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

Matthias Prestel; Caroline Prell-Schicker; Tom Webb; Rainer Malik; Barbara Lindner; Natalie Ziesch; Monika Rex-Haffner; Simone Röh; Thanatip Viturawong; Manuel Lehm; Michal Mokry; Hester den Ruijter; Saskia Haitjema; Yaw Asare; Flavia Söllner; Maryam Ghaderi Najafabadi; Rédouane Aherrahrou; Mete Civelek; Nilesh J Samani; Matthias Mann; Christof Haffner; Martin Dichgans

doi:10.1161/strokeaha.119.026112

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

Matthias Prestel, Caroline Prell-Schicker, Tom Webb, Rainer Malik, Barbara Lindner, Natalie Ziesch, Monika Rex-Haffner, Simone Röh, Thanatip Viturawong, Manuel Lehm, Michal Mokry, Hester den Ruijter, Saskia Haitjema, Yaw Asare, Flavia Söllner, Maryam Ghaderi Najafabadi, Rédouane Aherrahrou, Mete Civelek, Nilesh J Samani, Matthias MannChristof Haffner, Martin Dichgans

11 Citations (Scopus)

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.

Original language	English
Journal	Stroke
Volume	50
Issue number	10
Pages (from-to)	2651-2660
Number of pages	10
ISSN	0039-2499
DOIs	https://doi.org/10.1161/strokeaha.119.026112
Publication status	Published - 1 Oct 2019
Externally published	Yes

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Prestel, M., Prell-Schicker, C., Webb, T., Malik, R., Lindner, B., Ziesch, N., Rex-Haffner, M., Röh, S., Viturawong, T., Lehm, M., Mokry, M., den Ruijter, H., Haitjema, S., Asare, Y., Söllner, F., Najafabadi, M. G., Aherrahrou, R., Civelek, M., Samani, N. J., ... Dichgans, M. (2019). The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1. Stroke, 50(10), 2651-2660. https://doi.org/10.1161/strokeaha.119.026112

Prestel, M, Prell-Schicker, C, Webb, T, Malik, R, Lindner, B, Ziesch, N, Rex-Haffner, M, Röh, S, Viturawong, T, Lehm, M, Mokry, M, den Ruijter, H, Haitjema, S, Asare, Y, Söllner, F, Najafabadi, MG, Aherrahrou, R, Civelek, M, Samani, NJ, Mann, M, Haffner, C & Dichgans, M 2019, 'The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1', Stroke, vol. 50, no. 10, pp. 2651-2660. https://doi.org/10.1161/strokeaha.119.026112

@article{6c4d749991b8425390d75c8a8553fb78,

title = "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.",

author = "Matthias Prestel and Caroline Prell-Schicker and Tom Webb and Rainer Malik and Barbara Lindner and Natalie Ziesch and Monika Rex-Haffner and Simone R{\"o}h and Thanatip Viturawong and Manuel Lehm and Michal Mokry and {den Ruijter}, Hester and Saskia Haitjema and Yaw Asare and Flavia S{\"o}llner and Najafabadi, {Maryam Ghaderi} and R{\'e}douane Aherrahrou and Mete Civelek and Samani, {Nilesh J} and Matthias Mann and Christof Haffner and Martin Dichgans",

year = "2019",

month = oct,

day = "1",

doi = "10.1161/strokeaha.119.026112",

language = "English",

volume = "50",

pages = "2651--2660",

journal = "Stroke",

issn = "0039-2499",

publisher = "Lippincott Williams & Wilkins",

number = "10",

}

TY - JOUR

T1 - The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1

AU - Prestel, Matthias

AU - Prell-Schicker, Caroline

AU - Webb, Tom

AU - Malik, Rainer

AU - Lindner, Barbara

AU - Ziesch, Natalie

AU - Rex-Haffner, Monika

AU - Röh, Simone

AU - Viturawong, Thanatip

AU - Lehm, Manuel

AU - Mokry, Michal

AU - den Ruijter, Hester

AU - Haitjema, Saskia

AU - Asare, Yaw

AU - Söllner, Flavia

AU - Najafabadi, Maryam Ghaderi

AU - Aherrahrou, Rédouane

AU - Civelek, Mete

AU - Samani, Nilesh J

AU - Mann, Matthias

AU - Haffner, Christof

AU - Dichgans, Martin

PY - 2019/10/1

Y1 - 2019/10/1

N2 - 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.

AB - 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.

U2 - 10.1161/strokeaha.119.026112

DO - 10.1161/strokeaha.119.026112

M3 - Journal article

C2 - 31500558

SN - 0039-2499

VL - 50

SP - 2651

EP - 2660

JO - Stroke

JF - Stroke

IS - 10

ER -

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

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