Signaling by gain-of-function SHP-2 mutants in Noonan syndrome

描述（由申请人提供）：努南综合征（NS）是一种常染色体显性遗传病，发生的频率约为1:20 000活产。大约50%的NS患者包含人类PTPN11基因的功能获得突变，该基因编码含有SH2结构域的蛋白酪氨酸磷酸酶SHP-2。NS患者表现出多种临床表现，最明显的是先天性心脏病（CHD）。高达80%的NS患者发生冠心病，使得PTPN11/SHP-2突变成为冠心病最常见的非染色体原因。因此，酪氨酸磷酸化的改变是冠心病的基础。本研究的主要目标是揭示nsns相关的SHP-2突变如何引起冠心病的分子基础。尽管许多研究已经证实Ras/细胞外信号调节激酶1和2 （ERK1/2）通路的激活增强是ns介导的冠心病的原因，但ns相关SHP-2突变参与Ras/ERK1/2病理生理信号传导的确切机制尚不清楚。我们建议确定nsns相关SHP-2突变体的直接上游和下游靶点，并确定这些靶点是否参与nsns介导的冠心病的发展。在第一个目标中，我们已经确定了ns相关的SHP- 2突变体优先与含itim的跨膜糖蛋白相互作用。我们假设，ns相关的SHP-2突变体的膜邻近失调参与了引起Ras/ERK1/2信号传导的底物的混杂去磷酸化。这些ITIM/NS-SHP-2相互作用对Ras/ERK1/2信号的贡献将被定义。参与ns介导的Ras/ERK1/2活性的底物将因其参与ns相关的SHP-2突变信号而被鉴定和表征。在特定的目的二，ITIM含有跨膜糖蛋白已被确定为高酪氨酸磷酸化的小鼠NS模型。我们将识别ns诱导的酪氨酸激酶，并使用遗传和生化方法的结合来确定这种酪氨酸激酶是否会增强ERK1/2激活并随后导致ns相关的心脏缺陷。第三个目标将测试ns相关SHP-2突变体改变膜募集作为ns介导的心脏缺陷的决定因素的病理生理贡献。我们将通过采用遗传方法干扰NS-相关SHP-2向细胞膜的募集来实现这一目标。这些研究的完成将对ns相关SHP-2突变体在冠心病中的直接靶点产生新的见解，并可能揭示新的和已建立的信号分子在该病中的意想不到的作用。识别与冠心病相关的靶点也将揭示新的治疗策略模式，以及评估nsds相关冠心病的预后工具。

DESCRIPTION (provided by applicant): Noonan syndrome (NS) is an autosomal dominant disorder that occurs with a frequency of ~ 1:2,000 live births. Approximately 50% of NS patients contain a gain-of-function mutation in the human PTPN11 gene which encodes for the SH2 domain-containing protein tyrosine phosphatase, SHP-2. NS patients exhibit a diverse array of clinical manifestations, most notably, congenital heart disease (CHD). CHD occurs in up to 80% of NS patients, making PTPN11/SHP-2 mutations the most common non-chromosomal cause of CHD. Therefore, altered tyrosyl phosphorylation underlies the basis for CHD. The broad goal of this research is to uncover the molecular basis for how NS-associated SHP-2 mutations give rise to CHD. Although much work has established that enhanced activation of the Ras/extracellular signal-regulated kinases 1 and 2 (ERK1/2) pathway is causal to NS-mediated CHD the precise mechanisms through which NS-associated SHP-2 mutations engage in pathophysiological signaling to Ras/ERK1/2 remains unknown. We propose to identify the direct upstream and downstream targets of NS-associated SHP-2 mutants and determine if these targets are involved in the development of NS-mediated CHD. In the first aim, we have identified that NS-associated SHP- 2 mutants interact preferentially with ITIM-containing transmembrane glycoproteins. We hypothesize that dysregulated membrane proximity by NS-associated SHP-2 mutants engages promiscuous dephosphorylation of substrates that evoke Ras/ERK1/2 signaling. The contribution of these ITIM/NS-SHP-2 interactions to signal to Ras/ERK1/2 will be defined. The substrates involved in NS-mediated Ras/ERK1/2 activity will be identified and characterized for their involvement in NS-associated SHP-2 mutant signaling. In specific aim two, the ITIM containing transmembrane glycoproteins have been identified to be hypertyrosyl phosphorylated in a mouse model of NS. We will identify the NS-induced tyrosine kinase(s) and using a combination of genetic and biochemical approaches determine whether this tyrosine kinase(s) propagates enhanced ERK1/2 activation and subsequently NS-related cardiac defects. The third aim will test the pathophysiological contribution of altered membrane recruitment of NS-associated SHP-2 mutants as a determinant of NS-mediated cardiac defects. We will accomplish this by employing genetic approaches to interfere with the recruitment of NS- associated SHP-2 to the membrane. The completion of these studies will yield new insight into the direct targets of NS-associated SHP-2 mutants in CHD, and may reveal unanticipated roles, for new and established, signaling molecules in this disease. The identification of targets involved in CHD will also reveal new modes of therapeutic strategies in which to treat, and prognostic tools in which to evaluate, NS-related CHD.