Periodic Reporting for period 5 - MODVASC (Endothelial RNA Modifications in Vascular Homeostasis and Disease)
Période du rapport: 2023-03-01 au 2024-03-31
In an ageing society more and more people are spending more years living with chronic conditions like atherosclerosis resulting in an enormous health toll. Atherosclerosis, a major cause or morbidity and mortality in our world, is a multifactorial vascular disease driven by endothelial dysfunction.
The vascular endothelium, a monolayer of endothelial cells (ECs), constitutes the inner cellular layer of arteries, veins and capillaries forming a critical interface between blood and tissue. Depending on their location within the circulatory system, vascular ECs may act as sentinel cells sensing microenvironmental cues and as gatekeepers controlling the influx and outflux of circulating molecules, nutrients and immune cells from circulating blood to target tissues and vice versa. This strategic role of ECs makes them as the master regulator of blood-brain and blood-body barrier function, organ-organ and cell-cell crosstalk, immune system activation and organ function in health and disease.
Older people who smoke, have high blood pressure, high glucose or lipid levels in blood and are overweight are at high risk for future cardiovascular events including myocardial infarction and stroke due to endothelial inflammation of the vascular wall leading to atherosclerotic plaque erosion or rupture and subsequently arterial thrombosis. By endothelial inflammation it is meant the EC-triggered activation of the immune system leading to recruitment of circulating immune cells into the vascular wall and development of atherosclerotic plaques. Depending on which arterial bed is affected, atherosclerosis of the coronary arteries leads to acute and chronic coronary syndromes, of the peripheral arteries to peripheral arterial disease and of the cerebrovascular arteries to stroke. All these diseases may lead to organ dysfunction including heart failure, claudication (pain in the legs or arms that occurs while walking or using the arms) and dementia that significantly limit health- and lifespan. Despite recent medical advances and guideline-suggested treatment, atherosclerosis remains an incurable disease. Targeting chronic vascular inflammation seems an obvious solution, but inhibition of one EC signaling pathway is not enough to completely abolish endothelial inflammation due to the presence of aberrant signaling pathways that all lead to an EC pro-inflammatory and pro-atherosclerotic phenotype. Thus, the search for upstream checkpoints of EC immune activation has been the “holy grail” of scientific research in cardiovascular biology and medicine. Understanding the exact molecular mechanisms that orchestrate endothelial cell function and response to environmental stimuli is thus essential for the discovery and development of novel biomarkers and therapeutic strategies in vascular disease.
RNA modifications are (non)reversible, enzymatic ribonucleic acid (RNA) base modifications that expand the RNA alphabet from the 4 canonical nucleotides to more than 170. Among the most abundant RNA modifications in mammals is m6A RNA methylation. RNA modifications may control all aspects of RNA life affecting in a well-coordinated manner several cellular signaling pathways within the same cells. This unique feature of RNA modifications make them ideal upstream coordinators of hundreds of genes involved in several signaling pathways affecting cellular function in health and disease.
Adenosine methylation in the N6 position (m6A) is the most prominent RNA modification in eukaryotes and is mediated by METTL3, METT14 and WTAP writer complex. M6A RNA methylation is a highly conserved RNA modification among organisms and its presence in our cells has been shown to be essential for life. Accumulating recent evidence suggests that m6A RNA methylation controls cellular processes by interfering with the RNA – protein interaction and, thus, RNA fate. The ERC project MODVASC – endothelial RNA modifications in vascular health and disease- aimed at answering the following scientific questions: Is m6A RNA methylation important for EC function in health and disease? Does m6A RNA methylation affect EC gene expression and cellular signaling in response to pro-inflammatory stimuli? How does m6A RNA methylation control RNA fate? Are m6A RNA methylation levels altered in patients with human atherosclerosis? Which genes are differentially methylated in vascular disease?
The vascular endothelium, a monolayer of endothelial cells (ECs), constitutes the inner cellular layer of arteries, veins and capillaries forming a critical interface between blood and tissue. Depending on their location within the circulatory system, vascular ECs may act as sentinel cells sensing microenvironmental cues and as gatekeepers controlling the influx and outflux of circulating molecules, nutrients and immune cells from circulating blood to target tissues and vice versa. This strategic role of ECs makes them as the master regulator of blood-brain and blood-body barrier function, organ-organ and cell-cell crosstalk, immune system activation and organ function in health and disease.
Older people who smoke, have high blood pressure, high glucose or lipid levels in blood and are overweight are at high risk for future cardiovascular events including myocardial infarction and stroke due to endothelial inflammation of the vascular wall leading to atherosclerotic plaque erosion or rupture and subsequently arterial thrombosis. By endothelial inflammation it is meant the EC-triggered activation of the immune system leading to recruitment of circulating immune cells into the vascular wall and development of atherosclerotic plaques. Depending on which arterial bed is affected, atherosclerosis of the coronary arteries leads to acute and chronic coronary syndromes, of the peripheral arteries to peripheral arterial disease and of the cerebrovascular arteries to stroke. All these diseases may lead to organ dysfunction including heart failure, claudication (pain in the legs or arms that occurs while walking or using the arms) and dementia that significantly limit health- and lifespan. Despite recent medical advances and guideline-suggested treatment, atherosclerosis remains an incurable disease. Targeting chronic vascular inflammation seems an obvious solution, but inhibition of one EC signaling pathway is not enough to completely abolish endothelial inflammation due to the presence of aberrant signaling pathways that all lead to an EC pro-inflammatory and pro-atherosclerotic phenotype. Thus, the search for upstream checkpoints of EC immune activation has been the “holy grail” of scientific research in cardiovascular biology and medicine. Understanding the exact molecular mechanisms that orchestrate endothelial cell function and response to environmental stimuli is thus essential for the discovery and development of novel biomarkers and therapeutic strategies in vascular disease.
RNA modifications are (non)reversible, enzymatic ribonucleic acid (RNA) base modifications that expand the RNA alphabet from the 4 canonical nucleotides to more than 170. Among the most abundant RNA modifications in mammals is m6A RNA methylation. RNA modifications may control all aspects of RNA life affecting in a well-coordinated manner several cellular signaling pathways within the same cells. This unique feature of RNA modifications make them ideal upstream coordinators of hundreds of genes involved in several signaling pathways affecting cellular function in health and disease.
Adenosine methylation in the N6 position (m6A) is the most prominent RNA modification in eukaryotes and is mediated by METTL3, METT14 and WTAP writer complex. M6A RNA methylation is a highly conserved RNA modification among organisms and its presence in our cells has been shown to be essential for life. Accumulating recent evidence suggests that m6A RNA methylation controls cellular processes by interfering with the RNA – protein interaction and, thus, RNA fate. The ERC project MODVASC – endothelial RNA modifications in vascular health and disease- aimed at answering the following scientific questions: Is m6A RNA methylation important for EC function in health and disease? Does m6A RNA methylation affect EC gene expression and cellular signaling in response to pro-inflammatory stimuli? How does m6A RNA methylation control RNA fate? Are m6A RNA methylation levels altered in patients with human atherosclerosis? Which genes are differentially methylated in vascular disease?
In MODVASC, we identified that m6A RNA methylation is essential for both vascular development and growth as well as for maintenance of vascular homeostasis. We identified the methylated RNA molecules and the cellular processes that are affected by m6A RNA methylation. We discovered the regulatory effect of m6A RNA methylation in the expression of the messenger RNAs (mRNAs) which are the RNA molecules that carry the genetic information from nucleus to cytoplasm and unraveled the underlying molecular mechanism. Next, we characterized the role of methylated genes in EC function in health and disease. We mapped the m6A RNA methylome in ECs and in blood cells derived from patients with atherosclerosis and characterized the translational and clinical value of m6A RNA methylation writing machinery in human atherosclerotic vascular disease.
MODVASC findings are presented in several international conferences and are integrated into manuscripts that are published or currently prepared for publication in peer-reviewed journals. Identifying the dynamically methylated RNAs in ECs and characterizing how m6A RNA methylation mediates a precise adjustment of RNA life to environmental stimuli has unraveled the presence of a fundamental master regulator of EC coordinated responses to inflammatory stress with important implications for the development of atherosclerosis.
MODVASC findings are presented in several international conferences and are integrated into manuscripts that are published or currently prepared for publication in peer-reviewed journals. Identifying the dynamically methylated RNAs in ECs and characterizing how m6A RNA methylation mediates a precise adjustment of RNA life to environmental stimuli has unraveled the presence of a fundamental master regulator of EC coordinated responses to inflammatory stress with important implications for the development of atherosclerosis.
Our findings made the following major contributions in our understanding of the master RNA regulators of EC function in health and disease. Specifically:
1) Paradigm shift for the maintenance of EC homeostasis: Before the ERC project MODVASC is performed, we thought that the maintenance of EC homeostasis is mediated by effector proteins. During the execution of the ERC project MODVASC we got to know that m6A RNA methylation of specific endothelial genes and their post-transcriptional regulation is essential for the maintenance of vascular endothelial homeostasis in health.
2) Paradigm shift for the vascular endothelial control of inflammation in atherosclerotic vascular disease: Till now we knew that endothelial cells respond passively to microenvironmental changes by increasing their transcriptional programme of several genes. After the execution of the ERC project MODVASC we got to know about the presence of endogenous nucleotide chemical modifications in RNA molecules that may effectively rewrite the transcriptional programme of hundreds of genes affecting EC responses to environmental cues. This mechanism is of fundamental importance in atherosclerotic vascular disease.
3) Progress beyond the state of the art: by utilizing several modern technologies we managed to map m6A RNA methylation sites at single nucleotide level unraveling the first methylome mapping of RNA molecules in health and disease.
1) Paradigm shift for the maintenance of EC homeostasis: Before the ERC project MODVASC is performed, we thought that the maintenance of EC homeostasis is mediated by effector proteins. During the execution of the ERC project MODVASC we got to know that m6A RNA methylation of specific endothelial genes and their post-transcriptional regulation is essential for the maintenance of vascular endothelial homeostasis in health.
2) Paradigm shift for the vascular endothelial control of inflammation in atherosclerotic vascular disease: Till now we knew that endothelial cells respond passively to microenvironmental changes by increasing their transcriptional programme of several genes. After the execution of the ERC project MODVASC we got to know about the presence of endogenous nucleotide chemical modifications in RNA molecules that may effectively rewrite the transcriptional programme of hundreds of genes affecting EC responses to environmental cues. This mechanism is of fundamental importance in atherosclerotic vascular disease.
3) Progress beyond the state of the art: by utilizing several modern technologies we managed to map m6A RNA methylation sites at single nucleotide level unraveling the first methylome mapping of RNA molecules in health and disease.