Assessment of the Cardiac Noncoding Transcriptome by Single-Cell RNA Sequencing Identifies FIXER , a Conserved Profibrogenic Long Noncoding RNA

Abstract

BACKGROUND: Cardiac fibroblasts have crucial roles in the heart. In particular, fibroblasts differentiate into myofibroblasts in the damaged myocardium, contributing to scar formation and interstitial fibrosis. Fibrosis is associated with heart dysfunction and failure. Myofibroblasts therefore represent attractive therapeutic targets. However, the lack of myofibroblast-specific markers has precluded the development of targeted therapies. In this context, most of the noncoding genome is transcribed into long noncoding RNAs (lncRNAs). A number of lncRNAs have pivotal functions in the cardiovascular system. lncRNAs are globally more cell-specific than protein-coding genes, supporting their importance as key determinants of cell identity. METHODS: In this study, we evaluated the value of the lncRNA transcriptome in very deep single-cell RNA sequencing. We profiled the lncRNA transcriptome in cardiac nonmyocyte cells after infarction and probed heterogeneity in the fibroblast and myofibroblast populations. In addition, we searched for subpopulation-specific markers that can constitute novel targets in therapy for heart disease. RESULTS: We demonstrated that cardiac cell identity can be defined by the sole expression of lncRNAs in single-cell experiments. In this analysis, we identified lncRNAs enriched in relevant myofibroblast subpopulations. Selecting 1 candidate we named FIXER (fibrogenic LOX -locus enhancer RNA), we showed that its silencing limits fibrosis and improves heart function after infarction. FIXER mechanistically associates with CBX4, an E3 SUMO (small ubiquitin-like modifier) protein ligase and a transcription factor, target CBX4 to the promoter of the transcription factor RUNX1 to regulate its expression and thereby the whole fibrogenic gene program. FIXER is conserved in humans, supporting its translational value. CONCLUSIONS: Our results demonstrated that lncRNA expression is sufficient to identify the various cell types composing the mammalian heart. Focusing on cardiac fibroblasts and their derivatives, we identified lncRNAs uniquely expressed in myofibroblasts. In particular, the lncRNA FIXER represents a novel therapeutic target for cardiac fibrosis.

Cell type specific long non-coding RNA targets identified by integrative analysis of single-cell and bulk colorectal cancer transcriptomes

“…Firstly, lncRNAs’ low overall expression and incomplete annotation catalogues could potentially hinder their prioritization in scRNA-seq datasets. In future studies, this limitation can be addressed by performing de novo transcriptome assembly in matched bulk transcriptomes as was shown in other contexts 22 . Moreover, the functional experiments are focused on a subset of identified lncRNAs, and further studies are needed to explore the functional significance of other differentially expressed lncRNAs.…”

Enhanced TfR1 Recognition of Myocardial Injury after Acute Myocardial Infarction with Cardiac Fibrosis via Pre-Degrading Excess Fibrotic Collagen

“…Acute myocardial infarction (MI) is a severe clinical emergency in which necrotic areas are replaced by fibrous tissue due to the limited regenerative capacity of cardiomyocytes, resulting in ventricular remodeling with severe effects on cardiac function [1]. Myocardial remodeling after MI is an important contributor to cardiac dysfunction, increased complications, and increased mortality, and myocardial fibrosis plays a key role in this process [2].…”