Protein Tyrosine Phosphatase Shp2 and Neonatal Cardiac Innervation

Indiana University Indianapolis, Indianapolis IN

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Abstract

DESCRIPTION (provided by applicant): Using a unique conditional knockout (cKO) of Src homology protein tyrosine phosphatase 2 (Shp2) in post-migratory neural crest (NC) via our Periostin-Cre line, we generated a novel fully penetrant mouse model of diminished sympathetic cardiac innervation and neonatal death. These mice are indistinguishable from control littermates at birth and exhibit no structural cardiac anomalies observable by histology. However, 12 days after birth, cKOs develop sinus bradycardia as observed via electrocardiogram (ECG) and 100% of cKOs die by 3 weeks. The sinus bradycardia and prolonged PR intervals observed by ECG can predispose to arrhythmogenesis which may lead to the neonatal death observed in our Shp2 cKO mouse model. Shp2 is known to exert effects through many different signaling cascades and therefore plays diverse roles in many human diseases. Thus, we propose two aims to examine the role that Shp2 and downstream mediators play in our sympathetic innervation phenotype. Aim 1 will test our hypothesis that disruption within the NGF-SHP2- ERK signaling pathway is directly responsible for the diminished sympathetic innervation of the heart during development and define the functional consequences of that defective innervation as it relates to lethality. We will accomplish this aim via combined functional, molecular, and histological evaluation designed to characterize the Shp2 cKO phenotype from in utero stages to death at 3 weeks of age. Since no gross phenotype is evident until the first week of life, we expect that our longitudinal study will allowus to sequentially describe the events leading up to death and identify the role(s) that Shp2 and downstream targets play in this dynamic process. Our preliminary data as well as previously published works indicate a prominent role for pERK as a key downstream element involved in establishing sympathetic innervation. Therefore, Aim 2 will examine the direct role of pERK in mediating the Shp2 cKO phenotype. Aim 2 tests the hypothesis that lineage-specific restoration of Erk1/2 signaling will rescue the diminished sympathetic innervation phenotype. This will be accomplished by bypassing Shp2 to directly activate the downstream Erk1/2 pathway by expressing a constitutively active MEK1 (directly upstream of ERK) to replenish diminished pERK levels within the Shp2 lineage restricted cKO cells. Significantly, preliminary data demonstrates that Peri-Cre;caMEK1 mice on a wildtype background are hyperinnervated and induce both pERK and TH (sympathetic nerve marker) upregulation in vivo. Thus, we expect CAG-MEK1;Peri-Cre;Shp2f/f neonates will exhibit rescue of sympathetic innervation in the heart as assessed molecularly by TH levels via IHC and Western as well as functionally by HR measurements made by ECG and echocardiogram.