LINOLEIC ACID METABOLITES WITH PROTEIN TYROSINE KINASES & PHOSPATASES

本项目的重点是考察(S)通过 前列腺素和亚油酸代谢产物促进EGF有丝分裂 成纤维细胞系中的信号。在我们之前对叙利亚仓鼠的研究中 胚胎(SHE)细胞，我们发现亚油酸产品刺激EGF- 依赖于正常细胞的DNA合成，但不能增强有丝分裂 肿瘤抑制基因缺失变异的SHE细胞的免疫应答 功能。对这些结果的一个可信的解释是， 肿瘤抑制因子(-)细胞株缺乏EGF反应元件 与亚油酸化合物相互作用的信号转导途径。 这些结果表明，亚油酸盐产品可能在控制点起作用。 在肿瘤抑制基因与EGF有丝分裂信号相互作用的地方， 尤其是蛋白质酪氨酸磷酸化途径。 我们最近更详细地研究了信号的表达 表皮生长因子的蛋白质及其酪氨酸磷酸化程度 两种SHE细胞变异体中的信号通路。一个细微的差别 在蛋白质水平上观察到，除了较高水平的 在supB‘变异体中观察到EPS8。含酪氨酸的SH-2 磷酸酶PTP-1-D与suB+AS中磷酸化的EGFR结合 预期的，但不结合的磷酸化的EGFR在suB‘细胞。 纯化的人PTP-1-D与SHE细胞孵育确实导致 两个变异体的EGFR的结合。此数据和其他数据指向一个 在supB‘细胞中存在PTP-1-D突变或缺陷。 此外，在supB+变异体中，PTP-1-D的活性更高 与supB‘细胞相比。我们正在克隆PTP-1-D 和来自SHE细胞的EGFR受体。我们也在威慑如果 13(S)-HpODE改变PTP-1-D的活性或结合 和EGFR一起。

The focus of this project is to examine the mechanism(s) by which prostaglandins and linoleic acid metabolites potentiate the EGF mitogenic signal in fibroblast cell lines. In our previous work with Syrian hamster embryo (SHE) cells, we found that linoleate products stimulate EGF- dependent DNA synthesis in normal cells, but do not enhance the mitogenic response in variant SHE cells that had lost tumor suppressor gene function. One plausible interpretation of these results is that the tumor suppressor (-) cell line lacks the responsive element in the EGF signal transduction pathway which interacts with linoleate compounds. These results suggest that linoleate products may act at control points where tumor suppressor genes interact with EGF-mitogenic signals, in particular, the protein tyrosine phosphorylation pathway. We have recently examined in more detail the expression of signaling proteins and the extent of their tyrosine phosphorylation of the EGF signaling pathway in the two SHE cell variants. One minor differences in protein levels was observed with the exception that higher level of eps8 was observed in the supB' variant. The SH-2 containing tyrosine phosphatase PTP-1-D binds to the phosphorylated EGFR in the supB+ as expected but does not bind to the phosphorylated EGFR in the supB' cells. Incubation of human purified PTP-1-D with SHE cells does result in the bind of the EGFR of both variants. This and other data point to a mutation or defect in the PTP-1-D present in the supB' cells. Furthermore, the activity of the PTP-1-D is greater in the supB+ variant compared to the supB' cell. We are in the process of cloning the PTP-1-D and the EGFR receptor from the SHE cells. We are also deterring if the 13(S)-HpODE alters either the activity or the association of the PTP-1-D with the EGFR.