Gastrointestinal Peptide Signaling through PKC/PKD

通过 PKC/PKD 的胃肠肽信号传导

• 批准号: 8074526
• 负责人: JUAN ENRIQUE ROZENGURT
• 金额: $ 32.07万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-03-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8074526
• 关键词: 70-kDa Ribosomal Protein S6 Kinases; Agonist; Applications Grants; Binding; Biological Process; Bombesin; Cell Differentiation process; Cell Proliferation; Cell Proliferation Regulation; Cell physiology; Cells; Chemicals; Chronic; Development; Disease; Distal; Epithelial Cells; Epithelium; Event; Family; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Gastrin releasing peptide; Gastrointestinal Hormone Receptors; Gastrointestinal tract structure; Goals; Growth; In Vitro; Indium; Inflammatory; Intestines; Investigation; Laboratories; Malignant Neoplasms; Mediating; Mitogen-Activated Protein Kinases; Modeling; Molecular Mechanisms of Action; Pathogenesis; Pathway interactions; Peptide Signal Sequences; Peptides; Phase; Phosphorylation; Phosphotransferases; Play; Process; Protein Isoforms; Protein Kinase C; Proteins; Public Health; Receptor Activation; Regulation; Role; Signal Transduction; Signal Transduction Pathway; Stimulation of Cell Proliferation; Testing; Therapeutic Intervention; Transgenic Mice; Transgenic Organisms; biological systems; cell growth; citrate carrier; experience; gastrointestinal; in vivo; interest; mTOR protein; migration; mouse model; mutant; novel; protein kinase D; response; tumorigenesis

DESCRIPTION (provided by applicant): G protein-coupled receptors (GPCR) agonists, including gastrointestinal (GI) peptides, promote rapid contractile and/or secretory responses in their target cells. The discovery that GI peptides can also induce cell proliferation identified a novel role for these informational molecules. Recent evidence indicates that the mitogenic effects of GPCR agonists are relevant for a number of critical biological processes, including development, compensatory growth and tumorigenesis. The broad, long-term objective of this proposal remains to elucidate the signal transduction pathways that mediate GPCR-induced cell proliferation. Protein kinase C (PKC) occupies a pivotal role in the signal transduction pathways that mediate numerous cellular responses elicited by GI peptides including cell proliferation and modulation of GPCR signaling. However, the events occurring downstream of specific isoforms of PKC remain elusive and our own results demonstrated the existence of PKC-independent pathways leading to mitogenesis. The present challenge is to identify PKC-dependent and PKC-independent downstream targets for GPCRs that transmit signals to the cell interior that participate in the regulation of cell proliferation. Recently, protein kinase D (PKD), cloned in our laboratory, has emerged as a major element in the signal transduction pathways leading to GPCR-induced cell proliferation. Our previous studies demonstrated rapid PKD activation in a broad range of biological systems, including intestinal epithelial cells, via PKC- dependent PKD activation loop phosphorylation on Ser744 and Ser748. In the current proposal, we develop a model that envisages sequential PKD activation involving an early PKC-dependent and a late PKC- independent phase of PKD activation via transphosphorylation and autophosphorylation mechanisms. Additional studies with cells in culture and transgenic mice that express elevated levels of PKD protein in the distal small intestinal and colonic epithelium indicate that PKD plays a major role in mediating proliferation of intestinal epithelial cells, both in vitro and in vivo. Consequently, our central hypothesis is that GPCR agonists activate a PKD-mediated phosphorylation cascade that plays a critical role in the signal transduction of GPCR-induced cellular proliferation. An additional hypothesis is that PKD signaling also operates in the regulation of migration of intestinal epithelial cells. This proposal will examine the following Specific Aims: 1) Characterize early and late phases of PKD activation in response to GPCR activation in intestinal epithelial cells; 2) Characterize the role of PKD in the proliferation and migration of intestinal epithelial cells; 3) Determine the role of PKD in intestinal cell proliferation in vivo using transgenic expression of PKD. Relevance to Public Health: Gastrointestinal (GI) peptides are chemical messengers that play critical roles in the regulation of cell proliferation and are also implicated in the pathogenesis of important diseases, including chronic inflammatory processes and cancer. In the current application, we propose to continue our studies elucidating the regulation and function of protein kinase D (PKD), a molecule that is emerging as a key player in transmitting GI peptide signals in their target cells. These studies have important implications not only for understanding fundamental regulatory mechanisms in the GI tract but also for identifying PKD as a novel target for therapeutic intervention in diseases characterized by either excessive or defective cell proliferation.

描述（由申请人提供）：G蛋白偶联受体（GPCR）激动剂，包括胃肠道（GI）肽，促进其目标细胞中快速收缩和/或分泌反应。 GI肽还可以诱导细胞增殖发现这些信息分子的新作用。最近的证据表明，GPCR激动剂的有丝分裂作用与许多关键的生物学过程有关，包括发育，补偿性生长和肿瘤发生。该提案的广泛，长期目标仍然是阐明介导GPCR诱导细胞增殖的信号转导途径。 蛋白激酶C（PKC）在信号转导途径中占据关键作用，介导GI肽引起的许多细胞反应，包括细胞增殖和GPCR信号的调节。然而，发生的PKC特异性同工型下游发生的事件仍然难以捉摸，我们自己的结果证明了PKC独立的途径的存在，导致有丝分裂发生。目前的挑战是识别PKC依赖性和PKC独立的下游靶标的GPCR，将信号传输到参与细胞增殖调节的细胞内部。 最近，克隆在我们的实验室中的蛋白激酶D（PKD）已成为信号转导途径的主要元素，导致GPCR诱导的细胞增殖。我们先前的研究表明，通过PKC依赖性PKD激活环磷酸化在Ser744和Ser748上，包括肠上皮细胞在内的广泛生物系统中的快速PKD激活。在当前的建议中，我们开发了一个模型，该模型设想了涉及PKC依赖性早期和晚期PKC-独立阶段PKD激活的序列PKD激活的模型，该模型通过磷酸化和自磷酸化机制进行了PKD激活的晚期独立阶段。对培养细胞和转基因小鼠的细胞进行的其他研究表达了远端小肠和结肠上皮中PKD蛋白水平升高的研究表明，PKD在体外和体内介导肠上皮细胞的增殖中起主要作用。因此，我们的中心假设是GPCR激动剂激活了PKD介导的磷酸化级联反应，该级联反应在GPCR诱导的细胞增殖的信号转导中起关键作用。另一个假设是，PKD信号在调节肠上皮细胞的迁移中也起作用。该建议将检查以下特定目的：1）表征PKD激活的早期和晚期，以响应肠道上皮细胞中的GPCR激活； 2）表征PKD在肠上皮细胞增殖和迁移中的作用； 3）使用PKD的转基因表达确定PKD在体内肠细胞增殖中的作用。 与公共卫生有关：胃肠道（GI）肽是化学信使，在调节细胞增殖中起着关键作用，也与重要疾病的发病机理有关，包括慢性炎症过程和癌症。在当前的应用中，我们建议继续我们的研究，以阐明蛋白激酶D（PKD）的调节和功能，蛋白质激酶D（PKD）是一种在其目标细胞中传输GI肽信号的关键参与者的分子。这些研究不仅对理解胃肠道的基本调节机制具有重要意义，而且还将PKD鉴定为以过度或有缺陷的细胞增殖为特征的疾病治疗干预的新靶标。