Structurally distinct membrane-associated and soluble forms of GH-binding protein in the mouse

Abstract

It has previously been shown that the large increase in GH-binding capacity of mouse liver microsomes during pregnancy is due largely to an increase in the amount of GH-binding protein (GHBP), with a more modest increase in GH receptor (GHR). Here we show that mouse liver GHBP is predominantly present as a membrane-associated protein structurally distinct from the soluble form of GHBP present in serum. Liver GHBP is associated with both intracellular membranes and the plasma membrane. Membrane-associated GHBP and soluble GHBP appear to be identical polypeptides distinguished by the addition of different N-glycans to asparagine residues. The pattern of release of GHBP from membranes by various treatments indicates that GHBP associates with membranes through noncovalent interactions with one or more membrane protein, but not with GHR. Covalent crosslinking provides evidence for several GHBP-associated membrane polypeptides, with molecular masses ranging from 58 kDa to over 200 kDa.These studies in the mouse and similar studies in the rat suggest that GHBP is an important cell-surface receptor for GH in the liver of these species. We postulate that an arginine-glycine-aspartic acid sequence found on rat and mouse GHBP but absent in other species is responsible for the association of GHBP with the plasma membrane by binding to one or more integrins on the surface of liver cells.

Differential regulation of membrane associated-growth hormone binding protein (MA-GHBP) and growth hormone receptor (GHR) expression by growth hormone (GH) in mouse liver

“…2). The MA-GHBP forms from every mouse line studied showed the same molecular weight, which was different from the soluble GHBP molecular forms, corroborating what has been previously described by us and other authors [23,25,26].…”

“…To act as a negative regulator, MA-GHBP needs to be associated with the extracellular surface of the plasma membrane, although not every form of the MA-GHBP is at the extracellular membrane [23,26]. Frick et al [26] have shown that GHBP accounts for 50% of GH binding to plasma membrane of rat adipocytes, but most of MA-GHBP is associated with intracellular membranes, principally with the endoplasmic reticulum [23,43].…”

“…Frick et al [26] have shown that GHBP accounts for 50% of GH binding to plasma membrane of rat adipocytes, but most of MA-GHBP is associated with intracellular membranes, principally with the endoplasmic reticulum [23,43]. This MA-GHBP differs from soluble GHBP, either from serum or cytosol, since MA-GHBP forms show lower Mr (37 and 42) than the soluble GHBP forms (43 and 48) [22,25,43], but far from the Mr estimated for the aminoacidic sequence of the protein backbone (31 kDa).…”

“…These isoforms conserve the transmembrane domain but have a short cytoplasmic domain and are thus unable to transduce GH signal [17][18][19]21]. In rodents, GHBP was found associated to cellular membranes, representing the predominant GHR short form (MA-GHBP) [22][23][24][25][26]. MA-GHBP contains the extracellular domain of the GHR but lacks both the transmembrane and cytoplasmic domains, which are replaced by a short ''hydrophilic tail'' produced by alternative splicing that can be useful to specifically detect rodent GHBP.…”

Structure and regulation of expression of the mouse GH receptor

“…Recent studies have demonstrated that mouse liver GHBP is predominantly present as a membrane-associated protein structurally distinct from the soluble form of GHBP present in serum (Cerio et al 2002). Liver GHBP is associated with both intracellular membranes and the plasma membrane.…”