Genome-Wide Dynamics of Nascent Noncoding RNA Transcription in Porcine Heart After Myocardial Infarction

Abstract

Background— Microarrays and RNA sequencing are widely used to profile transcriptome remodeling during myocardial ischemia. However, the steady-state RNA analysis lacks in sensitivity to detect all noncoding RNA species and does not provide separation between transcriptional and post-transcriptional regulations. Here, we provide the first comprehensive analysis of nascent RNA profiles of mRNAs, primary micro-RNAs, long noncoding RNAs, and enhancer RNAs in a large animal model of acute infarction. Methods and Results— Acute infarction was induced by cardiac catheterization of domestic swine. Nuclei isolated from healthy, border zone, and ischemic regions of the affected heart were subjected to global run-on sequencing. Global run-on sequencing analysis indicated that half of affected genes are regulated at the level of transcriptional pausing. A gradient of induction of inflammatory mediators and repression of peroxisome proliferator-activated receptor signaling and oxidative phosphorylation was detected when moving from healthy toward infarcted area. In addition, we interrogated the transcriptional regulation of primary micro-RNAs and provide evidence that several arrhythmia-related target genes exhibit repression at post-transcriptional level. We identified 450 long noncoding RNAs differently regulated by ischemia, including novel conserved long noncoding RNAs expressed in antisense orientation to myocardial transcription factors GATA-binding protein 4, GATA-binding protein 6, and Krüppel-like factor 6. Finally, characterization of enhancers exhibiting differential expression of enhancer RNAs pointed a central role for Krüppel-like factor, MEF2C, ETS, NFY, ATF, E2F2, and NRF1 transcription factors in determining transcriptional responses to ischemia. Conclusions— Global run-on sequencing allowed us to follow the gradient of gene expression occurring in the ischemic heart and identify novel noncoding RNAs regulated by oxygen deprivation. These findings highlight potential new targets for diagnosis and treatment of myocardial ischemia.

Dark Side of the Deep Heart

“…Specifically, they derived this information from the difference of Pol II density between the promoter region and the gene body, respectively, from 0 to 200 and from 200 bp to the end of the gene. 12 Indeed, they found an increase of pausing for repressed transcripts associated to mitochondrial function and muscle contraction and a decrease of pausing in transcribed transcripts involved in inflammation, cell motility, and necrosis, giving an evident view of the transcriptional dynamics ongoing in an ischemic heart. 12 Their experimental approach enriched our knowledge about the transcriptome remodeling occurring during myocardial ischemia exploiting the high resolution of GRO-Seq and indicated to focus the attention especially on ncRNA species.…”

“…12 Indeed, they found an increase of pausing for repressed transcripts associated to mitochondrial function and muscle contraction and a decrease of pausing in transcribed transcripts involved in inflammation, cell motility, and necrosis, giving an evident view of the transcriptional dynamics ongoing in an ischemic heart. 12 Their experimental approach enriched our knowledge about the transcriptome remodeling occurring during myocardial ischemia exploiting the high resolution of GRO-Seq and indicated to focus the attention especially on ncRNA species. The authors were able to identify the primary miRNAs expressed into the ischemic heart after their expression changes after ischemia, pointing out for the first time the transcriptional regulation of miRNAs 12 in this condition.…”

“…In a study published in the present issue of Circulation: Cardiovascular Genetics, Kaikkonen et al 12 report the results obtained from GRO-Seq in a large animal model of acute infarction. The authors adopted the GRO-Seq to determine the profiles of nascent RNAs associated to myocardial infarction, scanning the transcriptional activity occurring in nuclei derived from different areas of the ischemic heart at 1 day after ischemia.…”

“…The authors adopted the GRO-Seq to determine the profiles of nascent RNAs associated to myocardial infarction, scanning the transcriptional activity occurring in nuclei derived from different areas of the ischemic heart at 1 day after ischemia. 12 By this approach, they found the induction of inflammatory markers (like tumor necrosis factor [TNF] and transforming growth factor β1 [TGFβ1]) and the downmodulation of genes involved in oxidative phosphorylation, cardiac muscle contraction, and peroxisome proliferator-activated receptor (PPAR) signaling. Interestingly, this study revealed for the first time the presence of a gradient of differentially regulated transcripts moving from the healthy myocardium to the hypoxic border zone up to the deep ischemic area.…”

“…Interestingly, this study revealed for the first time the presence of a gradient of differentially regulated transcripts moving from the healthy myocardium to the hypoxic border zone up to the deep ischemic area. 12 Moreover, they took advantage from GRO-Seq to calculate the transcriptional pausing ratio. Specifically, they derived this information from the difference of Pol II density between the promoter region and the gene body, respectively, from 0 to 200 and from 200 bp to the end of the gene.…”