In contrast to cells from normal tissues, tumor cells exhibit anchorage-independent growth. By specifically identifying the mechanisms which regulate anchorage-dependent cellular proliferation, it may be possible to gain an understanding of the basis of anchorage-independent tumor cell proliferation. Anchorage- dependent survival and proliferation of normal, diploid cells are mediated in part by the integrin family of adhesion receptors, which include the fibronectin receptor, integrin alpha5beta1. However, several studies have demonstrated that the proliferation and tumorigenicity of transformed cells can also be regulated by the integrin. Recently, we have shown that the unoccupied integrin alpha5beta1 negatively regulates tumor cell proliferation in a dominant fashion in vitro and in vivo. This growth inhibition is accompanied by transcriptional inhibition of immediate early genes and transcriptional induction of a growth arresting gene, in a manner that is reversible by ligation of alpha5beta1 by cellular attachment to fibronectin. Expression of the unoccupied integrin in these tumor cells leads to changes in signals transduced in these cells, which include inactivation of a novel MAP kinase. Cellular attachment to fibronectin then activates the well known form of MAP kinase, ERK 2. These results suggest that integrins can impact signal transduction events and cell behavior when they are unoccupied as well as occupied. In this proposal, the mechanisms bv which the integrin alpha5beta1 regulates anchorage-dependent cellular proliferation. signal transduction. and transcription will be examined in alpha 5 expressing tumor cells. First, the roles of cell attachment to fibronectin and receptor crosslinking in the regulation of cellular proliferation will be examined in alpha5 transfected tumor cells to determine the critical features of integrin ligation that are sufficient to reverse this growth inhibition. Second. the roles of the alpha5 subunit cytoplasmic and extracellular domains in growth, signal transduction, and transcriptional regulation will be established using tumor cells transfected with alpha5 subunits containing deletions and mutations as well as domains from other integrins. Third. the role of the MAP kinases in the regulation of tumor cell proliferation by the integrin alpha5beta1 will be examined in transfected tumor cells by expression of dominant active and inactive upstream regulators of MAP kinases. In addition, this regulation of proliferation and MAP kinase activation by integrin alpha5beta1 will be examined in additional tumor cell types. By obtaining an understanding of the mechanisms by which this and other integrins can regulate cellular growth, it may ultimately be possible to develop strategies to control the growth of tumors.
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