Cardiomyocyte Lineage Specification in Adult Human Cardiac Precursor Cells Via Modulation of Enhancer-Associated Long Noncoding RNA Expression

SummaryThe mechanisms controlling differentiation in adult cardiac precursor cells (CPCs) are still largely unknown. In this study, CPCs isolated from the human heart were found to produce predominantly smooth muscle cells but could be redirected to the cardiomyocyte fate by transient activation followed by inhibition of NOTCH signaling. NOTCH inhibition repressed MIR-143/145 expression, and blocked smooth muscle differentiation. Expression of the microRNAs is under control of CARMEN, a long noncoding RNA associated with an enhancer located in the MIR-143/145 locus and target of NOTCH signaling. The CARMEN/MIR-145/143 axis represents, therefore, a promising target to favor production of cardiomyocytes in cell replacement therapies

The long noncoding RNA Wisper controls cardiac fibrosis and remodeling

Long noncoding RNAs (lncRNAs) are emerging as powerful regulators of cardiac development and disease. However, our understanding of the importance of these molecules in cardiac fibrosis is limited. Using an integrated genomic screen, we identified Wisper (Wisp2 super-enhancer–associated RNA) as a cardiac fibroblast–enriched lncRNA that regulates cardiac fibrosis after injury. Wisper expression was correlated with cardiac fibrosis both in a murine model of myocardial infarction (MI) and in heart tissue from human patients suffering from aortic stenosis. Loss-of-function approaches in vitro using modified antisense oligonucleotides (ASOs) demonstrated that Wisper is a specific regulator of cardiac fibroblast proliferation, migration, and survival. Accordingly, ASO-mediated silencing of Wisper in vivo attenuated MI-induced fibrosis and cardiac dysfunction. Functionally, Wisper regulates cardiac fibroblast gene expression programs critical for cell identity, extracellular matrix deposition, proliferation, and survival. In addition, its association with TIA1-related protein allows it to control the expression of a profibrotic form of lysyl hydroxylase 2, implicated in collagen cross-linking and stabilization of the matrix. Together, our findings identify Wisper as a cardiac fibroblast–enriched super-enhancer–associated lncRNA that represents an attractive therapeutic target to reduce the pathological development of cardiac fibrosis in response to MI and prevent adverse remodeling in the damaged heart

Genome-wide profiling of the cardiac transcriptome after myocardial infarction identifies novel heart-specific long non-coding RNAs

Aim Heart disease is recognized as a consequence of dysregulation of cardiac gene regulatory networks. Previously, unappreciated components of such networks are the long non-coding RNAs (lncRNAs). Their roles in the heart remain to be elucidated. Thus, this study aimed to systematically characterize the cardiac long non-coding transcriptome post-myocardial infarction and to elucidate their potential roles in cardiac homoeostasis. Methods and results We annotated the mouse transcriptome after myocardial infarction via RNA sequencing and ab initio transcript reconstruction, and integrated genome-wide approaches to associate specific lncRNAs with developmental processes and physiological parameters. Expression of specific lncRNAs strongly correlated with defined parameters of cardiac dimensions and function. Using chromatin maps to infer lncRNA function, we identified many with potential roles in cardiogenesis and pathological remodelling. The vast majority was associated with active cardiac-specific enhancers. Importantly, oligonucleotide-mediated knockdown implicated novel lncRNAs in controlling expression of key regulatory proteins involved in cardiogenesis. Finally, we identified hundreds of human orthologues and demonstrate that particular candidates were differentially modulated in human heart disease. Conclusion These findings reveal hundreds of novel heart-specific lncRNAs with unique regulatory and functional characteristics relevant to maladaptive remodelling, cardiac function and possibly cardiac regeneration. This new class of molecules represents potential therapeutic targets for cardiac disease. Furthermore, their exquisite correlation with cardiac physiology renders them attractive candidate biomarkers to be used in the clini

Regulatory characterisation of the novel gene, myocyte stress 1

Myocyte stress 1 (ms1) is a striated muscle actin binding protein required for muscle specific activity of the myocardin related transcription factor (MRTF)/serum response factor (SRF) transcriptional pathway. Previous work in our group demostrated that cardiac ms1 is transiently up-regulated after pressure overload suggesting a possible role in the initial signalling of the hypertrophic response. Subsequent studies have supported this and demonstrated that ms1 plays an important role in cardiac development and physiology. To date, little is known about the molecular mechanisms that govern striated muscle specific expression of ms1. In order to delineate ms1 regulation and function, a strategy of comparative in silico analysis coupled with experimental characterisation was used. In silico analysis identified four genomic intervals of potential regulatory function designated PP, UP1, UP2 and UP3. Using in vitro and in vivo appraoches, important cardiac regulatory roles for these domains were defined. The PP domain represents the basal promoter and is required for all regulatory contexts. This domain serves to intergrate context specific regulatory signals from the distal UP2 and UP3 domains. Within the heart the cardiac transcription factor GATA4, and the calcineurin singalling pathway confer cardiac regulatory function on the PP, UP2 and UP3 domains. Within skeletal muscle, MyoD binding sites within the PP and UP1 domain were identified, which mediate temporal induction of ms1 during myogenesis. Both cardiac and skeletal regulatory processes were dependent on epigenetic phenomena with histone acetylation being a major determinant for ms1 expression. Collectively, these findings demonstrate that ms1 transcriptional regulation is mediated by the complex interplay of context specific regulatory domains and binding factors. Therefore through ms1, important striated muscle gene regulatory networks (GRNs) (GATA4, Mef2 and MyoD GRNs) can integrate with SRF, thus exquisitely controlling biological processes in muscle. It is proposed that dysregulation of ms1 expression may result in pathological phenotypes. Therefore, the insights obtained here may allow for the therapeutic manipulation of ms1 expression in pathological settings and potentially lead to effective paliatation of such phenotypes

Getting to the heart of genomic dark matter

<p>Slides to my recent presentation at the ESC Congress in Barcelona on the genomewide profiling of novel lncRNAs in the mammalian heart after myocardial infraction</p

Sequence deep or go home - LncRNA dicovery

<p>Here we demonstrate using simulations how de novo discovery of novel cardiac lncRNAs (which have unique functional and regulatory characteristics) requires extremely deep RNA-Seq. 300-400 million 100bp-PE reads is what we suggest for identifying cardiac specific lncRNAs with highly specialised cardiogenic functions</p

Unzipping genomic 'dark matter' in the heart - New l(i)ncs to development and disease

<p>Slides from my presentation at the European Cardiovascular Science Center in Frankfurt, Germany.</p

Small and long non-coding RNAs in cardiac homeostasis and regeneration

Cardiovascular diseases and in particular heart failure are major causes of morbidity and mortality in the Western world. Recently, the notion of promoting cardiac regeneration as a means to replace lost cardiomyocytes in the damaged heart has engendered considerable research interest. These studies envisage the utilization of both endogenous and exogenous cellular populations, which undergo highly specialized cell fate transitions to promote cardiomyocyte replenishment. Such transitions are under the control of regenerative gene regulatory networks, which are enacted by the integrated execution of specific transcriptional programs. In this context, it is emerging that the non-coding portion of the genome is dynamically transcribed generating thousands of regulatory small and long non-coding RNAs, which are central orchestrators of these networks. In this review, we discuss more particularly the biological roles of two classes of regulatory non-coding RNAs, i.e. microRNAs and long non-coding RNAs, with a particular emphasis on their known and putative roles in cardiac homeostasis and regeneration. Indeed, manipulating non-coding RNA-mediated regulatory networks could provide keys to unlock the dormant potential of the mammalian heart to regenerate. This should ultimately improve the effectiveness of current regenerative strategies and discover new avenues for repair. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Cardiac Pathways of Differentiation, Metabolism and Contraction