Abstract 12880: Neddylation Regulates Murine Cardiomyocyte Maturation

Abstract

From birth to adulthood cardiomyocytes (CMs) undergo a complex maturation process characterized by drastic changes in cell size, ultrastructure, metabolism and gene expression. The molecular mechanisms regulating CM maturation remain poorly defined. Neddylation is a novel post-translational modification that covalently attaches a small ubiquitin-like protein, NEDD8, to target proteins. We found that deletion of NAE1, a critical NEDD8 E1 enzyme that triggers neddylation, in embryonic and perinatal mouse hearts via Nkx2.5 Cre and aMHC Cre , respectively, led to myocardial hypoplasia, heart failure and early lethality. The severe cardiac phenotypes are accompanied by disorganized sarcomere structure and defective fetal-to-adult gene transition, suggesting a possible role for neddylation in CM maturation. To rule out the confounding effect of secondary changes caused by perturbed CM differentiation and cardiac stress response in the mutant hearts, we used AAV9-cTnT-Cre to achieve mosaic knockout of NAE1 in postnatal hearts. High-dose AAV9 deleted NAE1 in 80% CMs and caused heart failure, whereas low-dose AVV9 deleted NAE1 in 30% CMs and preserved heart function. In the context of preserved cardiac function, CMs lacking NAE1 showed severe disrupted transverse tubules (T-tubules) network, which was further exacerbated in hearts with modest dysfunction. In contrast, T-tubule morphology was largely maintained in CMs with intact NAE1 expression from low-dose AAV9 infected hearts, as well as in CMs with deletion of one allele of NAE1, indicating a key, cell-autonomous role of neddylation in T-tubule maturation. Biochemical and histological analyses further revealed dysregulated metabolic and sarcomeric gene expression and disrupted sarcomere ultrastructure in high-dose AAV9 infected hearts, supporting defective CM maturation. In vitro, pharmacological inhibition of neddylation also impaired myofibril assembly in isolated neonatal and adult CMs. Together, our data reveal an important role of neddylation in CM maturation in the developing heart, warranting further investigations of detailed posttranslational mechanisms underlying CM maturation.