The ras proto-oncogenes function in normal cell proliferation, while their oncogenic counterparts cause transformation of mammalian cells in culture, and are among the most commonly found oncogenes in human tumors. The effector molecule for the ras oncoprotein and the mechanism by which ras causes cell transformation are not known, however, the similarity of the ras protein to other signal transduction molecules suggests they may have common targets. One group of targets for known signal transducers, such as GTP-binding proteins, are plasma membrane ion channels. Preliminary data suggest that ras also modulates the function of a calcium-activated potassium channel in fibroblast cell lines, and that modulation of channel activity affects the proliferative state of the cell. The long-term objectives of this project are to understand the effect of ras at a well-defined target molecule, the calcium-activated potassium channel, and to use this information to probe the role of channel-oncogene interaction in cell transformation. The effect of ras proteins on channel physiology in normal and ras-transformed cells will be investigated using single channel and whole-cell patch clamp recording techniques, in conjunction with intracellular and cell-free membrane patch application of purified ras and specific ras inhibitor proteins. Characterization of the channel will be completed so that the modulatory influence of ras and the effect of the channel on cell proliferation can be better understood. In addition, candidate mechanisms to describe the interaction of ras and the potassium channel will be considered, including direct binding of ras to the channel protein, or modulation at a distance via diffusable second messenger or intervening enzymes. The results of this study will provide a molecular physiological basis for understanding the oncogenic action of ras.
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