Abstract
Reactivation of fetal-specific genes and isoforms occurs during heart failure. However, the underlying molecular mechanisms and the extent to which the fetal program switch occurs remains unclear. Limitations hindering transcriptome-wide analyses of alternative splicing differences (i.e. isoform switching) in cardiovascular system (CVS) tissues between fetal, healthy adult and heart failure have included both cellular heterogeneity across bulk RNA-seq samples and limited availability of fetal tissue for research. To overcome these limitations, we have deconvoluted the cellular compositions of 996 RNA-seq samples representing heart failure, healthy adult (heart and arteria), and fetal-like (iPSC-derived cardiovascular progenitor cells) CVS tissues. Comparison of the expression profiles revealed that reactivation of fetal-specific RNA-binding proteins (RBPs), and the accompanied re-expression of 1,523 fetal-specific isoforms, contribute to the transcriptome differences between heart failure and healthy adult heart. Of note, isoforms for 20 different RBPs were among those that reverted in heart failure to the fetal-like expression pattern. We determined that, compared with adult-specific isoforms, fetal-specific isoforms encode proteins that tend to have more functions, are more likely to harbor RBP binding sites, have canonical sequences at their splice sites, and contain typical upstream polypyrimidine tracts. Our study suggests that compared with healthy adult, fetal cardiac tissue requires stricter transcriptional regulation, and that during heart failure reversion to this stricter transcriptional regulation occurs. Furthermore, we provide a resource of cardiac developmental stage-specific and heart failure-associated genes and isoforms, which are largely unexplored and can be exploited to investigate novel therapeutics for heart failure.
Highlights
Heart failure is associated with increased expression of fetal-specific genes including cardiac ion channels [1], as well as fetal-specific isoforms of genes with pivotal roles in cardiac development, including TTN and SCN5A [2–4]
To overcome the limitations of sample heterogeneity and isoform detection, we have taken advantage of 786 GTEx samples collected from 352 adults [22] and 180 iPSCORE samples from 139 individuals [14]. Analyzing these 966 bulk RNA-seq samples representing cardiovascular system (CVS) tissues at two developmental stages and three distinct types, we found that the vast majority of RNA binding proteins (RBPs) are expressed at higher levels in the fetallike induced pluripotent stem cell (iPSC)-CVPC compared with adult CVS tissues and play a larger role regulating the expression of fetal-specific isoforms than adult-specific isoforms
The iPSC-CVPCs were associated with a higher cell division rate [27–29] and a lower pseudotime score [30], compared with the two adult CVS tissues (Fig S2F–S2H), which is congruent with the findings of earlier studies showing iPSC-CVPCs resemble fetal-like CVS tissue [14,16]
Summary
Heart failure is associated with increased expression of fetal-specific genes including cardiac ion channels [1], as well as fetal-specific isoforms of genes with pivotal roles in cardiac development, including TTN and SCN5A [2–4]. A transcriptome-wide analyses of RBP expression and alternative splicing differences in cardiovascular system (CVS) tissues between fetal and adult (healthy and diseased) stages, has yet to be conducted. Several limitations have hindered transcriptome-wide analyses of alternative splicing differences (i.e. isoform switching) between fetal and adult CVS tissues (heart and arteria). Heterogeneity across bulk RNA-seq samples of the same tissue type limits the power to identify gene and isoform expression differences between different tissues and development stages. Bulk RNA-seq experiments average gene expression across the population of cells in a given sample and thereby prevent the ability to capture isoform expression variability across different cell types. The limited availability of human fetal heart tissues for research makes it impossible to obtain the large sample sizes required for the statistical power to perform transcriptome-wide analyses of the reexpression of fetal-specific isoforms. We and other have shown that induced pluripotent stem cell (iPSC) derived cardiovascular progenitor cells (iPSC-CVPC) show transcriptomic, epigenomic, morphological, structural and functional properties of their fetal counterparts, including their cellular composition being a mixture of fetal-like cardiomyocytes and epicardial-derived cells (EPDCs) including myofibroblasts, vascular smooth muscle cells and endothelial cells, and can be used as a surrogate for fetal cardiac tissues for developmental, genetic and pharmacological studies [14,16–21]
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