Receptor binding activity and in vitro biological activity of the human FSH charge isoforms as disclosed by heterologous and homologous assay systems- Implications for the structure-function relationship of the FSH variants

Follicle-stimulating hormone (FSH) is produced and secreted in multiple mol ecular forms. These isoforms differ in their oligosaccharide structures, wh ich determine the particular behavior of a given variant in in vitro and in vivo systems. Employing heterologous cell assay systems, this and other la boratories have shown that highly sialylated human FSH variants exhibit low er receptor binding/immunoactivity as well as in vitro bioactivity/immunoac tivity relationships than their less sialylated counterparts. It is not kno wn, however, whether this characteristic behavior of the FSH isoforms is re produced by homologous assay systems, in which unique variants of the recep tor are presumptively expressed. To gain further insights into the structur e-activity relationship of the various FSH isoforms, we analyzed the capaci ty of nine charge isoforms obtained after high-resolution chromatofocusing (pH window, 7.10 to <3.80) of anterior pituitary glycoprotein extracts to b ind and activate their cognate receptor expressed by naturally occurring he terologous cell systems (rat granulosa cells and seminiferous tubule homoge nates) as well as by human embryonic kidney-derived 293 (HEK-293) cells tra nsfected with the human FSH (FSH-R) receptor cDNA. In both (heterologous an d homologous) receptor assay systems, the isoforms displaced I-125-labeled FSH from the receptor in a dose-response manner; however, whereas in the he terologous systems, the receptor binding activity varied according to the e lution pH value/sialic content of the isoforms, with the less acidic varian ts exhibiting higher receptor binding activity (r = 0.851 and 0.495 [p < 0. 01 and p < 0.05] for the granulosa cell and testicular homogenate receptor assay systems, respectively) than the more acidic/sialylated analogs, in th e homologous assay, this relationship was practically absent (r = 0.372, p N.S.). The capacity of the isoforms to induce androgen aromatization by rat granulosa cells followed the same trend shown by its corresponding recepto r assay system (r = 0.864, p < 0.01). Interestingly and in contrast to the results observed in the homologous receptor binding assay, the ability of t he isoforms to induce cAMP production by HEK-293 cells varied according to their elution pH value, with the more sialylated isoforms exhibiting lower potency than their less acidic counterparts (r = 0.852, p < 0.01). The resu lts yielded by the heterologous assays suggest that the different potency o f the isoforms to elicit a biological effect in a naturally occurring recep tor system depends primarily on the particular affinity of the receptor mol ecule for each isoform. The existence of a clear dissociation between recep tor binding and signal transduction in the homologous system indicate that this later function is rather related to the different ability of the FSH g lycosylation variants to induce and/or stabilize distinct receptor conforma tions that may permit preferential or different degrees of activation/inhib ition of a given signal transduction pathway. Thus, the human FSH receptor- transducer system apparently possesses sufficient versatility to respond in a different manner to glycosylation-dependent diverse FSH signals.