Human Shp2 (Ptpn11) mutations and cardiac valve development

人类 Shp2 (Ptpn11) 突变与心脏瓣膜发育

• 批准号: 7629640
• 负责人: BENJAMIN G. NEEL
• 金额: $ 27万
• 依托单位: UNIVERSITY HEALTH NETWORK
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-16 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7629640
• 关键词: Address; Adult; Affect; Alleles; Apoptosis; Binding; Biochemical; Biological; Biological Assay; Cardiac; Cardiac Jelly; Cell Proliferation; Cell Shape; Cells; Complex; Congenital Abnormality; Congenital Heart Defects; Data; Defect; Development; Disease; Dominant-Negative Mutation; EGF gene; ERBB2 gene; Endocardium; Epidermal Growth Factor Receptor; Epithelial; ErbB Receptor Family Protein; ErbB4 gene; Event; Exhibits; Extracellular Matrix; Genes; Genetic; Growth Factor Receptors; Heart Atrium; Heart Valves; Heparin Binding; Heregulin; Human; Integral Membrane Protein; Invaded; Knock-in Mouse; LEOPARD Syndrome; Laboratories; MAP Kinase Modules; Medical; Mesenchymal; Mesenchyme; Mitogen-Activated Protein Kinases; Modeling; Morbidity - disease rate; Mus; Mutate; Mutation; Myocardial; Myocardium; Newborn Infant; Noonan Syndrome; PTPN11 gene; Pathogenesis; Pathway interactions; Peptides; Phenotype; Play; Process; Production; Proliferating; Protein Tyrosine Kinase; Protein Tyrosine Phosphatase; Proteins; Publishing; Receptor Protein-Tyrosine Kinases; Research Personnel; Role; Series; Signal Pathway; Signal Transduction; Single-Gene Defect; Specific qualifier value; Staging; Structure; Syndrome; Tamoxifen; Testing; Time; Tyrosine Phosphorylation; Variant; Vascular Endothelial Growth Factors; Ventricular septum; base; cell transformation; cell type; congenital heart disorder; formycin triphosphate; gain of function mutation; human PTPRT protein; insight; loss of function; man; mortality; mouse model; mutant; notch protein; programs; receptor; recombinase; src Homology Region 2 Domain

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：先天性心脏病（CHD）是最常见的先天缺陷类型。瓣膜缺陷由异常的心内膜垫发育造成，占CHD的三分之一，也是成人发病率/死亡率的重要原因。尽管在描述小鼠和人类驱动瓣膜发生的单个基因缺陷中取得了进展，这些基因如何调节发育以及如何通过疾病相关的突变改变正常发育，但仍在很大程度上未知。蛋白酪氨酸磷酸化，由蛋白酪氨酸激酶（PTK）和蛋白质 - 酪氨酸磷酸酶（PTP）控制，是一种关键的细胞调节机制。许多生长因子受体都是跨膜PTK，并且有几种与瓣膜发生有关。由Myocardjal细胞产生的血管内皮生长因子（VEGF）抑制心内膜 - 介体过渡（EMT），这是一个主要的初始事件，其中专门的心内膜心脏内膜病！细胞转变为侵入垫子基质并增殖的间充质细胞。 ERBB3很可能与ERBB2（HER2）合作并响应此处的糖蛋白（HRG），可促进EMT和/或间充质增殖。相反，通过EGFR（ERBB1）作用的肝素结合EGF（HB-EGF）终止间充质细胞增殖，并在阀重塑中起关键作用。非受体PTP SHP2（PTPN11）在阀发育中也起着至关重要的作用。 SHP2缺乏增强了EGFR功能在小鼠中的影响，从而导致异常瓣膜增厚。此外，约有50％的常染色体显性疾病NPONAN综合征（NS）是CHD最常见的非染色体原因，是由PTPN11突变引起的。结构，酶学和生化研究以及我们实验室中产生的小鼠NS模型表明，NS突变是功能收益等位基因，可增强ERK MAP激酶在发育中的心脏库中的激活。 PTPN11突变还引起豹综合征（LS），该突变表现出心脏缺陷的重叠频谱。但是我们最近表明，与NS等位基因不同，LS突变体是PTP无效的，并且是损害ERK激活的主要阴性突变体。基于这些数据，我们假设NS和LS突变通过在瓣膜发生期间在不同时间对不同的RTK途径作用，从而导致相似的心脏瓣膜缺陷。我们提出了一种合并的生化，细胞生物学和遗传学方法，以解决有关人PTPN11突变引起的心脏缺陷发病机理的关键问题。 AIM 1将使用诱导的敲门方法来确定NS突变体作用引起瓣膜缺陷的细胞类型和发育间隔。在AIM 2中，我们将研究一系列等位基因SHP2敲入等位基因，询问特定的PTPN11突变是否影响NS心脏表型，并产生LS的敲入小鼠模型，以检验LS突变体在瓣膜发生过程中后来起作用以抑制EGFR信号/重塑的假设。最后，AIM 3将使用小鼠模型，外植体测定和药理剂的组合来测试NS等位基因增强对RAS/ERK途径的ERBB2/3信号传导的假设，从而增强EMT并可能增强间质增殖。我们的结果应该对CHD的发病机理产生新的见解，并可能对NS和LS的治疗具有重要意义。