Morphology of Ras-Transformed Cells Becomes Apparently Normal Again with Tyrosine Kinase Inhibitors without a Decrease in the Ras-GTP Complex1

Abstract

Radicicol, an inhibitor of protein-tyrosine kinase, was found to cause morphological reversion of v-Ha-ras-transformed NIH3T3 fibroblasts and T24 human urinary bladder carcinoma cells that contain an activated ras mutation. The network of actin stress fibers was restored during the treatment with radicicol. A similar morphological change was observed with another protein-tyrosine kinase inhibitor, herbimycin A. Radicicol did not cause any changes in the proportion of the active GTP binding form of p21ras or its subcellular localization. These results rule out the possibility that the morphological reversion by radicicol is due to direct or indirect inhibition of the p21ras function. Cycloheximide and actinomycin D inhibited the morphological change by radicicol, suggesting that the induced transcription of a gene(s) followed by de novo protein synthesis is required for suppression of the transformed phenotype in ras-transformed cells by tyrosine kinase inhibitors.

Targeting of the protein chaperone, HSP90, by the transformation suppressing agent, radicicol

“…With this goal in mind, a biotinylated derivative of radicicol was synthesized. The structure of radicicol (shown in Figure 1b) contains an epoxide residue that is critical for its antifungal properties (Ayer et al, 1980) and may play a role in its biological activity as a tyrosine kinase inhibitor (Kwon et al, 1992a). In addition, radicicol, like many other tyrosine kinase inhibitors, possesses phenolic ring structures that may be necessary for its function.…”

“…These observations are especially germane in light of recent ®ndings that radicicol treatment of K-Ras transformed cells leads to the selective degradation of Raf-1 and a concomitant suppression of the transformed phenotype (Soga et al, 1998). In addition, previous studies have shown that radicicol treatment attenuates the MAP kinase pathway in vivo (Zhao et al, 1995), but has little or no e ect on the in vitro activity of MAP kinase (Kwon et al, 1995). In its inactive state Raf exists as a cytosolic multi-protein complex, associated with proteins such as HSP90, Cdc37/p50 and 14-3-3.…”

“…Previous studies have documented that radicicol has the capacity to suppress transformation by a variety of oncogenes such as Src, Ras and Mos (Kwon et al, 1992b(Kwon et al, , 1995Zhao et al, 1995). However, the molecular mechanism of action of radicicol is currently unknown.…”

“…Other approaches target the Ras signal transduction pathway as a means to control its activity (reviewed in Marshall, 1995a). An example of an agent that suppresses Rastransformation by the latter approach, is radicicol (Kwon et al, 1995;Soga et al, 1998;Zhao et al, 1995). In addition to Ras-transformed cells, radicicol also suppresses the transformed phenotype of cells expressing activated Src (Kwon et al, 1992b) and Mos oncogenes (Zhao et al, 1995).…”

“…In addition to Ras-transformed cells, radicicol also suppresses the transformed phenotype of cells expressing activated Src (Kwon et al, 1992b) and Mos oncogenes (Zhao et al, 1995). In both Ras-and Mostransformed cells radicicol treatment led to the generalized attenuation of the Ras-Raf-MAP kinase pathway in vivo (Zhao et al, 1995), but had no e ects on the kinase activity of MAP kinase kinase (MAPKK) or MAP kinase in vitro (Kwon et al, 1995). This apparent paradox adds to the conundrum of the mechanism of transformation suppression by radicicol.…”