Diacylglycerol Kinase Delta in Growth and Development

描述(申请人提供)：表皮生长因子受体(EGFR)调节细胞的增殖、迁移、凋亡和分化。许多类型的癌症具有异常高的EGFR活性，这有助于它们的生长，研究表明，抑制EGFR可以缩小肿瘤大小。我们建立了二酰甘油激酶(DGK)Delta基因敲除小鼠，发现DGK Delta通过调节蛋白激酶C酶来调节EGFR的表达和活性。这项提议有三个目的，将帮助我们了解DGKDelta是如何调节EGFR信号的，并将测试干扰DGKDelta功能的潜在抗肿瘤效果。在目标1中，我们将确定DGKDelta如何调节EGFR的表达。我们发现，在DGKDelta缺陷细胞中，EGFR泛素化增强，PKC活性增加。此外，我们发现PKCalpha是适当的EGFR泛素化所必需的。综上所述，这些数据使我们假设DGKDelta通过PKCalpha调节EGFR泛素化。我们将在目标1中测试这种可能性。我们还发现DGKDelta独立于EGFR表达的变化来调节EGFR的活性。在DGKDelta缺陷的细胞和组织中，我们发现EGFR中的苏氨酸残基的磷酸化增强。已知这种磷酸化可以抑制EGFR的活性，我们的数据表明PKCDelta与此有关。这导致我们假设DGKDelta通过抑制EGFR的PKC Delta磷酸化来调节EGFR的活性。在目标2中，我们将测试这种可能性。最后，在目标3中，我们将使用细胞培养和小鼠异种移植模型来测试干扰DGKDelta功能对肿瘤发生的影响。总之，我们的实验将阐明DGKDelta是如何调节EGFR信号的，将定义DGKDelta在肿瘤形成中的作用，并将测量干扰DGKDelta功能的抗肿瘤效果。公共卫生相关性：EGFR是一个重要的癌症靶点。我们发现了一种新的方法来调节它的表达和活性。EGFR的这些特性对它促进癌症至关重要，我们的实验可能会发现新的抗癌疗法。因此，我们的建议与公共卫生具有重要的相关性。

DESCRIPTION (provided by applicant): The epidermal growth factor receptor (EGFR) regulates proliferation, migration, apoptosis, and differentiation of cells. Many types of cancer have abnormally high EGFR activity that contributes to their growth, and studies have shown that inhibiting EGFR can reduce tumor size. We made diacylglycerol kinase (DGK) delta knockout mice and discovered that by regulating protein kinase C enzymes, DGKdelta modulates the expression and activity of EGFR. This proposal has three aims that will help us understand how DGKdelta regulates EGFR signaling and will test the potential anti-tumor effects of disrupting the function of DGKdelta. In Aim 1, we will determine how DGKdelta modulates EGFR expression. We found enhanced EGFR ubiquitination and increased PKC activity in DGKdelta deficient cells. Additionally, we discovered that PKCalpha is required for proper EGFR ubiquitination. Together, these data led us to hypothesize that DGKdelta regulates EGFR ubiquitination through PKCalpha. We will test this possibility in Aim 1. We also discovered that DGKdelta modulates EGFR activity independently of changes in EGFR expression. In DGKdelta deficient cells and tissues, we found enhanced phosphorylation of a threonine residue in EGFR. This phosphorylation is known to inhibit EGFR activity and our data indicate that PKCdelta is responsible. This led us to hypothesize that DGKdelta regulates EGFR activity by inhibiting PKCdelta phosphorylation of EGFR. In Aim 2, we will test this possibility. Finally, in Aim 3, we will use cell culture and mouse xenograft models to test the impact on tumorigenesis of disrupting the function of DGKdelta. Collectively, our experiments will clarify how DGKdelta regulates EGFR signaling, will define the contribution of DGKdelta to tumor formation, and will measure the anti-tumor effects of disrupting the function of DGKdelta. PUBLIC HEALTH RELEVANCE: EGFR is an important cancer target. We have found a novel way to regulate its expression and activity. These properties of EGFR are fundamentally important for it to promote cancer and our experiments might uncover new anti-cancer therapies. Thus, our proposal has important relevance to public health.