MECHANISTIC STUDY OF THE ROLE OF TYROSINE KINASE SIGNAL

Src和Ras的激活与多种肿瘤的发生和转移密切相关。然而，这些oneogenes在药物敏感性和侵袭过程中的作用仍有待澄清。我们研究了Src和Ras在这些过程中的作用，使用HAG-1人上皮细胞系转染v-src和激活H-ras。首先，我们通过将含有显性负性Ras和Rac（DN/Ras和DN/Rac）的腺病毒载体引入这些细胞中来研究Ras和Rac（一种作用于Ras下游的小GT3结合蛋白）的潜在作用。DN/Ras和DN/Rac均能降低src转染细胞的侵袭能力。DN/Rac可完全抑制src转染细胞在裸鼠体内的成瘤能力。这些结果提示Src、Ras、Rac均参与了肿瘤细胞的侵袭过程，Rac可能在这些侵袭和致瘤信号通路的下游起作用。下一篇：ex ...更多信息 研究了活化Src对靶向微管的抗癌药物泰索帝敏感性的影响。转染v-src的HAG/src 3-l细胞对泰索帝的敏感性是亲本细胞的7.0倍。HAG/src 3 -1细胞对泰索帝的敏感性可被除莠霉素A（HA）逆转，表明Src酪氨酸激酶增强了细胞对泰索帝的敏感性。用泰索帝处理HAG/src 3 -1细胞导致Bcl-2磷酸化和随后诱导的凋亡性细胞死亡，而在亲本或c-H-ras转染的HAG-1细胞中既不发生Bcl-2磷酸化也不发生凋亡。Bcl-2蛋白在v-src转染的细胞系中过表达。用HA处理HAG/src 3 -1细胞可降低Bcl-2的表达和磷酸化，并消除紫杉醇诱导的细胞凋亡，这表明Src酪氨酸激酶在紫杉醇诱导的细胞凋亡事件中具有潜在的作用。H-7和PI-3激酶抑制剂wortmannin既不改变紫杉醇敏感性，也不抑制紫杉醇诱导的细胞凋亡。这些数据表明，激活的Src增加泰索帝敏感性的能力将通过Src到下游信号转导途径Bcl-2磷酸化而发生的凋亡事件介导，而不是通过激活的Ras、PI-3激酶或蛋白激酶C。少

Activation of Src and Ras has been demonstrated be closely associated with the pathogenesis and metastatic potential of many human tumors. However, the role of these oneogenes in the drug sensitivity and invasion process remain to be clarified. We examined the role of Src and Ras on these processes, using HAG-1 human epithelial cell lines transfected with v-src and activated H-ras. First we examined the potential role of Ras and Rac, a small GTPase binding protein which acts downstream of Ras, by introducing adenoviral vector containing dominant negative Ras and Rac (DN/Ras and DN/Rac) into these cells. Both DN/Ras and DN/Rac reduced the invasive potential of src-transfected cells. Moreover, DN/Rac suppressed completely the tumorigenic potentials of src-transfected cells in nude mice. These results suggest that Src, Ras, Rac, all appeared to participate in the invasion processes, and that Rac may act downstream of these signaling pathways of invasion and tumorigenicity. Next we have ex … More amined the effect of activated Src on the sensitivity to taxotere, an anticancer drug targeting microtubules. As compared with parental HAG-1 cell line, v-src-transfected HAG/src3-l cells became 7.0-fold sensitive to taxotere. The taxotere sensitivity in HAG/src3-1 cells was reversed by herbimycin A (HA), indicating that Src tyrosine kinase augments sensitivity to taxotere. Treatment of HAG/src3-1 cells with taxotere resulted in phosphorylation of Bel-2 and subsequent induction of apoptotic cell death, whereas neither Bcl-2 phosphorylation nor apoptosis occurred in parental or c-H-ras-transfected HAG-1 cells. The Bcl-2 protein is overexpressed in v-src-transfected cell line. Treatment of HAG/src3-1 cells with HA reduced the expression and phosphorylation of Bcl-2, and abrogated taxotere-induced apoptosis, suggesting a potential role for Src tyrosine kinase in the taxotere-induced apoptotic events. H-7, and wortmannin, PI-3 kinaseinhibitor, neither altered taxoteresensitivity nor inhibited taxotere-induced apoptosis in these cells. These data indicate that the ability of activated Src to increase taxotere sensitivity would be mediated by apoptotic events occurring through Src to downstream signal transduction pathways toward Bcl-2 phosphorylation, but not by activated Ras, PI-3 kinase or protein kinase C. Less