Chimaerins-receptors for diacylgly and phorbal esters

二酰基和佛巴酯的嵌合蛋白受体

• 批准号: 7665264
• 负责人: MARCELO G. KAZANIETZ
• 金额: $ 5.65万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-30 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7665264
• 关键词: 1,2-diacylglycerol; Actins; Adhesions; Affect; Alleles; Animal Model; Binding; Breast; Breast Cancer Cell; C-terminal; Cancer Etiology; Cell Cycle Progression; Cell physiology; Cyclin D1; Cytoskeleton; DAG/PE-Binding Domain; Data; Diglycerides; Disease Progression; Down-Regulation; EGF gene; Epidermal Growth Factor Receptor; Esters; Etiology; Family; Funding; GTPase-Activating Proteins; Generations; Goals; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Heregulin; Human; Hydrolysis; Isoenzymes; Knock-in Mouse; Knock-out; Knockout Mice; Laboratories; Ligands; Link; Lipids; Malignant - descriptor; Malignant Neoplasms; Mammary Neoplasms; Mediating; Methylation; Modeling; Monomeric GTP-Binding Proteins; N-terminal; Names; Neoplasm Metastasis; Neoplasms; Nude Mice; Pathway interactions; Personal Satisfaction; Phenotype; Phorbol Esters; Phospholipase C; Phosphorylation; Phosphorylation Site; Phosphotyrosine; Play; Positioning Attribute; Protein Binding; Protein Family; Protein Kinase C; Protein Overexpression; Protein-Serine-Threonine Kinases; Proteins; Public Health; Receptor Activation; Receptor Protein-Tyrosine Kinases; Regulation; Role; Second Messenger Systems; Signal Transduction; Site; Specimen; Tertiary Protein Structure; Tetradecanoylphorbol Acetate; Therapeutic; Translating; Tumor Promoters; Tumor Suppressor Proteins; Work; Xenograft procedure; cell motility; citrate carrier; concept; malignant breast neoplasm; migration; mouse model; mutant; novel; phorbol ester receptor; promoter; receptor; response; second messenger; src Homology Region 2 Domain; tumor progression; tumorigenesis

DESCRIPTION (provided by applicant): This competing renewal focuses on the characterization of an intracellular receptor for the phorbol ester tumor promoters and the second messenger diacylglycerol (DAG), ¿2-chimaerin, and its relevance in the etiology of breast cancer. With the support of this RO1 we unambiguously established the concept of "divergence in DAG signaling", which implies that DAG and phorbol ester tumor promoters can trigger responses via direct activation of effectors that do not belong to the protein kinase C (PKC) family, considered until not so long ago the only intracellular receptors for DAG and phorbol esters. ¿2-chimaerin possesses 3 well-defined regions: a N-terminal SH2 domain of unknown function (present only in a2- and ¿2-chimaerins), a C1 domain (responsible for DAG and phorbol ester binding), and a C-terminal GTPase Activating Protein (GAP) domain that has the ability to accelerate the hydrolysis of GTP from the small G-protein Rac, leading to Rac inactivation. We established that ¿2-chimaerin is a direct effector of the EGF receptor via the DAG pathway, and that activation of ¿2-chimaerin serves as a "brake" that limits the activation of Rac in response to tyrosine kinase receptor stimulation. Moreover, we determined that in breast cancer cells ¿2-chimaerin negatively modulates Rac1-mediated responses, including actin cytoskeleton reorganization, migration, invasion, proliferation, cell cycle progression, and cyclin D1 induction in response to ErbB receptor ligands, including EGF and heregulin. Remarkably, preliminary studies revealed that ¿2-chimaerin is down-regulated in human breast cancer cells and specimens, suggesting a potential tumor suppressor role for this DAG-regulated Rac-GAP. As an extension of our previous work we now propose 4 aims. In Aim 1 we will determine the relevance of ¿-chimaerin in breast cancer progression using xenografts and also 1ith knock-out and knock-in mice for a hyperactive ¿2-chimaerin mutant (I130A allele). In Aim 2 we will analyze the expression of ¿2-chimaerin in a panel of human mammary tumors and their matched normal counterparts in order to validate our preliminary evidence that ¿2-chimaerin is down-regulated in breast cancer. We will also focus on promoter methylation as the potential cause for this down-regulation. In Aim 3 we will assess the regulation of ¿2-chimaerin activity and function by phosphorylation, as data revealed a PKC phosphorylation potentially modulates the Rac-GAP activity of ¿2-chimaerin. Lastly, in Aim 4 we will identify and validate potential proteins that interact with ¿2-chimaerin via the SH2 domain and determine the functional consequences of such interactons. The identification of a DAG/phorbol ester receptor unrelated to the PKC family that negatively regulates responses associated with the progression of breast cancer, and probably other neoplasias, not only has fundamental connotations in cancer signaling, but it may also reveal novel aspects of the etiology of cancer and have important therapeutic implications. PUBLIC HEALTH RELEVANCE: In this proposal we will characterize a receptor for the phorbol ester tumor promoters and the lipid second messenger diacylglycerol (DAG) named ¿2-chimaerin. We found that this protein negatively regulates important cellular functions involved in tumorigenesis and metastasis, and that it is down-regulated in breast cancer, suggesting a potential relationship with breast cancer etiology. This proposal will investigate the relevance of ¿2-chimaerin in breast cancer progression as well as key regulatory mechanisms that control its activation and function.

描述（通过应用程序提供）：这种竞争的更新重点是对凤凰酯肿瘤启动子的细胞内受体的表征和第二胺二酰基甘油（DAG），€2-Chimaerin及其在乳腺癌病因中的相关性。在此RO1的支持下，我们明确地建立了“ DAG信号发散”的概念，这意味着DAG和Phorbol Ester肿瘤启动子可以通过直接激活不属于蛋白激酶C（PKC）家族的效应来触发响应，直到不久前，它是唯一的细胞内受感受器的DAG和Phorbol esters esters的响应。 2-奇马素具有3个定义明确的区域：一个未知功能的N端SH2结构域（仅存在于A2-和2-吉马素中），一个C1结构域（负责DAG和Phorbol酯结合），以及C-末端的GTPase GTPase激活蛋白（GAP）较小的GTP gtp gtp gtp gtp，失活。我们确定 - 2-奇马素是通过DAG途径的EGF受体的直接效应子，并且2-奇马素的激活是一种“制动”，它限制了RAC对酪氨酸激酶受体刺激的响应。此外，我们确定在乳腺癌细胞中»2-奇马素对Rac1介导的反应进行负调节，包括肌动蛋白细胞骨架重组，迁移，侵袭，侵袭，增殖，细胞周期进展和细胞周期蛋白D1诱导，以响应ERBB受体配体，包括EGF和这里的EGF。值得注意的是，初步研究表明，在人类乳腺癌细胞和标本中，2-奇马素被下调，这表明该DAG调节的RAC-GAP具有潜在的肿瘤抑制作用。作为我们以前的工作的扩展，我们现在提出了4个目标。在AIM 1中，我们将使用Xenographictics和1Ith敲除和敲除小鼠在乳腺癌进展中的相关性，用于2奇马蛋白突变体（I130a等位基因）。在AIM 2中，我们将分析在人类乳腺肿瘤和它们匹配的正常对应物中的2-奇马素的表达，以验证我们的初步证据表明€2-奇马素在乳腺癌中被下调。我们还将专注于启动子甲基化作为这种下调的潜在原因。在AIM 3中，我们将评估 „ 2-奇马素的活性和磷酸化的功能，因为数据表明PKC磷酸化可能会调节2-奇马素的RAC-GAP活性。最后，在AIM 4中，我们将通过SH2结构域识别和验证与2-奇马素相互作用的潜在蛋白质，并确定此类相互作用子的功能后果。与PKC家族无关的DAG/Phorbol酯受体的鉴定，对与乳腺癌进展以及可能其他肿瘤相关的反应产生负面调节，不仅具有癌症信号传导的基本含义，而且还可能揭示了癌症病因的新方面，并且具有重要的治疗疗法。公共卫生相关性：在此提案中，我们将描述佛波尔酯肿瘤启动子的受体和名为2-Chimaerin的脂质二胺二酰基甘油（DAG）。我们发现该蛋白质对肿瘤发生和转移的重要细胞功能负面调节，并且在乳腺癌中被下调，这表明与乳腺癌病因有潜在的关系。该建议将研究„ 2-奇马素在乳腺癌进展中的相关性以及控制其激活和功能的关键调节机制。