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Production of insulin‐like growth factor II (IGF‐II) and different forms of IGF‐binding proteins by HT‐29 human colon carcinoma cell line
Author(s) -
Culouscou JeanMichel,
RemacleBonnet Maryse,
Garrouste Françoise,
Fantini Jacques,
Marvaldi Jacques,
Pommier Gilbert
Publication year - 1990
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.1041430302
Subject(s) - radioimmunoassay , growth factor , insulin like growth factor 2 , cell culture , somatomedin , insulin like growth factor , insulin like growth factor binding protein , biology , receptor , molecular mass , size exclusion chromatography , biochemistry , chemistry , microbiology and biotechnology , genetics , enzyme
The serum‐free medium conditioned by the human colon cancer cell line HT‐29 contains insulin‐like growth factors (IGF) that are entirely complexed to binding proteins (IGF‐BP). Gel filtration in acid conditions of the cell‐conditioned medium permits separation of IGF‐BP from two molecular forms of IGF of 15,000 and 7,500 M r . As determined by ligand blotting, IGF‐BP are heterogeneous and constituted of three molecular forms of 31,000, 28,000, and 26,000 M r . Using IGF‐I and IGF‐II radioreceptor assays, IGF‐I radioimmunoassay (RIA), and competitive protein‐binding assay specific for IGF‐II, it is shown that the IGF‐type eluting in 15 K and 7.5 K position from gel filtration is restricted to IGF‐II. Its concentration is ∼6 ng/10 6 HT‐29 cells with 60% present as a high‐molecular‐weight form of IGF‐II. This large 15 K IGF molecule is devoided of any IGF‐binding activity and might represent incomplete processing of pro‐IGF‐II peptide. By contrast, the level of IGF‐I detected by RIA is barely measurable and considered negligible (0.57 pg/10 6 HT‐29 cells). Although these IGF‐II‐like peptides exhibit a growth‐promoting activity on FR3T3 fibroblasts, they cannot stimulate, as recombinant IGF‐I or IGF‐II, 3 H‐thymidine incorporation into DNA of HT‐29 cells, whatever the experimental conditions used. Finally, we have shown that IGF binding is restricted predominantly to the basolateral domain of the cell membrane by using HT‐29‐D4 clonal cells, derived from the parental HT‐29 cell line, maintained in a differentiated state by culture in a medium in which glucose is replaced by galactose.