Epidermal growth factor receptors associated to cytoskeletal elements of epidermoid carcinoma (A431) cells.
The structural interaction of the epidermal growth factor (EGF) receptor and the cytoskeleton of A431 cells has been studied using a monoclonal anti-EGF receptor antibody. This has been done with immunogold labeling using a variety of electron microscopical preparation procedures and EGF binding studies. By providing an image of the membrane-associated cytoskeleton, the dry cleavage method reveals a preferential localization of EGF receptors superimposed upon cytoskeletal filaments. The colocalization of gold particles with cytoskeletal filaments is not affected when pre-labeled cells are extracted with the non-ionic detergent Triton X-100, as visualized by dry cleavage. Using surface replication, this treatment results in visualization of the cytoskeleton. In these latter preparations, it is also observed that EGF receptor-coupled gold particles remain associated with cytoskeletal elements. Moreover, Triton extraction performed before immunogold labeling of EGF receptors demonstrates that isolated cytoskeletons contained binding sites for anti-EGF receptor antibodies. Using stereo micrographs of replica's obtained from these isolated cytoskeletons, it is shown that gold-labeled EGF receptors are exclusively present on the cortical membrane-associated region of the cytoskeleton and not on more intracellular-located filaments. Scatchard analysis of EGF binding to cells fixed with glutaraldehyde and treated with Triton X-100 before and after EGF binding indicates that a high affinity EGF binding site is associated with the Triton X-100 insoluble cytoskeleton.
D000911:Antibodies, Monoclonal; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000648:ultrastructure; D002460:Cell Line; D003599:Cytoskeleton / Q000378:metabolism*; D066246:ErbB Receptors; D005404:Fixatives; D006801:Humans; D008854:Microscopy, Electron / Q000379:methods; D011092:Polyethylene Glycols; D011956:Receptors, Cell Surface / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D005404:Fixatives; D011956:Receptors, Cell Surface; D011092:Polyethylene Glycols; D066246:ErbB Receptors