Activin and inhibin binding to the soluble extracellular domain of activinreceptor II

Activins and inhibins belong to the transforming growth factor-beta-like su perfamily of growth and differentiation factors that exert pleiotropic effe cts in many target tissues. Heteromeric association of activin with two str ucturally related receptor serine/threonine kinases, activin receptor types I and II, initiates downstream signaling events. The extracellular domain of type II mouse activin receptor (ActRII ECD) was expressed in the baculov irus system, purified in three steps by lectin affinity, anion exchange, an d reverse phase chromatography, and further characterized by mass spectrome try. The reduction in the apparent size of the purified ActRII ECD on SDS-P AGE after treatment with glycosidases provided evidence for N- and O-linked oligosaccharides. Specific receptor/ligand complexes of [I-125]activin A t o ActRII ECD or [I-125]ActRII ECD to activin A were analyzed by cross-linki ng and immunoprecipitation. Two major radiolabeled bands were observed on S DS-PAGE with mobilities consistent with the expected size of ActRII ECD/bet a(A) or ActRII ECD/beta(A)beta(A). When inhibin A was cross-linked to [I-12 5]ActRII ECD, a slower migrating complex corresponding to ActRII ECD/beta(A )alpha was also observed. The apparent dissociation constant (K-d) for acti vin A binding to ActRII ECD was 2-7 nM. This K-d value is approximately an order of magnitude greater than that of the full-length membrane-associated type II receptor. Treatment of cultured rat anterior pituitary cells with ActRII ECD attenuated FSH secretion in response to exogenous activin A or e ndogenous activin B. These data indicate that the soluble ActRII ECD has st ructural determinants that are sufficient for high affinity Ligand binding.