ADAMs: Key regulators of EGFR signaling

ADAM：EGFR 信号传导的关键调节因子

• 批准号: 7922909
• 负责人: Carl Peter Blobel
• 金额: $ 29.75万
• 依托单位: HOSPITAL FOR SPECIAL SURGERY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7922909
• 关键词: Address; Affect; Amphiregulin; Animals; Biological Assay; Calmodulin; Cells; Chemicals; Chimera organism; Cleaved cell; DTR gene; Development; Dimerization; Disease; Disintegrins; Doctor of Philosophy; Drug Design; EGF gene; Epidermal Growth Factor Receptor; Epiregulin; Event; G-Protein-Coupled Receptors; Goals; Hand; Knockout Mice; Ligand Binding; Ligands; MAP kinase activator; Malignant Neoplasms; Mammary gland; Membrane; Metalloproteases; Mus; Paracrine Communication; Pathway interactions; Phorbol Esters; Protein Family; Proteins; Proteolysis; Receptor Protein-Tyrosine Kinases; Receptor Signaling; Regulation; Research; Role; Signal Pathway; Signal Transduction; Structure; Testing; Transgenic Mice; Work; base; betacellulin; enzyme substrate; human PHEMX protein; in vivo; inhibitor/antagonist; insight; intercellular communication; overexpression; receptor; response

Proteolysis has emerged as a key posttranslational regulator of ligands of the epidermal growth factor receptor (EGFR), a tyrosine kinase receptor with important roles in development and diseases such as cancer. All EGFR-ligands are made as membrane anchored precursors whose ectodomains frequently require proteolytic release or "shedding" to trigger EGFR-signaling. Metalloproteases of the ADAM (a disintegrin and metalloprotease) protein family have key roles in shedding six EGFR-ligands, and mice lacking ADAM17 resemble animals lacking the EGFR, or animals lacking the ADAM17 substrates TGFa, HB-EGF and amphiregulin. The main goal of the proposed research is to elucidate the mechanism underlying the critical role of ADAMs in shedding and activating ligands of the EGFR. Specifically, we will: 1) Perform a structure/function analysis to understand the substrate selectivity and regulation ofADAMslO and 17. We will generate chimera between ADAMslO and 17 as well as between the ADAM10 substrate EGF and the ADAM17 substrate TGFa to identify which domains of these enzymes and substrates are required for their substrate selectivity. Moreover, we will establish how different activators and inhibitors of intracellular signaling pathways affect the function of ADAMslO and 17. 2) Study the role ofADAM17 in juxtacrine signaling. Signaling via the EGFR is unusual in that it requires two separate ligand binding events before the occupied receptors can dimerize. Uncleaved membrane tethered ligands are predicted to impede receptor dimerization at low ligand concentrations, which might explain why cleavage of these ligands is critical for juxtacrine signaling (cell-cell signaling) under certain conditions. However, clustering or overexpression of ligands might allow receptor dimerization and thus juxtacrine signaling even when they are not cleaved. We will test this hypothesis by assessing how low or high concentrations of uncleavable TGFa, or of TGFa that is clustered by the tetraspanin CD9 or by chemical inducers of dimerization, affect EGFR-signaling. 3) Address the in vivo relevance of the results of aim 2 using conditional knockout mice forADAM17 crossed with transgenic mice expressing different levels of TGFa in the mammary gland. We anticipate that the proposed studies will provide exciting new insights into the upstream regulation of the EGFR pathway by proteolysis of its ligands. Because EGFR-signaling has a crucial role in diseases such as cancer, we hope this work will uncover new targets for the design of drugs that can affect EGFR signaling.

蛋白水解已成为表皮生长因子配体的关键翻译后调节因子 受体（EGFR），一种酪氨酸激酶受体，在发育和疾病中发挥重要作用，例如 癌症。所有 EGFR 配体都是作为膜锚定前体制成的，其胞外域经常 需要蛋白水解释放或“脱落”来触发 EGFR 信号传导。 ADAM 的金属蛋白酶（a 解整合素和金属蛋白酶）蛋白家族在脱落六种 EGFR 配体和小鼠中发挥关键作用 缺乏 ADAM17 类似于缺乏 EGFR 的动物，或缺乏 ADAM17 底物 TGFa 的动物， HB-EGF 和双调蛋白。拟议研究的主要目标是阐明其机制 ADAM 在脱落和激活 EGFR 配体中发挥关键作用。具体来说，我们将： 1) 进行结构/功能分析以了解 ADAMs10 的底物选择性和调节 和 17. 我们将在 ADAMs10 和 17 之间以及 ADAM10 底物之间生成嵌合体 EGF 和 ADAM17 底物 TGFa 来识别这些酶和底物的哪些结构域 其底物选择性所需。此外，我们将确定不同的激活剂和抑制剂如何 细胞内信号传导途径影响 ADAMs10 和 17 的功能。 2)研究ADAM17在近分泌信号传导中的作用。通过 EGFR 发出信号是不寻常的，因为它需要两个 在占据的受体二聚化之前分离配体结合事件。未裂解的膜系留 预计配体在低配体浓度下会阻碍受体二聚化，这可能解释了为什么 在某些条件下，这些配体的裂解对于近分泌信号传导（细胞间信号传导）至关重要。 然而，配体的聚集或过度表达可能会导致受体二聚化，从而导致近分泌 即使它们没有被切割，也会发出信号。我们将通过评估低或高来检验这个假设 不可切割的 TGFa 的浓度，或由四跨膜蛋白 CD9 或化学物质聚集的 TGFa 的浓度 二聚化诱导剂，影响 EGFR 信号传导。 3) 使用 ADAM17 杂交的条件敲除小鼠解决目标 2 结果的体内相关性 转基因小鼠的乳腺中表达不同水平的 TGFa。 我们预计拟议的研究将为上游监管提供令人兴奋的新见解 EGFR 途径通过其配体的蛋白水解作用。因为 EGFR 信号传导在疾病中起着至关重要的作用，例如 癌症，我们希望这项工作能够发现设计能够影响 EGFR 信号传导的药物的新靶点。