Trafficking of the EGF receptor to the nucleus: Mechanisms and Effects

描述（由申请人提供）：肝脏显示出独特的生长和再生能力。例如，在三分之二的肝脏被切除后的几天到几周内发生完全的肝再生。慢性肝细胞损伤可导致肝再生调节受损，从而导致肝细胞癌，这是世界上最常见的恶性肿瘤之一。母公司授权的假设是HGF通过c-met通过诱导肝细胞核内InsP 3介导的Ca 2+信号来调节肝脏中的生长。该FIRCA应用将研究这是否是受体酪氨酸激酶（RTK）在一系列组织中的更一般的作用机制。具体而言，该FIRCA申请的假设是表皮生长因子受体（EGFR）与c-met一样，通过在细胞核内诱导InsP 3介导的Ca 2+信号来调节细胞生长，并且EGFR的这种作用介导常见恶性肿瘤中的细胞增殖。这一假设将通过以下具体目标进行检验：1。将确定EGFR是否以及如何到达常见恶性肿瘤的细胞核。我们将测试是否有一个亚群的EGFR在小窝交通的细胞核。将通过细胞分级分离研究监测受体的细胞内移动。在肝细胞中鉴定的途径将在源自乳腺癌、肺癌、前列腺癌和结肠癌的细胞中进行测试。2. EGF是否以及如何增加细胞核中的Ca 2+将被确定。靶向InsP 3缓冲液将用于确定EGF是否像HGF一样特异性诱导细胞核内的InsP 3形成。RNA干扰技术将用于比较EGF和HGF激活的PLC亚型，并确定这些PLC亚型是否在细胞类型之间存在差异。3.将确定细胞核Ca 2+信号在EGF诱导的细胞生长中的作用。我们将确定EGF诱导的细胞增殖是否通过阻断（a）EGFR向细胞核的移动，（B）EGF诱导的细胞核中Ca 2+信号的形成，或（c）细胞核内Ca 2+依赖性蛋白如CaMK II的活化而被破坏。这些研究将揭示生长因子及其相应的受体酪氨酸激酶如何控制完整细胞中的核Ca 2+，并确定其在调节肿瘤生长中可能发挥的独特作用。本研究将主要在巴西UFMG与Dawidson Gomes合作进行，作为NIH P01 DK 57751项目1的扩展。 公共卫生相关性：肝脏和其他器官内细胞的生长受生长因子及其受体（称为受体酪氨酸激酶）调节。这些研究将揭示生长因子及其相应的受体酪氨酸激酶如何控制细胞核内的信号传导，并确定该信号传导途径在各种肿瘤中发生的异常生长中可能发挥的独特作用。

DESCRIPTION (provided by applicant): The liver displays a unique ability to grow and regenerate. For example, complete hepatic regeneration occurs within days to weeks after two-thirds of the liver has been resected. Chronic hepatocellular damage can lead to impaired regulation of liver regeneration, which results in hepatocellular carcinoma, one of the most common malignancies in the world. The hypothesis of the parent grant is that HGF, via c-met, regulates growth in the liver by inducing InsP3-mediated Ca2+ signals within the nucleus of hepatocytes. This FIRCA application would investigate whether this is a more general mechanism of action of receptor tyrosine kinases (RTKs) across a range of tissues. Specifically, the hypothesis of this FIRCA application is that the Epidermal Growth Factor receptor (EGFR), like c-met, regulates cell growth by inducing InsP3-mediated Ca2+ signals within the nucleus, and that this action of EGFR mediates cell proliferation in common malignancies. This hypothesis will be tested through the following specific aims: 1. whether and how the EGFR reaches the nucleus in common malignancies will be determined. We will test whether a sub-population of EGFRs in caveolae traffic to the nucleus. Intracellular movement of the receptor will be monitored by as well as by cell fractionation studies. Pathways identified in liver cells will be tested in cells derived from breast, lung, prostate, and colon cancers. 2. Whether and how EGF increases Ca2+ in the nucleus will be determined. Targeted InsP3 buffers will be used to determine whether EGF, like HGF, specifically induces InsP3 formation within the nucleus. RNA interference techniques will be used to compare PLC isoforms activated by EGF and HGF, and to determine whether these PLC isoforms vary among cell types. 3. The role of nuclear Ca2+ signals in EGF-induced cell growth will be determined. We will determine whether EGF-induced cell proliferation is disrupted by blocking either (a) movement of EGFR to the nucleus, (b) EGF-induced formation Ca2+ signals in the nucleus, or (c) activation of Ca2+-dependent proteins within the nucleus, such as CaMKII. These studies will reveal how growth factors and their corresponding receptor tyrosine kinases control nuclear Ca2+ in intact cells, and identify the distinct role this may play in regulating tumor growth. This research will be performed primarily at UFMG in Brazil in collaboration with Dawidson Gomes as an extension of Project 1 of NIH P01 DK57751. PUBLIC HEALTH RELEVANCE: Growth of cells within the liver and other organs is regulated by growth factors and their receptors, known as receptor tyrosine kinases. These studies will reveal how growth factors and their corresponding receptor tyrosine kinases control signaling within the cell nucleus, and identify the distinct role this signaling pathway may play in the abnormal growth that occurs in a wide variety of tumors.