Adult mammals poorly regenerate injured hearts. In contrast, adult zebrafish possess a remarkable capacity to regenerate damaged hearts. Combined with available genetic tools, this capacity makes zebrafish a powerful system for deciphering the mechanisms underlying heart regeneration. Upon injury, the endocardium and epicardium are rapidly activated to secrete paracrine factors that facilitate heart regeneration. Heart regeneration research has focused mostly on identifying the secreted factors that trigger regenerative programs, yet little is known about the regulatory mechanisms controlling their expression, a proximal step in the regenerative process. Proper spatiotemporal regulation of these genes is essential for the intricate and tightly coordinated processes underlying cardiac regeneration. Thus, elucidating gene regulatory mechanisms governing injury-responsive gene expression will provide insights into potential therapeutic strategies based on stimulating paracrine effectors. Previously, we identified leptin b (lepb) as an injury-induced factor secreted by the endocardium. Importantly, we showed that the cardiac regeneration enhancer linked to lepb (LEN) is robustly activated by cardiac injury, maintains activity during regeneration, and then returns to a nave state upon completion of regeneration. We also identified a cardiac injury-responsive enhancer linked to interleukin 11a (il11a), which encodes a proregenerative cytokine structurally similar to lepb. We showed that zebrafish injury-responsive enhancers drive reporter-gene expression in injured mouse hearts, indicating that the mechanisms mediating injury-induced enhancer function are evolutionarily conserved. Based on these findings, we propose two aims to decipher gene regulatory mechanisms governing heart regeneration. The central hypothesis is that cardiac injury-responsive enhancers establish the transcriptional state of genes encoding paracrine factors that facilitate heart repair.
Aim 1 will determine the transcriptional mechanisms underlying cardiac injury-responsive enhancer activity. We discovered that cardiac LEN (cLEN) is not only activated by injury but also actively repressed in the absence of injury. We will employ transgenic assays, genetic and pharmacological approaches, as well as epigenomic and computational analyses to test the hypothesis that repression and activation mechanisms collectively determine cardiac injury-responsive enhancer function to confer injury-dependent cardiac gene transcription.
Aim 2 will use loss-of-function and gain-of-function studies to define how il11a controls heart regeneration and to dissect its injury-responsive enhancer. Elucidating the transcriptional mechanisms and function of crucial enhancer-regulated factors secreted upon cardiac injury will transform knowledge on how injury signals are transduced to facilitate heart regeneration.
Our knowledge of heart regeneration is increasing, yet our understanding of which genes are key drivers and how regeneration signals control these genes are incompletely defined. We propose to characterize how cardiac injury induces the expression of paracrine factors that facilitate heart regeneration. Our results will have profound implications for understanding the heart regenerative process and developing innovative strategies to promote heart regeneration in mammals.
