Congenital heart disease (CHD) is the most common type of birth defect. Valvuloseptal defects, which result from aberrant endocardial cushion development, comprise up to one third of CHD, and also are a significant cause of adult morbidity/mortality. Although progress has been made in delineating single gene defects that perturb valvulogenesis in mouse and man, how these genes regulate development, and how normal development is altered by disease-associated mutations, remain largely unknown. Protein tyrosine phosphorylation, controlled by protein-tyrosine kinases (PTKs) and protein-tyrosine phosphatases (PTP), is a key cellular regulatory mechanism., Many growth factor receptors are transmembrane PTKs, and several are implicated in valvulogenesis. Vascular endothelial growth factor (VEGF), produced by myocardjal cells, inhibits endocardial-mesechymal transition (EMT), a key initial event in which specialized endocardia! cells transform into mesenchymal cells that invade the cushion matrix and proliferate. ErbB3, most likely partnering with ErbB2 (HER2) and responding to heregulin (HRG), promotes EMT and/or mesenchymal proliferation. Conversely, heparin-binding EGF (HB-EGF), acting via the EGFR (ErbB1), terminates mesenchymal cell proliferation and plays a key role in valve remodeling. The non-receptor PTP, Shp2 (PTPN11), also plays an essential role in valve development. Shp2 deficiency enhances the effect of EGFR loss of function in mice, resulting in abnormal valve thickening. Furthermore, ~50% of cases of the autosomal dominant disorder Nponan syndrome (NS), the most common non-chromosomal cause of CHD, is caused by PTPN11 mutations. Structural, enzymologic, and biochemical studies, and a mouse NS model generated in our laboratory, indicate that NS mutations are gain-of-function alleles that enhance Erk MAP kinase activation in developing cardiac cushions. PTPN11 mutations also cause LEOPARD syndrome (LS), which exhibits an overlapping spectrum of cardiac defects. But we showed recently that unlike NS alleles, LS mutants are PTP-inactive, and act as dominant negative mutants that impair Erk activation. Based on these data, we hypothesize that NS and LS mutations cause similar cardiac valve defects by acting in opposing ways on distinct RTK pathways at different times during valvulogenesis. We propose a combined biochemical, cell biological and genetic approach to address key questions about the pathogenesis of cardiac defects caused by human PTPN11 mutations.
Aim 1 will use an inducible knock-in approach to determine the cell type and developmental interval in which NS mutants act to cause valvuloseptal defects.
In Aim 2, we will study an allelic series of Shp2 knock-in alleles, asking if the specific PTPN11 mutation affects the NS cardiac phenotype, and generate a knock-in mouse model of LS to test the hypothesis that LS mutants act later during valvulogenesis to inhibit EGFR signaling/remodeling. Finally, Aim 3 will use a combination of mouse models, explant assays, and pharmacologic agents to test the hypothesis that NS alleles enhance ErbB2/3 signaling to the Ras/Erk pathway, thereby enhancing EMT and possibly mesenchymal proliferation. Our results should yield new insights into the pathogenesis of CHD, and may have important implications for the therapy of NS and LS.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL083273-04
Application #
7789554
Study Section
Cardiovascular Differentiation and Development Study Section (CDD)
Program Officer
Schramm, Charlene A
Project Start
2007-07-16
Project End
2011-04-30
Budget Start
2010-05-01
Budget End
2011-04-30
Support Year
4
Fiscal Year
2010
Total Cost
$270,000
Indirect Cost
Name
University Health Network
Department
Type
DUNS #
208469486
City
Toronto
State
ON
Country
Canada
Zip Code
M5 2-M9
Yin, Jiani C; Platt, Mathew J; Tian, Xixi et al. (2017) Cellular interplay via cytokine hierarchy causes pathological cardiac hypertrophy in RAF1-mutant Noonan syndrome. Nat Commun 8:15518
Cordeddu, Viviana; Yin, Jiani C; Gunnarsson, Cecilia et al. (2015) Activating Mutations Affecting the Dbl Homology Domain of SOS2 Cause Noonan Syndrome. Hum Mutat 36:1080-7
Winter, Georg E; Rix, Uwe; Carlson, Scott M et al. (2012) Systems-pharmacology dissection of a drug synergy in imatinib-resistant CML. Nat Chem Biol 8:905-912
Wu, Xue; Yin, Jiani; Simpson, Jeremy et al. (2012) Increased BRAF heterodimerization is the common pathogenic mechanism for noonan syndrome-associated RAF1 mutants. Mol Cell Biol 32:3872-90
Wu, Xue; Simpson, Jeremy; Hong, Jenny H et al. (2011) MEK-ERK pathway modulation ameliorates disease phenotypes in a mouse model of Noonan syndrome associated with the Raf1(L613V) mutation. J Clin Invest 121:1009-25
Marin, Talita M; Keith, Kimberly; Davies, Benjamin et al. (2011) Rapamycin reverses hypertrophic cardiomyopathy in a mouse model of LEOPARD syndrome-associated PTPN11 mutation. J Clin Invest 121:1026-43
Lee, Icksoo; Pecinova, Alena; Pecina, Petr et al. (2010) A suggested role for mitochondria in Noonan syndrome. Biochim Biophys Acta 1802:275-83
Stewart, Rodney A; Sanda, Takaomi; Widlund, Hans R et al. (2010) Phosphatase-dependent and -independent functions of Shp2 in neural crest cells underlie LEOPARD syndrome pathogenesis. Dev Cell 18:750-62
Araki, Toshiyuki; Chan, Gordon; Newbigging, Susan et al. (2009) Noonan syndrome cardiac defects are caused by PTPN11 acting in endocardium to enhance endocardial-mesenchymal transformation. Proc Natl Acad Sci U S A 106:4736-41
Kontaridis, Maria I; Yang, Wentian; Bence, Kendra K et al. (2008) Deletion of Ptpn11 (Shp2) in cardiomyocytes causes dilated cardiomyopathy via effects on the extracellular signal-regulated kinase/mitogen-activated protein kinase and RhoA signaling pathways. Circulation 117:1423-35