FIBRONECTIN IN CULTURED RAT KERATINOCYTES - DISTRIBUTION, SYNTHESIS, AND RELATIONSHIP TO CYTOSKELETAL PROTEINS

Abstract

Whether epidermal cells can synthesize fibronectin and whether the distribution of this glycoprotein is related to the adhesion and cytoskeletal organization of these cells was investigated. The production of fibronectin by newborn rat epidermal cells was shown by indirect immunofluorescence staining of cultures grown in the absence of a feeder layer using an antiserum which had been cross-adsorbed with fetal calf serum proteins to remove antibodies which recognized serum fibronectin. The distribution of fibronectin in areas of cell-cell and cell-substratum contact, characteristically in the form of short radial stitches, was examined in more detail using immunoelectron microscopy with colloidal gold as marker. This showed the close proximity of fibronectin to the cell membrane, with the ventral surface and fine cellular processes showing the heaviest labelling, and also revealed evidence of a relationship between external fibronectin and internal structure in epidermal cells. Immunofluorescence showed that tonofilaments (keratin) and microtubules were present as fibrillar arrays but were not related to fibronectin distribution. Vimentin and desmin were absent. Actin was distributed as a circumferential bundle of filaments, with finer strands running radially to the edge. The latter were reminiscent of the radial fibronectin stitches, and a spatial correspondence between fibronectin and actin was confirmed by double-label immunofluorescence which revealed many instances of overlap and colinearity of actin and fibronectin filaments. Evidently, keratinocytes can contribute to the formation of the basement membrane in skin. The localization of fibronectin and its close association with actin also suggests that it is involved in keratinocyte adhesion and is related to the internal organization of these cells.