Role of JAK2-PAK1 interaction in prolactin-dependent signaling

JAK2-PAK1 相互作用在催乳素依赖性信号传导中的作用

• 批准号: 8325709
• 负责人: MARIA DIAKONOVA
• 金额: $ 29.89万
• 依托单位: UNIVERSITY OF TOLEDO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8325709
• 关键词: Actins; Adipocytes; Affect; Alveolar; Anterior Pituitary Gland; Apoptosis; Astrocytes; Basic Science; Binding; Biological; Biological Process; Breast; Breast Cancer Cell; CCND1 gene; Cancer Center; Cancer Research Project; Cancer cell line; Cell Survival; Cells; Cellular Morphology; Cessation of life; Clinical Sciences; Columbidae; Consult; Cyclin D1; Cytokine Receptors; Cytoskeleton; DNA Sequence Rearrangement; Data; Development; Dissection; Endocrine; Epithelial Cells; Etiology; Event; Functional disorder; Gene Expression Regulation; Genes; Genetic Transcription; Gland; Goals; Grant; Growth Factor; Hormones; Human; JAK2 gene; Lacrimal gland structure; Lactation; Letters; Life Expectancy; Ligand Binding; Link; Liver; Location; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of lung; Mammary Neoplasms; Mammary gland; Mediating; Membrane; Michigan; Milk; Molecular; Molecular Target; Multiprotein Complexes; Names; Normal Cell; Oryctolagus cuniculus; Ovary; Pancreas; Pathway interactions; Peptide Mapping; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Physician Executives; Pituitary Gland; Play; Production; Prolactin; Prolactin Receptor; Prostate; Protein Tyrosine Kinase; Protein-Serine-Threonine Kinases; Proteins; Receptor Activation; Receptor Signaling; Recruitment Activity; Regulation; Research; Risk; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Signaling Protein; Specimen; Structure of beta Cell of islet; Submandibular gland; T-Lymphocyte; Translational Research; Tumorigenicity; Tyrosine; Tyrosine Phosphorylation; United States; Universities; Woman; Work; adapter protein; autocrine; base; biological adaptation to stress; cancer cell; cancer diagnosis; cancer type; carcinogenesis; cell motility; cell type; design; human disease; in vivo; interest; malignant breast neoplasm; mammary epithelium; mammary gland development; medical schools; neoplastic cell; novel; novel therapeutic intervention; oncology; overexpression; protein protein interaction; public health relevance; receptor; receptor binding; response; tumor; tumor progression; two-dimensional

DESCRIPTION (provided by applicant): In normal mammary development, the hormone prolactin (PRL) is critical for alveolar proliferation and differentiation. Increasing evidence supports the involvement of PRL in breast cancer, the leading type of cancer in women and the second leading cause (after lung cancer) of cancer death among women. In 2008, 40,480 women are expected to die from breast cancer in the U.S. The prolactin receptor (PRLR) is detected in 80% of human breast cancers and is overexpressed in breast cancer cells. Normal and tumor mammary epithelial cells synthesize PRL and PRLR, thus the PRL could behave as an autocrine growth factor for human breast cancer cells. These results suggest the need for a more complete understanding of PRLR signaling in breast cancer. Tyrosine (Tyr) kinase JAK2 was identified as a PRLR-bound signaling molecule. Identification of the proteins recruited to the PRLR-JAK2 and dissection of the signaling pathways that are subsequently activated will ultimately provide a basis for understanding PRL action. Preliminary data demonstrate that the serine-threonine kinase PAK1 associates with and is Tyr phosphorylated by JAK2. Two-dimensional peptide mapping identified three Tyr(s) of PAK1 which are phosphorylated by JAK2. Tyr phosphorylation of PAK1 by JAK2 was also shown to increase cell motility. In this grant we propose to examine the hypothesis prolactin-dependent JAK2 phosphorylation of PAK1 regulates PAK1 activity. Activated PAK1 regulation of target proteins may depend on phosphorylation events or/and protein-protein interactions, leading to the formation of a multiprotein complex that modulates the actin cytoskeleton, increases cell motility and invasiveness, mediates cyclin D1 gene transcription and affects tumorigenicity of human breast cancer cells. Aim1 will determine the role of JAK2-phosphorylated PAK1 in regulating PRL-dependent actin cytoskeleton rearrangement, cell motility and invasiveness. Aim2 will determine the role of PAK1 in regulating PRL-activated cyclin D1 gene transcription. Finally, Aim3 will determine whether JAK2 phosphorylation of PAK1 affects the tumorigenicity of human breast cancer cells in vivo. Because both PAK1 and PRL have been implicated in breast cancer, the proposed studies may ultimately fill out the existing gap between upstream PRL-PRLR-JAK2 events and downstream PAK1-dependent functions in our understanding of the mechanism of human breast cancer. Tyr phosphorylation of PAK1 by JAK2 is likely to represent a novel molecular target in the search for the etiology and treatment of human breast cancer. PUBLIC HEALTH RELEVANCE: Prolactin (PRL) was discovered in 1928 as a pituitary factor able to stimulate mammary gland development and lactation in rabbits, as well as the production of crop milk in pigeons. A few years later, the name prolactin was given, based on its ability to stimulate milk production. More than 300 separate biological activities have been attributed to PRL. These biological functions are mediated by specific membrane receptors. These receptors are non tyrosine kinases, they transduce the signal via associated kinases that are recruited by the receptor and activated upon ligand binding. One of these kinases is JAK2 tyrosine kinase. We have recently shown that another protein - PAK1 - is a novel substrate of JAK2. We showed that JAK2 binds to PAK1 and makes PAK1 more active. Activated PAK1 contributes to better cell survival and cell migration. However, how these two proteins work together and the precise mechanism of their action is unknown. PRL is also involved in breast cancer. Breast cancer is the most commonly diagnosed cancer in women - nearly 1 in 3 (30%) of all cancers in women occur in the breast. Based on the current life expectancy for women in the United States, 1 out of 9 women will develop breast cancer in her lifetime - a risk that was 1 out of 14 in 1960. Much is still unknown regarding the molecular biological mechanism by which a normal cell becomes a cancer cell. Our long term goal is to understand the molecular mechanisms of PRL action and disregulation of which leads to human diseases including breast cancer. Toward this aim, we have started to analyze the relationship between JAK2 and PAK1. In the current proposal we will study how JAK2 and PAK1 increase cell migration and cell survival, and which genes are regulated by these two proteins. Understanding how these proteins work together will help to design new therapeutic approaches and possibly drugs for treatment of different human diseases and breast cancer.

描述（由申请人提供）：在正常乳房发育中，催乳素激素（PRL）对于肺泡增殖和分化至关重要。越来越多的证据支持 PRL 与乳腺癌有关，乳腺癌是女性的主要癌症类型，也是女性癌症死亡的第二大原因（仅次于肺癌）。 2008年，美国预计将有40,480名女性死于乳腺癌。催乳素受体(PRLR)在80%的人类乳腺癌中被检测到，并且在乳腺癌细胞中过度表达。正常和肿瘤乳腺上皮细胞合成PRL和PRLR，因此PRL可以作为人类乳腺癌细胞的自分泌生长因子。这些结果表明需要更全面地了解乳腺癌中的 PRLR 信号传导。酪氨酸 (Tyr) 激酶 JAK2 被鉴定为 PRLR 结合信号分子。鉴定招募到 PRLR-JAK2 的蛋白质并剖析随后激活的信号通路将最终为理解 PRL 作用提供基础。初步数据表明，丝氨酸-苏氨酸激酶 PAK1 与 JAK2 结合并被 JAK2 磷酸化。二维肽图谱鉴定出 PAK1 的三个酪氨酸被 JAK2 磷酸化。 JAK2 对 PAK1 的 Tyr 磷酸化也被证明可以增加细胞运动性。在这项资助中，我们建议检验 PAK1 催乳素依赖性 JAK2 磷酸化调节 PAK1 活性的假设。激活的PAK1对靶蛋白的调节可能依赖于磷酸化事件或/和蛋白质-蛋白质相互作用，导致多蛋白复合物的形成，调节肌动蛋白细胞骨架，增加细胞运动性和侵袭性，介导细胞周期蛋白D1基因转录并影响人乳腺癌细胞的致瘤性。 Aim1 将确定 JAK2 磷酸化的 PAK1 在调节 PRL 依赖性肌动蛋白细胞骨架重排、细胞运动和侵袭性中的作用。 Aim2 将确定 PAK1 在调节 PRL 激活的细胞周期蛋白 D1 基因转录中的作用。最后，Aim3将确定JAK2对PAK1的磷酸化是否影响人乳腺癌细胞体内的致瘤性。由于 PAK1 和 PRL 都与乳腺癌有关，因此拟议的研究可能最终填补上游 PRL-PRLR-JAK2 事件与下游 PAK1 依赖性功能之间现有的空白，以帮助我们了解人类乳腺癌的机制。 JAK2 对 PAK1 的 Tyr 磷酸化可能代表寻找人类乳腺癌病因和治疗的新分子靶点。 公共健康相关性：催乳素 (PRL) 于 1928 年被发现，是一种垂体因子，能够刺激兔子的乳腺发育和泌乳，以及鸽子的嗉囊奶的产生。几年后，由于其刺激产奶的能力，催乳素被命名为催乳素。 PRL 具有 300 多种不同的生物活性。这些生物学功能是由特定的膜受体介导的。这些受体是非酪氨酸激酶，它们通过相关激酶转导信号，这些激酶由受体募集并在配体结合时激活。这些激酶之一是 JAK2 酪氨酸激酶。我们最近证明另一种蛋白质 - PAK1 - 是 JAK2 的新型底物。我们发现 JAK2 与 PAK1 结合并使 PAK1 更加活跃。激活的 PAK1 有助于更好的细胞存活和细胞迁移。然而，这两种蛋白质如何协同工作以及它们作用的确切机制尚不清楚。 PRL 也与乳腺癌有关。乳腺癌是女性最常诊断的癌症 - 近三分之一 (30%) 的女性癌症发生在乳腺癌。根据目前美国女性的预期寿命，每 9 名女性中就有 1 人会在一生中患上乳腺癌——这一风险在 1960 年为 14 人中就有 1 人。关于正常细胞变成癌细胞的分子生物学机制，目前仍有很多未知数。我们的长期目标是了解 PRL 作用及其失调导致包括乳腺癌在内的人类疾病的分子机制。为了这个目标，我们开始分析JAK2和PAK1之间的关系。在当前的提案中，我们将研究 JAK2 和 PAK1 如何增加细胞迁移和细胞存活，以及哪些基因受这两种蛋白调节。了解这些蛋白质如何协同工作将有助于设计新的治疗方法和可能的药物来治疗不同的人类疾病和乳腺癌。