Soluble 34K Binding Protein Inhibits the Binding of Insulin-like Growth Factor I to Its Cell Receptors in Human Secretory Phase Endometrium: Evidence for Autocrine/Paracrine Regulation of Growth Factor Action*

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Abstract
The human secretory phase endometrium synthesizes and secretes a 34K insulin-like growth factor (IGF)-binding protein designated placental protein 12. We now report that membrane preparations of human endometrium possess IGF receptors that compete with the soluble binding protein for binding to IGF-I. Multipulication-stimulating activity and insulin were 1% and 0.1% as potent as recombinant (Thr59)IGF-I in inhibiting the binding of [126I](Thr59)IGF-I to endometrial membranes. Scatchard plots for the IGF-I binding data were curvilinear, and the apparent affinities [Ka=1.4.+-.0.2(.+-.SEM) .times.1.09 M-1] for (Thr59)IGF-I (high affinity site) did no change during the menstrual cycle. Affinity cross-linking of [125I](Thr59)IGF-I to endometrial membranes revealed two major bands with mol wt of 130K and 260K on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. The 130K band is consistent with the a.alpha.-subunit of the type I IGF receptor. The 260K band is either the type II IGF receptor or represents cross-linking (dimer) of two .alpha.-subunits of the type I receptor. A less intense band at about 40K was also seen in all membrane preparations. It comigrated with the cross-linked purified 34K binding protein. The band was more intensely labeled when the tracer was cross was cross-linked to proteins in the cytosol fractions of late secretory phase endometria. By specific RIA, the 34K binding protein was detected in the cytosol of late secretory phase endometrial only. Newly synthesized binding protein, which contaminated membrane preparations, caused an apparent increase in the binding of (Thr59)IGF-I to the membranes prepared from late secretory phase endometria when studied by competitive binding assay. In contrast, purified binding protein prevented the binding of [125I](Thr59)IGF-I to membrane receptors, as confirmed by affinity cross-linking. These results suggest that the 34K IGF-binding protein, secreted by the human endometrium in a cyclic fashion, has a significant role in inhibiting the receptor binding and, thus, the possible biological action of IGF-I in the endometrium in an autocrine/paracrine manner.

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