Molecular Medicine Epistatic Rescue of Nkx2.5 Adult Cardiac Conduction Disease Phenotypes by Prospero-Related Homeobox Protein 1 and HDAC3

Rationale: Nkx2.5 is one of the most widely studied cardiac-specific transcription factors, conserved from flies to man, with multiple essential roles in both the developing and adult heart. Specific dominant mutations in NKX2.5 have been identified in adult congenital heart disease patients presenting with conduction system anomalies and recent genome-wide association studies implicate the NKX2.5 locus, as causative for lethal arrhythmias (" sudden cardiac death ") that occur at a frequency in the population of 1 in 1000 per annum worldwide. Haploinsufficiency for Nkx2.5 in the mouse phenocopies human conduction disease pathology yet the phenotypes, described in both mouse and man, are highly pleiotropic, implicit of unknown modifiers and/or factors acting in epistasis with Nkx2.5/NKX2.5. Objective: To identify bone fide upstream genetic modifier(s) of Nkx2.5/NKX2.5 function and to determine epistatic effects relevant to the manifestation of NKX2.5-dependent adult congenital heart disease. Conclusions: Here we identify Prox1 as a direct upstream modifier of Nkx2.5 in the maintenance of the adult conduction system and rescue of Nkx2.5 conduction disease phenotypes. This study is the first example of rescue of Nkx2.5 function and establishes a model for ensuring electrophysiological function within the adult heart alongside insight into a novel Prox1-HDAC3-Nkx2.5 signaling pathway for therapeutic targeting in conduction disease. T he cardiac conduction system comprises a network of specialized cardiomyocytes that generate and propagate electric impulses throughout the muscle of the heart, punctuated by nodal tissue, to collectively pace and stimulate the synchronized contraction of the atria and ventricles and facilitate unidirectional blood flow. A number of cardiac transcription factors have been implicated in the maintenance of conduction system structure and function (reviewed in Ref. 1), of which Nkx2.5 is arguably the most well studied. However, despite a causative role for NKX2.5 in conduction disease and a substantial focus on Nkx2.5 in both the developing 2–5 and postnatal conduction system, 6 – 8 there have been no reports yielding insight into upstream modifier effects on Nkx2.5 activity or physiologically relevant factors identified to rescue Nkx2.5/NKX2.5 function in disease. The adult Nkx2.5 heterozygous mouse (Nkx2.5 Cre/ϩ) presents with background-dependent phenotypes, including atrial septal defects and atrioventricular (AV) conduction delay, which precisely mimic the clinical symptoms in humans with pathological mutations in NKX2.5. 9 Thus the mouse, in this instance, represents a bona fide model to study the manifestation of NKX2.5-derived human adult congenital heart disease. As such we focused on candidate factors that may regulate …