Translatome profiling reveals Itih4 as a novel smooth muscle cell-specific gene in atherosclerosis
Ravindran, Aarthi; Holappa, Lari; Niskanen, Henri; Skovorodkin, Ilya; Kaisto, Susanna; Beter, Mustafa; Kiema, Miika; Selvarajan, Ilakya; Nurminen, Valtteri; Aavik, Einari; Aherrahrou, Rédouane; Pasonen-Seppänen, Sanna; Fortino, Vittorio; Laakkonen, Johanna P; Ylä-Herttuala, Seppo; Vainio, Seppo; Örd, Tiit; Kaikkonen, Minna U (2024-01-30)
Oxford University Press
30.01.2024
Aarthi Ravindran, Lari Holappa, Henri Niskanen, Ilya Skovorodkin, Susanna Kaisto, Mustafa Beter, Miika Kiema, Ilakya Selvarajan, Valtteri Nurminen, Einari Aavik, Rédouane Aherrahrou, Sanna Pasonen-Seppänen, Vittorio Fortino, Johanna P Laakkonen, Seppo Ylä-Herttuala, Seppo Vainio, Tiit Örd, Minna U Kaikkonen, Translatome profiling reveals Itih4 as a novel smooth muscle cell–specific gene in atherosclerosis, Cardiovascular Research, Volume 120, Issue 8, May 2024, Pages 869–882, https://doi.org/10.1093/cvr/cvae028
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© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
https://creativecommons.org/licenses/by/4.0/
© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
https://creativecommons.org/licenses/by/4.0/
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:oulu-202405143488
https://urn.fi/URN:NBN:fi:oulu-202405143488
Tiivistelmä
Abstract
Aims:
Vascular smooth muscle cells (SMCs) and their derivatives are key contributors to the development of atherosclerosis. However, studying changes in SMC gene expression in heterogeneous vascular tissues is challenging due to the technical limitations and high cost associated with current approaches. In this paper, we apply translating ribosome affinity purification sequencing to profile SMC-specific gene expression directly from tissue.
Methods and results:
To facilitate SMC-specific translatome analysis, we generated SMCTRAP mice, a transgenic mouse line expressing enhanced green fluorescent protein (EGFP)-tagged ribosomal protein L10a (EGFP-L10a) under the control of the SMC-specific αSMA promoter. These mice were further crossed with the atherosclerosis model Ldlr−/−, ApoB100/100 to generate SMCTRAP−AS mice and used to profile atherosclerosis-associated SMCs in thoracic aorta samples of 15-month-old SMCTRAP and SMCTRAP-AS mice. Our analysis of SMCTRAP-AS mice showed that EGFP-L10a expression was localized to SMCs in various tissues, including the aortic wall and plaque. The TRAP fraction demonstrated high enrichment of known SMC-specific genes, confirming the specificity of our approach. We identified several genes, including Cemip, Lum, Mfge8, Spp1, and Serpina3, which are known to be involved in atherosclerosis-induced gene expression. Moreover, we identified several novel genes not previously linked to SMCs in atherosclerosis, such as Anxa4, Cd276, inter-alpha-trypsin inhibitor-4 (Itih4), Myof, Pcdh11x, Rab31, Serpinb6b, Slc35e4, Slc8a3, and Spink5. Among them, we confirmed the SMC-specific expression of Itih4 in atherosclerotic lesions using immunofluorescence staining of mouse aortic roots and spatial transcriptomics of human carotid arteries. Furthermore, our more detailed analysis of Itih4 showed its link to coronary artery disease through the colocalization of genome-wide association studies, splice quantitative trait loci (QTL), and protein QTL signals.
Conclusion:
We generated a SMC-specific TRAP mouse line to study atherosclerosis and identified Itih4 as a novel SMC-expressed gene in atherosclerotic plaques, warranting further investigation of its putative function in extracellular matrix stability and genetic evidence of causality.
Aims:
Vascular smooth muscle cells (SMCs) and their derivatives are key contributors to the development of atherosclerosis. However, studying changes in SMC gene expression in heterogeneous vascular tissues is challenging due to the technical limitations and high cost associated with current approaches. In this paper, we apply translating ribosome affinity purification sequencing to profile SMC-specific gene expression directly from tissue.
Methods and results:
To facilitate SMC-specific translatome analysis, we generated SMCTRAP mice, a transgenic mouse line expressing enhanced green fluorescent protein (EGFP)-tagged ribosomal protein L10a (EGFP-L10a) under the control of the SMC-specific αSMA promoter. These mice were further crossed with the atherosclerosis model Ldlr−/−, ApoB100/100 to generate SMCTRAP−AS mice and used to profile atherosclerosis-associated SMCs in thoracic aorta samples of 15-month-old SMCTRAP and SMCTRAP-AS mice. Our analysis of SMCTRAP-AS mice showed that EGFP-L10a expression was localized to SMCs in various tissues, including the aortic wall and plaque. The TRAP fraction demonstrated high enrichment of known SMC-specific genes, confirming the specificity of our approach. We identified several genes, including Cemip, Lum, Mfge8, Spp1, and Serpina3, which are known to be involved in atherosclerosis-induced gene expression. Moreover, we identified several novel genes not previously linked to SMCs in atherosclerosis, such as Anxa4, Cd276, inter-alpha-trypsin inhibitor-4 (Itih4), Myof, Pcdh11x, Rab31, Serpinb6b, Slc35e4, Slc8a3, and Spink5. Among them, we confirmed the SMC-specific expression of Itih4 in atherosclerotic lesions using immunofluorescence staining of mouse aortic roots and spatial transcriptomics of human carotid arteries. Furthermore, our more detailed analysis of Itih4 showed its link to coronary artery disease through the colocalization of genome-wide association studies, splice quantitative trait loci (QTL), and protein QTL signals.
Conclusion:
We generated a SMC-specific TRAP mouse line to study atherosclerosis and identified Itih4 as a novel SMC-expressed gene in atherosclerotic plaques, warranting further investigation of its putative function in extracellular matrix stability and genetic evidence of causality.
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