors that of inflammatory stimuli.