The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1

Abstract

Background and Purpose— Genome-wide association studies have identified the HDAC9 (histone deacetylase 9) gene region as a major risk locus for atherosclerotic stroke and coronary artery disease in humans. Previous results suggest a role of altered HDAC9 expression levels as the underlying disease mechanism. rs2107595, the lead single nucleotide polymorphism for stroke and coronary artery disease resides in noncoding DNA and colocalizes with histone modification marks suggestive of enhancer elements. Methods— To determine the mechanisms by which genetic variation at rs2107595 regulates HDAC9 expression and thus vascular risk we employed targeted resequencing, proteome-wide search for allele-specific nuclear binding partners, chromatin immunoprecipitation, genome-editing, reporter assays, circularized chromosome conformation capture, and gain- and loss-of-function experiments in cultured human cell lines and primary immune cells. Results— Targeted resequencing of the HDAC9 locus in patients with atherosclerotic stroke and controls supported candidacy of rs2107595 as the causative single nucleotide polymorphism. A proteomic search for nuclear binding partners revealed preferential binding of the E2F3/TFDP1/Rb1 complex (E2F transcription factor 3/transcription factor Dp-1/Retinoblastoma 1) to the rs2107595 common allele, consistent with the disruption of an E2F3 consensus site by the risk allele. Gain- and loss-of-function studies showed a regulatory effect of E2F/Rb proteins on HDAC9 expression. Compared with the common allele, the rs2107595 risk allele exhibited higher transcriptional capacity in luciferase assays and was associated with higher HDAC9 mRNA levels in primary macrophages and genome-edited Jurkat cells. Circularized chromosome conformation capture revealed a genomic interaction of the rs2107595 region with the HDAC9 promoter, which was stronger for the common allele as was the in vivo interaction with E2F3 and Rb1 determined by chromatin immunoprecipitation. Gain-of-function experiments in isogenic Jurkat cells demonstrated a key role of E2F3 in mediating rs2107595-dependent transcriptional regulation of HDAC9 . Conclusions— Collectively, our findings imply allele-specific transcriptional regulation of HDAC9 via E2F3 and Rb1 as a major mechanism mediating vascular risk at rs2107595.

Microbiota in cerebrovascular disease: A key player and future therapeutic target

“…HDAC9 polymorphisms are associated with large artery atherosclerosis and stroke, which is inhibited by butyrate, as previously mentioned. 101 Additionally, COL4A1 and COL4A2 polymorphisms are associated with hemorrhagic stroke, disrupting type IV collagen of the vascular basement protein, 102 where Cnm-positive S. mutans adhere. 87,103 APOE and TREM2 are the most influential genetic risk factors for AD and CAA.…”

Histone Deacetylases (HDACs) and Atherosclerosis: A Mechanistic and Pharmacological Review

“…The rs2107595 HDAC9 gene polymorphism leads to increased expression of HDAC9 in the internal carotid artery (Grbić et al, 2020) and plasma (Wang et al, 2016) and modulated gene expression in the blood of patients suffering from large vessel atherosclerotic stroke (Shroff et al, 2019). The polymorphism variant is significantly associated with AS (Prestel et al, 2019;Grbić et al, 2020) and may contribute to coronary AS and coronary artery disease risk (Wang et al, 2016). Consistently, HDAC9 deficiency in ApoE −/− mice resulted in evidently reduced lesion size in aortas and less advanced lesions (Azghandi et al, 2015).…”

Making fundamental scientific discoveries by combining information from literature, databases, and computational tools – An example

“… A-E) DNA-related occurrences of SAMD1 in the literature [9] , [13] , [14] , [15] , [16] . Those publications cannot be found by standard literature search but were discovered via internet research.…”