FIBRONECTIN REGULATION OF CARCINOMA APOPTOSIS

说明：纤维连接蛋白(FN)及其受体是肿瘤细胞存活的重要调节成分。我们已经确定FN的羧基末端的肝素结合区和选择性剪接的V区在这一过程中是重要的，因为含有突变的肝素结合区和FN的V区的FN微小蛋白(V+H-)诱导鳞状细胞癌(SCC)细胞的凋亡。相反，对应的野生型蛋白(V+H+)或其他不包含V区的FN微型蛋白促进这些细胞的存活。此外，在SCC细胞中，V+H介导的细胞凋亡率比正常的原代角质形成细胞要慢。这些数据表明，致瘤性使SCC细胞能够延迟其对改变的基质的反应而开始凋亡，FN的V区和肝素结合域调节这些细胞的生存。我们的初步数据表明，V+H蛋白诱导SCC细胞延迟凋亡的机制可能涉及硫酸软骨素蛋白多糖和整合素受体，以及P53和c-myc介导的信号。在原代成纤维细胞中，这种凋亡机制是由硫酸软骨素蛋白多糖、α4整合素和一个有趣的需要下调p53和c-myc的新途径介导的。此外，由于P53在这一机制中从细胞核重新定位到细胞膜，并且整合素相关信号分子粘着斑激酶(Pp125FAK)和c-jun氨基末端激酶(JNK)在这一途径中被抑制，这表明P53可能与整合素/FAK产生的信号进行通信。我们推测，SCC细胞通过细胞表面蛋白多糖和整合素受体来抵抗FN基质(V+H-)的改变而产生的抗凋亡作用。这启动了一条信号转导通路，导致FAK、P53和c-myc的下调。其次，由于其在调控SCC细胞侵袭、迁移和凋亡中的重要作用，我们推测FN的V区可能在SCC细胞的发病机制中起重要作用。我们将在以下特定目标中检验这些假说：(1)确定参与延缓V+H-FN蛋白诱导的细胞凋亡的SCC细胞表面受体，并将这些受体与原代角质形成细胞中存在的受体进行比较。(2)检测参与延缓V+H-FN蛋白诱导的细胞凋亡的SCC细胞信号反应。(3)检测FN选择性剪接V区在口腔低度和高度不典型增生及口腔癌组织中的表达。这些研究将有助于解释一些调节肿瘤细胞生物学的细胞-基质相互作用和信号机制，并可能为口腔鳞状细胞癌的发病机制提供见解。

DESCRIPTION: Fibronectin (FN) and its receptors are important regulatory components in tumor cell survival. We have determined that the carboxyl-terminal heparin-binding domain and alternatively spliced V region of FN are important to this process, since a FN miniprotein (V+H-) containing a mutated heparin-binding domain and the V region of FN induces apoptosis of squamous cell carcinoma (SCC) cells. In contrast, the counterpart wildtype protein (V+H+) or other FN miniproteins not containing the V region promote survival of these cells. Furthermore, in SCC cells, the rate of V+H--mediated apoptosis is delayed compared to normal primary keratinocytes. These data suggest that tumorigenicity has enabled the SCC cells to delay their onset of apoptosis in response to an altered matrix, and that the V region and heparin-binding domain of FN regulate survival of these cells. Our preliminary data suggest that the mechanism by which the V+H- protein induces delayed SCC cell apoptosis may involve chondroitin sulfate proteoglycan and integrin receptors, and p53 and c-myc mediated signals. In primary fibroblasts, this apoptotic mechanism is mediated by a chondroitin sulfate proteoglycan, the alpha4 integrin, and by an intriguing new pathway that requires downregulation of p53 and c-myc. In addition, since p53 relocalizes from the nucleus to the cell membrane in this mechanism, and since the integrin-associated signaling molecule focal adhesion kinase (pp125FAK) and c-Jun N-terminal kinase (JNK) are depressed in this pathway, this suggests that p53 may communicate with integrin/FAK generated signals. We posit that SCC cells resist apoptosis in response to an altered FN matrix (V+H-) via cell surface proteoglycan and integrin receptors. This initiates a signal transduction pathway that leads to downregulation of FAK, p53 and c-myc. Secondarily, because of its important role in regulating SCC cell invasion, migration, and apoptosis, we posit that the V region of FN may be important to SCC cell pathogenesis. We will test these hypotheses in the following Specific Aims: (1) Identify the SCC cell-surface receptors involved in delaying apoptosis induced by the V+H- FN protein in these cells and compare these receptors to those present in primary keratinocytes. (2) Examine the SCC cell signaling response involved in delaying apoptosis induced by the V+H- FN protein. (3) Examine the expression of the alternatively spliced V region of FN in low grade and high-grade oral dysplasia and oral cancer. These studies will help explain some of the cell-matrix interactions and signaling mechanisms that regulate tumor cell biology, and may provide insights into the pathogenesis of oral SCC.