Alterations in Interacting Signaling Pathways

相互作用信号通路的改变

• 批准号: 6949778
• 负责人: STUART H. YUSPA
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6949778
• 关键词: Caenorhabditis elegans; animal tissue; biological signal transduction; carcinogenesis; cell transformation; chemokine; enzyme activity; gene expression; genetic mapping; genetically modified animals; human tissue; inflammation; keratinocyte; laboratory mouse; oncogenes; phosphorylation; polymerase chain reaction; protein isoforms; protein kinase C; skin neoplasms; squamous cell carcinoma; tissue /cell culture

Ras oncogene mediated transformation of mouse keratinocytes to a benign squamous papilloma phenotype is associated with activation of PKC-alpha. Tumors produced by ras transformation of keratinocytes derived from skin targeted PKC-alpha transgenic mice produced in our laboratory grow faster and achieve a larger size, but do not convert to malignancy at a higher rate. Activation of a PKC-alpha transgene by TPA in mouse epidermis induced a marked intraepidermal neutrophilic infiltrate that mimics the inflammatory response in several human cutaneous diseases including psoriasis. Multiple chemokines are upregulated detected by RT-PCR and RPA assays, and supernatants from cultured transgenic keratinocytes contain chemotactic factors for murine neutrophils. In vitro chemotaxis assays indicate that relevant chemokines are downstream of TNF-alpha. Activation of PKC-alpha in transgenic skin causes epidermal apoptosis, and this is prevented in cultured transgenic keratinocytes by inhibiting AP-1 activity. Microarray analysis of TPA treated transgenic keratinocytes has revealed a spectrum of PKC-alpha regulated genes, including differentiation markers, chemokines/cytokines and proteases and these can be selectively altered by specific inhibitors of the AP-1 and NFkappaB pathways. This model continues to reveal unique insights into the regulation of cutaneous inflammation, the regulation of pro-inflammatory and pro-apoptotic pathways in skin and the contribution of inflammation to cutaneous carcinogenesis. Activation of PKC inactivates the AKT kinase in mouse keratinocytes through prevention or reversal of AKT phosphorylation on serine 473. Inhibition of PKC activity enhances IGF-1- mediated protection from UV induced apoptosis, presumably by enhancing AKT activity as detected by AKT phosphorylation. The specific PKC isoforms targeting AKT appear to be delta and/or epsilon. The genomic structures of C. elegans, mouse, rat and human PKCdelta were analyzed after sequencing a genomic fragment of mouse PKC. Overall exon-intron genomic structure was highly conserved among mammals while significantly diverged in C. elegans. A 1.7 kb region of the PKC-delta promoter revealed a hitherto unknown NFkappaB site, and TNF-alpha was shown to upregulate the expression of the gene. Disruption of the nuclear tumor suppressor gene PML function in skin by targeting the PML-RAR-alpha fusion oncogene to the skin of transgenic mice causes spontaneous skin tumor induction. This is associated with cutaneous retinoid deficiency. Tumor development is independent of ras gene mutations. When PML is targeted to the epidermis in transgenic mice, transgenic keratinocytes in vitro senesce more rapidly, adhere more tightly to collagen substrates and have altered proteolytic activity. In vivo, PML transgenic mice undergo spontaneous alopecia, and have delayed skin tumor formation, smaller tumors and a longer latency to carcinoma formation when treated topically with carcinogens.

RAS癌基因介导的小鼠角质形成细胞向良性鳞状乳头状瘤表型的转化与PKC-Alpha的激活有关。由源自皮肤靶向的PKC-α转基因小鼠衍生出的角质形成细胞的RAS转化产生的肿瘤，我们实验室产生的肿瘤生长速度更快并达到更大的尺寸，但不要以较高的速度转化为恶性肿瘤。 TPA在小鼠表皮中激活PKC-Alpha转基因会诱导明显的表皮性嗜中性粒细胞性浸润，以模仿包括牛皮癣在内的几种人皮肤疾病的炎症反应。 RT-PCR和RPA分析检测到多种趋化因子，并且来自培养的转基因角质形成细胞的上清液含有鼠嗜中性粒细胞的趋化因子。体外趋化性测定表明相关的趋化因子在TNF-Alpha的下游。 PKC-α在转基因皮肤中的激活会引起表皮凋亡，并且通过抑制AP-1活性，可以在培养的转基因角质形成细胞中预防这种凋亡。 TPA处理过的转基因角质形成细胞的微阵列分析揭示了PKC-Alpha调控基因的谱，包括分化标记物，趋化因子/细胞因子和蛋白酶，并且可以通过AP-1和NFKAPPAB途径的特定抑制剂选择性地改变这些基因。该模型继续揭示对皮肤炎症调节，皮肤促炎和促凋亡途径的调节以及炎症对皮肤致癌作用的贡献的独特见解。 PKC的激活通过预防或逆转AKT磷酸化的丝氨酸473对小鼠角质形成细胞中的Akt激酶。抑制PKC活性可以增强IGF-1介导的介导的保护，从而通过AKT磷酸化来增强紫外线诱导的凋亡，从而增强AKT AKT活性。靶向Akt的特定PKC同工型似乎是Delta和/或Epsilon。 测序小鼠PKC的基因组片段后，分析了秀丽隐杆线虫，小鼠，大鼠和人PKCDELTA的基因组结构。总体外显子基因组结构在哺乳动物中高度保守，而在秀丽隐杆线虫中则显着分歧。 PKC-DELTA启动子的1.7 kb区域显示迄今未知的NFKAPPAB位点，TNF-Alpha显示出上调基因的表达。通过将PML-RAR-ALPHA融合癌基因靶向转基因小鼠的皮肤，核肿瘤抑制基因PML功能的破坏会引起自发的皮肤肿瘤诱导。这与皮肤类视黄素缺乏有关。肿瘤发育与RAS基因突变无关。当PML靶向转基因小鼠的表皮时，经过的转基因角质形成细胞更快地粘附在胶原蛋白底物上，并改变了蛋白水解活性。在体内，PML转基因小鼠会自发性脱发，并延迟了皮肤肿瘤形成，较小的肿瘤和用癌细胞局部治疗的癌形成的较长潜伏期。