Investigation of NRG1 functions in endocardial cushion development in the heart

Albert Einstein College Of Medicine, Bronx NY

Investigators

Abstract

ABSTRACT: Heart valve disease affects 1-2% of the U.S. population, and some are due to congenital defects. The early embryonic heart consists of two layers, the inner endocardial layer, and the outer myocardial layer. These two cell types are separated by an extracellular matrix (ECM). During the formation of the valves, a subset of endocardial cells undergoes endocardial to mesenchymal transition (EndMT) and mesenchymal cells invade the ECM to form endocardial cushions as a key step in valve development. The atrioventricular canal (AVC) sits between the left ventricle and the atria and is one site of cushion formation in the early heart. The AVC cushions subsequently remodel into the atrioventricular valves. Neuregulin-1 (NRG1) is a signaling molecule that is strongly expressed in AVC endocardium throughout EndMT. However, the cellular and molecular mechanisms controlled by NRG1 in the AVC cushions are not fully understood. Cell-cell communication between the endocardium and myocardium leads to the initiation and progression of EndMT. NOTCH is a well-known example that is critical for these processes. NRG1 and NOTCH pathways have a complex relationship in many contexts. Our study aims to understand the interaction of these two pathways in the AVC cushions, as well as uncover novel transcriptional networks downstream of Nrg1 in cushion development. We will use two different Cre mouse lines, Tie2-Cre and Nfatc1Cre that are active in the endocardium, starting at different stages of mouse development, to inactivate Nrg1. Both lines are necessary to understand distinct roles of Nrg1 in initiation (Tie2- Cre) and progression of EndMT (Nfatc1Cre). We will look at the cardiac phenotypes and the interactions between NRG1 and NOTCH in vivo and using in vitro cell culture methods. My preliminary data shows that Nfatc1Cre/+ Nrg1f/f mutant embryos have hypoplastic cushions and I am comparing these with Tie2-Cre Nrg1f/f mutants. I hypothesize that NRG1 facilitates cell-cell communication between the cells of the AVC cushion to promote the initiation and progression of EndMT. Changes in cell signaling and communication will be reflected in transcriptional alteration including that of downstream effector genes. I have recently generated single-cell RNA- sequencing data of control and Nfatc1Cre/+ Nrg1f/f mutant hearts at E9.5. I plan to compare gene expression changes of control, Tie2-Cre, and Nfatc1Cre mutant hearts to understand more about cell-cell communication regulated by NRG1. ERBB receptors, which are activated by NRG1, are expressed on all three cell types of the AVC: endocardial, mesenchymal, and myocardial cells. I expect to identify new cellular mechanisms that function downstream of NRG1 in the endocardial cushions. Aims 1 and 2 will combine developmental genetics with single cell RNA-sequencing to understand how NRG1 signaling, and downstream transcriptional changes regulates this crosstalk.