Embryonal carcinoma (EC) cells are models of early embryonic ectoderm and are tumorigenic in adult hosts. EC cells can also """"""""normalize"""""""" when mixed with normal embryo cells and eventually can differentiate into all the tissues of the resulting chimeras. The role of the EGF receptor in development, differentiation and tumorigenesis is to the studied in EC cells and in embryos. My laboratory recently discovered that EC cells express intracellular, EGF-stimulated, 170 kd kinase activity recognized by our recently developed antibodies to purified mouse EGF receptors. This intriguing finding raised a great number of questions important to mechanisms that control early development. An important aim is to characterize intracellular receptors biochemically and to determine if they function and if they can be regulated in a manner similar to cell surface receptors. The latter appear when EC cells are treated for more than 24 hrs. with retinoic acid and will be useful for a comparison. The mechanism whereby EGF receptors are retained within the cell and their role will be examined after introducing exogenous EGF receptors by transfection with an expression vector that directs the synthesis of EGF receptors. Expression will be driven using the SV40 promoter together with dihyrofolate reductase directed selection and amplification. In addition, the effect of reduction of EGF receptor expression in EC cells using an antisense expression vector will be examined on the growth properties, tumorigenicity and ability to differentiate. Thus, one of our aims os to perturb EGF receptor levels so that observations of the subsequent behavior of these multi-functional cells will help elucidate receptor roles in these various activities. Some of the initial steps in the surface EGF receptors of EC and embryonic cells to activate c-fos and c-myc and how these may lead to mitogenesis and/or differentiation. Finally, we hypothesize a particular role for EGF receptors in the process of trophoblast invasion and implantation in the maternal uterus. This will be studied using trophoblast cells lines and blastocyst outgrowths.
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