Abstract P1064: Hippo Signaling Pathway Maintains Sinoatrial Node Homeostasis

Abstract

The sinoatrial node (SAN) functions as pacemaker of the heart to initiate and drive rhythmic heartbeats. The Hippo signaling pathway is a fundamental pathway for heart development and regeneration. Although abnormalities of Hippo pathway are associated with cardiac arrhythmias in human patients, yet its role in the SAN is unknown. We investigated key regulators of the Hippo pathway in the sinoatrial node by conditionally inactivating the Hippo signaling kinases Lats1 and Lats2 using the tamoxifen-inducible, cardiac conduction system (CCS)-specific Cre driver Hcn4 CreERT2 and Lats1 and Lats2 conditional knockout (CKO) alleles. In addition, the Hippo signaling effectors Yap and Taz were conditionally inactivated in the sinoatrial node. To determine the function of the Hippo signaling pathway in the sinoatrial node and other CCS components, we conducted a series of physiological and molecular experiments including telemetry ECG recording, echocardiography, Masson’s Trichrome staining, calcium imaging, immunostaining, Cleavage Under Targets and Tagmentation (CUT&Tag) sequencing using antibodies against Yap1 or H3K4me3, qRT-PCR, and Western blotting. We found that Lats1/2 inactivation caused severe sinoatrial node dysfunction (SND; sick sinus syndrome). Compared to the controls, Lats1/2 CKO mutants exhibited dysregulated calcium handling and increased fibrosis in the sinoatrial node, indicating Lats1/2 function through both cell-autonomous and non-cell-autonomous mechanisms. Notably, the Lats1/2 CKO phenotype was rescued by genetic deletion of Yap and Taz in the CCS, and these rescued mice had normal sinus rhythm and reduced fibrosis of the sinoatrial node, indicating that Lats1/2 function through Yap and Taz . CUT&Tag sequencing data showed that Yap regulates genes critical for calcium homeostasis such as Ryr2 and genes encoding paracrine factors important in intercellular communication and fibrosis induction such as Tgf-β1 and Tgf-β3 . Consistently, Lats1/2 CKO mutants had decreased Ryr2 expression and increased Tgf-β1 and Tgf-β3 expression compared with control mice. We reveal for the first time that the canonical Hippo-Yap pathway has a pivotal role in functional homeostasis of the sinoatrial node. We also provide novel insight into the genetic regulation of sinoatrial node homeostasis and identify new potential therapeutic targets to improve recovery from cardiac arrhythmias.