Preferential binding of insulin‐like growth factor‐II (IGF‐II) to a putative α2β2 IGF‐II receptor type in C2 myoblasts
- 1 September 1992
- journal article
- Published by Wiley in European Journal of Biochemistry
- Vol. 208 (2) , 273-279
- https://doi.org/10.1111/j.1432-1033.1992.tb17183.x
Abstract
We have studied insulin‐like‐growth‐factor (IGF) binding in two subclones of the C2 myogenic cell line. In the permissive parental subclone, myoblasts differentiate spontaneously into myotubes in medium supplemented with fetal calf serum. Unlike permissive myoblasts, inducible myoblasts require high concentrations of insulin (1.6 μM) or lower concentrations of IGF‐I (25 nM) to differentiate, and expression of MyoD1 is not constitutive. IGF receptors were studied in microsomal membranes of proliferating and quiescent myoblasts and myotubes. IGF‐II binding was also studied in inducible myoblasts transfected with the MyoD1 cDNA (clone EP5).Both inducible and permissive cells exhibited a single class of binding sites with similar affinity for IGF‐I (Kd 0.8–1.2 nM). Affinity cross‐linking of [125I]IGF‐I to microsomal membranes, under reducing conditions, revealed a binding moiety with an apparent molecular mass of 130 kDa in permissive cells and 140 kDa in inducible cells, which corresponded to the α subunit of the IGF‐I receptor.In permissive quiescent myoblasts, linear Scatchard plots suggested that [125I]IGF‐II bound to a single class of binding sites (Kd 0.6 nM) compatible with binding to the IGF‐II/M6P receptor. This was confirmed by affinity cross‐linking experiments showing a labeled complex with an apparent molecular mass of 260 kDa and 220 kDa when studied under reducing and non‐reducing conditions, respectively.In contrast, competitive inhibition of [125I]IGF‐II binding to inducible quiescent myoblasts generated curvilinear Scatchard plots which could be resolved into two single classes of binding sites. One of them corresponded to the IGF‐II/M6P receptor (Kd 0.2 nM) as evidenced by cross‐linking experiments. The second was the binding site of highest affinity (Kd 0.04 nM) which was less inhibited by IGF‐I than by IGF‐II and was not inhibited by insulin. It migrated in SDS/PAGE at a position equivalent a molecular mass of 140 kDa, under reducing conditions, and at approximately 300 kDa, under non‐reducing conditions. The labeling of this atypical binding moiety was not inhibited by anti(IGF‐II/M6P‐receptor) immunoglobulin. It was also observed in permissive and inducible myoblasts at proliferating stage. It was absent for permissive quiescent myoblasts and from permissive and inducible myotubes. Forced expression of MyoD1 in inducible cells (EP5 cells) dramatically reduced [125I]IGF‐II binding to this atypical receptor.It emerges from these experiments that C2 cells express a putative α2β2 IGF‐II receptor structurally related to the insulin/IGF‐I receptor family. It is present in myoblasts but not in myotubes. The possible relationship between expression of this receptor and expression of MyoD1 as well as myoblast differentiation is discussed.Keywords
This publication has 51 references indexed in Scilit:
- Insulin-like growth factor-II/mannose 6-phosphate receptor is incapable of activating GTP-binding proteins in response to mannose 6-phosphate, but capable in response to insulin-like growth factor-IIBiochemical and Biophysical Research Communications, 1990
- Control of myogenesis in the mouse myogenic C2 cell line by medium composition and by insulin: Characterization of permissive and inducible C2 myoblastsDifferentiation, 1988
- Specificity of insulin-like growth factor binding to type-II IGF receptors in rabbit mammary gland and hypophysectomized rat liverBiochemical and Biophysical Research Communications, 1987
- Demonstration and structural comparison of receptors for insulin-like growth factor-I and -II (IGF-I and -II) in brain and blood-brain barrierBiochemical and Biophysical Research Communications, 1987
- Heterogeneity of insulin-like growth factor-I affinity for the insulin-like growth factor-II receptor: Comparison of natural, synthetic and recombinant DNA-derived insulin-like growth factor-IBiochemical and Biophysical Research Communications, 1987
- Purification of the Insulin-like growth factor II (IGF-II) receptor from an IGF-II-producing cell line, and generation of an antibody which both immunoprecipitates and blocks the type 2 IGF receptorBiochemical and Biophysical Research Communications, 1986
- IGF-II binding on human lymphoid cells: Demonstration of a common high affinity receptor for insulin like peptidesBiochemical and Biophysical Research Communications, 1984
- pH dependence of dissociation of the ovine prolactin rabbit mammary receptor complexBiochemical and Biophysical Research Communications, 1983
- A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye bindingAnalytical Biochemistry, 1976
- Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4Nature, 1970