Prolyl isomerase function during Jak Stat signaling in breast cancer

描述(申请人提供)：受体相关的酪氨酸激酶，如JAK2和Src，作为配体结合的近端介体。大量数据表明，这两种蛋白在乳腺癌的发病机制中都有重要作用，但导致其激活的机制尚不清楚。我们的研究集中在与乳腺癌相关的催乳素受体(PRLr)上，发现这些酶的激活需要肽性Pro异构酶(PPI)，亲环素A(CypA)。这些发现表明，CypA的PPI活性参与了这种受体-激酶复合体的构象重组，从而有助于其激活。更多的研究表明，PRLr、JAK2、Src和CypA之间形成的多聚体复合体是配体诱导的PRLr相关信号复合体激活所必需的。在翻译水平上，我们对CypA基因敲除模型的评估以及PPI抑制剂环孢素A(CsA)和NIM811在体外和体内成功应用于乳腺癌模型，进一步证实了这些结果。综上所述，我们的发现导致我们假设，这些蛋白质之间的分子间相互作用和CypA的PPI活性导致了配体结合后JAK2和Src的触发，并且这些事件与乳腺癌的发病机制高度相关。这一假说将在以下三个具体目标中得到验证：首先，将使用乳腺癌细胞内的突变、过表达和敲除方法来评估PRLr/JAK2/Src/CypA中蛋白质-蛋白质相互作用的功能作用。其次，将使用体外激酶以及可溶性和固态核磁共振谱来评估受CypA PPI活性调节的PRLr/JAK2/Src复合体的构象状态。第三，将在基于遗传和异种移植的小鼠模型中使用获得和功能丧失的方法，以评估PRLr复合体中的蛋白质相互作用和PPI功能在乳腺癌发病机制中的作用。这些研究具有重要的意义，因为这将为我们理解受PPI活性CypA调控的受体JAK2/Src激活过程中的结构/功能关系提供分子基础，并通过乳腺癌的细胞到小鼠模型来解释这些发现。

DESCRIPTION (provided by applicant): Receptor-associated tyrosine kinases, such as Jak2 and Src, serve as proximal mediators of ligand binding. Abundant data indicate that both of these kinases significantly contribute to the pathogenesis of breast cancer, and yet the mechanisms leading to their activation has remained uncertain. Our research, which has focused on the breast cancer-relevant receptor for prolactin (PRLr), has revealed that the peptidly prolyl isomerase (PPI), cyclophilin A (CypA) is required for the activation of these kinases. These findings would indicate that PPI activity of CypA is involved in a conformational restructuring of this receptor-kinase complex that contributes to its activation. Additional studie have revealed that the formation of a multimeric complex between the PRLr, Jak2, Src, and CypA is necessary of ligand-induced activation of the PRLr-associated signaling complex. At a translational level, these results have been further corroborated by our evaluation of a CypA knockout model and the successful use in breast cancer models of the PPI inhibitors cyclosporine A (CsA) and NIM811 both in vitro and in vivo. Taken together, our findings lead us to hypothesize that the intermolecular interactions between these proteins and the PPI activity of CypA result in the triggering of Jak2 and Src following ligand engagement, and that such events are highly relevant to the pathogenesis of breast cancer. This hypothesis will be tested in three specific aims, as follows: First, the functional role of protein- protein interactions withinthe PRLr/Jak2/Src/CypA will be evaluated using mutagenic, overexpression, and knockdown approaches within breast cancer cells. Second, in vitro kinase and soluble and solid state NMR spectroscopy will be used to assess conformer status with the PRLr/Jak2/Src complex as regulated by the PPI activity of CypA. Third, both gain- and loss-of-function approaches will be used in genetic and xenograft-based murine models to assess the role of protein interactions and PPI function within the PRLr complex during the pathogenesis of mammary cancer. The studies proposes are highly significant in that the will provide a molecular foundation for our understanding of the structure/function relationships during receptor-Jak2/Src activation as modulated by the PPI activity CypA and translate these discoveries through cellular to mouse models of breast cancer.