A deletion in the extracellular domain of the alpha platelet-derived growth factor (PDGF) receptor differentially impairs PDGF-AA and PDGF-BB binding affinities.

M.A Heidaran,J.C Yu,R.A Jensen,J.H Pierce,S.A Aaronson

doi:10.1016/s0021-9258(19)50668-8

Abstract

32D cells transfected with the human alpha platelet-derived growth factor receptor (alpha PDGFR) bind PDGF-AA, -AB, and -BB isoforms with high affinity, and the binding of each can be efficiently competed by all three isoforms. In an effort to develop better understanding of spatial relationships of binding sites for PDGF-AA and -BB, we constructed an alpha PDGFR mutant which deleted amino acids 150-189 within its extracellular domain. This mutant showed a marked decrease in high affinity binding sites for PDGF-AA without comparable alteration in affinity for PDGF-BB. These findings imply that the high affinity binding sites for PDGF-AA and PDGF-BB in the alpha PDGFR extracellular domain are not structurally coincident.

Highlights

Platelet-derived growth factor (PDGF)’ is a potent mitogen involved in the normal proliferation and differentiation of a variety of connective tissue and cell types [1,2]
PDGF-AA activates only aPDGFRs, while PDGF-BB interacts at high affinity with dimers involving eitherorboth receptors
These results indicated that theaRA150-19m0utant was expressed a t a level comparable with that of the aPDGFR in32D cell transfectants

Summary

EXPERIMENTAL PROCEDURES

Platelet-derived growth factor (PDGF)’ is a potent mitogen involved in the normal proliferation and differentiation of a variety of connective tissue and cell types [1,2]. Thereis accumulating evidence that PDGF activation involves recruitment of receptor dimers [7,8,9,10,11]. PDGF-AA activates only aPDGFRs, while PDGF-BB interacts at high affinity with dimers involving eitherorboth receptors. Accumulating evidence indicates that PDGF-AB activates aPDGFR or aPPDGFR dimers but does not induce. A t each cloning step, restriction mapping and sequence analysis were performed to ensure identityof the deletion mutant. For Scatchard analysis and cross-competition studies, 32D cells were incubated in the presence of labeled PDGF-BB (40 ng/ml) or PDGF-. AA (40 ng/ml), after preincubation with varying concentrations of unlabeled PDGF ligand as indicated Data obtained from these experiments were analyzed by the method of Scatchard [16] using Thecells were harvested by an automatic cell harvestor and counted ina Beckman LS1701 counter

RESULTS

DNA synthesis was measured by by

DISCUSSION