Insulin‐like growth factors bind with high affinity to specific binding proteins in extracellular fluids. To identify structural characteristics of IGF‐binding proteins that might define their physiological roles, we determined the complete primary structure of a novel human IGF‐binding protein (IGFBP‐2) from a cloned cDNA. The cDNA encodes a 328 amino acid IGF‐binding protein precursor which contains a 39‐residue signal peptide. The mature 289 amino acid IGFBP‐2 has a predicted Mr of 31,325. Chinese hamster ovary (CHO) cells stably transformed with the IGFBP‐2 cDNA secreted a 36 kd protein which bound, with different affinities, IGFII and IGFI, but did not bind insulin. The predicted protein sequence of this IGF‐binding protein shares extensive amino acid homology (greater than 85%) with the IGF‐binding protein secreted by rat BRL‐3A cells, but less than 40% homology with human IGFBP‐1. Therefore IGFBP‐2, and not IGFBP‐1 as previously suggested, represents the human homologue of the rat BRL‐BP (alpha IGFBP‐2). Moreover, from alignment of the predicted protein sequences of IGFBP‐1 and IGFBP‐2, extensive conservation of the distribution of cysteine residues is observed. Although the overall amino acid homology shared by these proteins is not high, we suggest that they represent a family of structurally related human IGFBPs. Southern blot analysis of human DNA demonstrates that IGFBP‐2 is encoded by a single‐copy gene, different from that of IGFBP‐1.

Rat serum contains two major forms of insulin-like growth factor (IGF) binding proteins (BPs) that have apparent mol wts of about 35,000 and 150,000. We have isolated a cDNA clone encoding an IGF-BP whose N-terminal sequence is completely homologous to the NH2-terminal of the Buffalo rat liver cells-3A BP. The 270 amino acid mature protein has a predicted mol wt of 29,500. It contains a cysteine rich domain at each end of the molecule and an Arg-Gly-Asp (RGD) tripeptide motif near its C-terminus which suggests that this BP might associate with integrin cell surface receptors. The mature protein shares only partial homology with two published human IGF-BPs. Northern blot analysis shows that its mRNA is abundant in several fetal tissues, in adult brain, testes, ovaries, and kidney. Expression in the liver is high in fetal life but decreases to a barely detectable level in adulthood. However, upon hypophysectomy, the mRNA level increases at least 20-fold which suggests a hormonal regulation for the hepatic production of this small IGF-BP.

The goal of this study was to find out whether GH or insulin regulate the mRNA expression of the fetal binding protein of insulin-like growth factor (IGFBP-2). Primary hepatocytes from adult rats were used as a test system. IGFBP-2 mRNA was abundant in cells cultured in the absence of hormones and markedly reduced in cultures containing insulin. Addition of GH had no effect on IGFBP-2 mRNA levels although the cells are responsive to GH as demonstrated by a GH mediated elevation of IGF l mRNA levels. Half-maximal down-regulation of IGFBP-2 mRNA levels occurred at an insulin concentration of 1 to 2 x 10(-10) M. The finding that insulin is a potent negative regulator of hepatic IGFBP-2 mRNA levels suggests a physiologically important regulatory link between the two hormones insulin and IGF l.

Hepatic mRNA levels of insulin-like growth factor I (IGF I) and of the fetal, nonglycosylated 32 kDa IGF-binding protein (BP) were analysed in diabetic, diabetic insulin-and IGF I-treated rats as well as in age-matched, healthy control animals. IGF I mRNA levels are reduced in diabetic rats and increased by insulin treatment. In contrast, the infusion of IGF I does not significantly upregulate IGF I mRNA levels. Fetal IGF BP mRNA expression is very low in healthy control animals, but high levels are found in diabetic rats. Insulin therapy lowers fetal IGF BP mRNA levels, whereas IGF I has no effect. We propose that insulin is a major regulator of the 32 kDa IGF BP levels in adult rats.Insulin-like growth factor I; mRNA, Protein, IGF-binding; (Streptozotocin-diabetic rat)

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