Premium
Properties and fate of plasma fibronectin bound to the tissue culture substratum
Author(s) -
Haas Robert,
Culp Lloyd A.
Publication year - 1982
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041130217
Subject(s) - fibronectin , incubation , chemistry , polyacrylamide gel electrophoresis , cell culture , tissue culture , blood proteins , in vivo , in vitro , gel electrophoresis , biochemistry , cell , chromatography , biology , enzyme , microbiology and biotechnology , genetics
Abstract Plasma fibronectin (pFn) is a serum protein which, when adsorbed to a glass or plastic substratum, mediates the adhesion of fibroblasts in culture. We have studied some of the details of its adsorption and subsequent fate. By using 125 l‐labeled pFn, we show that a substratum incubated with pFn adsorbs approximately 0.4 μg/cm 2 pFn (a monomolecular layer), and one incubated with medium containing serum adsorbs approximately 7 ng/cm 2 pFn (a 12‐fold enrichment relative to a random selection of the soluble proteins). SDS‐polyacrylamide gel electrophoresis (SDS‐PAGE) suggests the bound serum proteins (eluted with SDS) are primarily BSA and β‐globulins. The bound pFn adheres so tightly, though, that most resists elution, as assayed (1) with pFn radioiodinated before binding, (2) with pFn radioiodinated after binding, or (3) by the cell spreading activity of the bound pFn retained after SDS treatment. Under culture conditions, there is a continuous “turnover” of substratum‐bound pFn: soluble pFn can bind to a serum‐coated substratum, while bound pFn is gradually removed by incubation with serum proteins. The presence of fibroblasts increases the rate of this removal several‐fold. By SDS‐PAGE the material removed (as well as that eluted from the substratum with SDS after cell detachment) is intact pFn or large (possibly proteolytically generated) fragments. Thus, pFn binds preferentially to the tissue culture substratum, but can be removed subsequently by the combined action of cells and other serum proteins.