Periodic Reporting for period 4 - MODVASC (Endothelial RNA Modifications in Vascular Homeostasis and Disease)
Okres sprawozdawczy: 2021-09-01 do 2023-02-28
Endothelial cells (ECs) cover the entire inner layer of the circulation which delivers blood, oxygen and nutrients in all organs, regulating homeostasis maintenance and organ regeneration after injury and inhibiting or enabling blood cell invasion to injured tissues controlling by this way tissue healing or injury processes. All known cardiovascular risk factors induce a process called, endothelial cell activation, which ultimately leads to atherosclerosis, the major cause of mortality in western world. Currently, atherosclerosis may be manageable with the aid of prescribed standard medication which should be taken for the entire lifespan but this disease has not been yet cured. The socioeconomic burden stemming from this modern world disease is admittedly huge. 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 174 (a brief overview of RNA modifications is available here: http://modomics.genesilico.pl/modifications/). Importantly, they occur intrinsically in all of our cells. Among the most frequent RNA modifications are two different chemical processes both happening on a specific RNA nucleotide, adenosine, and they are termed deamination and methylation. My and other laboratories have reported that RNA modifications control all aspects of RNA life and are thus involved in several cellular processes and functions. The adenosine deamination to inosine is a process called RNA editing and is catalyzed by tasked enzymes, the adenosine deaminases acting on RNAs (ADARs). My laboratory has shown that adenosine deamination regulates cathepsin S mRNA stability by controlling the recruitment of the stabilizing RNA-binding protein called HuR onto the cathepsin S mRNA. Interestingly, we revealed a strong association of this RNA modification with vascular disease, documenting, for the first time, the importance of RNA modifications in cardiovascular system pathology. Adenosine methylation in the N6 position (m6A) is the most prominent RNA modification in eukaryotes and is catalyzed by a set of enzymes acting by being scavenged in another protein hub. This set of enzymes are termed METTL3, METT14 and WTAP. 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.
Identifying the dynamically methylated RNAs of our cells and characterizing how m6A mediates a precise adjustment of RNA life to environmental stimuli will undoubtedly contribute towards the understanding of human disease. This may open the road to the development of new disease biomarkers and therapeutic strategies. MODVASC aims to explore the role of m6A RNA methylation in vascular endothelial cell function and in vascular disease. Specifically, MODVASC will characterize: a) the endothelial methylome, b) the regulatory effects of m6A RNA methylation in endothelial gene expression, c) the role of m6A RNA methylation machinery in vascular development and maintenance of vascular function and d) the regulation of m6A RNA methylation 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 174 (a brief overview of RNA modifications is available here: http://modomics.genesilico.pl/modifications/). Importantly, they occur intrinsically in all of our cells. Among the most frequent RNA modifications are two different chemical processes both happening on a specific RNA nucleotide, adenosine, and they are termed deamination and methylation. My and other laboratories have reported that RNA modifications control all aspects of RNA life and are thus involved in several cellular processes and functions. The adenosine deamination to inosine is a process called RNA editing and is catalyzed by tasked enzymes, the adenosine deaminases acting on RNAs (ADARs). My laboratory has shown that adenosine deamination regulates cathepsin S mRNA stability by controlling the recruitment of the stabilizing RNA-binding protein called HuR onto the cathepsin S mRNA. Interestingly, we revealed a strong association of this RNA modification with vascular disease, documenting, for the first time, the importance of RNA modifications in cardiovascular system pathology. Adenosine methylation in the N6 position (m6A) is the most prominent RNA modification in eukaryotes and is catalyzed by a set of enzymes acting by being scavenged in another protein hub. This set of enzymes are termed METTL3, METT14 and WTAP. 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.
Identifying the dynamically methylated RNAs of our cells and characterizing how m6A mediates a precise adjustment of RNA life to environmental stimuli will undoubtedly contribute towards the understanding of human disease. This may open the road to the development of new disease biomarkers and therapeutic strategies. MODVASC aims to explore the role of m6A RNA methylation in vascular endothelial cell function and in vascular disease. Specifically, MODVASC will characterize: a) the endothelial methylome, b) the regulatory effects of m6A RNA methylation in endothelial gene expression, c) the role of m6A RNA methylation machinery in vascular development and maintenance of vascular function and d) the regulation of m6A RNA methylation in vascular disease.
The first half of the funding period progressed in accordance with the suggested timeline and milestones presented in the agreed proposal. We identified the methylated RNA molecules by m6A-RNA immunoprecipitation followed by RNA-sequencing of endothelial cell RNA. We next discovered with the help of deep RNA sequencing 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. We identified several cellular processes that may be affected by m6A RNA methylation. By using a combination of gain and loss of m6A enzymes function experiments, we have established the pivotal involvement of m6A in endothelial cell function. Furthermore, we have increased our understanding with regards to the specific molecular mechanism through which m6A regulates endothelial cell function. This was enabled by the transcriptome-wide analysis we performed in endothelial cells using the methylated RNA-sequencing technology established in my laboratory. Our first studies show that m6A RNA methylation may be critically involved in atherosclerotic cardiovascular disease and currently we conduct extensive mechanistic studies. The in vivo relevance of our findings will be further elucidated during the second half of our funding period.
Our findings until now reveal the importance of m6A RNA methylation for the endothelial cell homeostasis and function, which was the main goal of MODVASC, thus expanding the current knowledge. In the last part of MODVASC, we aim to enhance our understanding towards:
1) the in vivo implications of m6A RNA methylation-associated vascular endothelial homeostasis using the new genetic model we generated.
2) the elucidation of potentially common regulatory pathways controlling endothelial cell homeostasis and disease by using the established transcriptome-wide technologies.
3) the clinical value of m6A RNA methylation in atherosclerotic cardiovascular disease as well as inflammatory vascular diseases.
1) the in vivo implications of m6A RNA methylation-associated vascular endothelial homeostasis using the new genetic model we generated.
2) the elucidation of potentially common regulatory pathways controlling endothelial cell homeostasis and disease by using the established transcriptome-wide technologies.
3) the clinical value of m6A RNA methylation in atherosclerotic cardiovascular disease as well as inflammatory vascular diseases.