Insulin‐induced decrease in 5′‐nucleotidase activity in skeletal muscle membranes

Dearey

Pünter

1993

Biochemical Journal

Sulphonylurea drugs stimulate glucose transport and metabolism in muscle and fat cells in vitro. The molecular basis for the insulin-mimetic extrapancreatic effects of these oral antidiabetic therapeutic agents is unknown at present. Here we demonstrate that incubation of 3T3 adipocytes with the novel sulphonylurea, glimepiride, causes a time- and concentration-dependent release of the glycosylphosphatidylinositol (GPI)-anchored ecto-proteins, 5′-nucleotidase, lipoprotein lipase and a 62 kDa cyclic AMP (cAMP)-binding protein from the plasma membrane into the culture medium. The change in the localization is accompanied by conversion of the membrane-anchored amphiphilic proteins into their soluble hydrophilic versions, as judged by pulse-chase experiments and Triton X-114 partitioning, and by appearance of anti-cross-reacting determinant (CRD) immunoreactivity of the released proteins as shown by Western blotting. Metabolic labelling of cells with myo-[14C]inositol demonstrates that inositol is retained in the major portion of released lipoprotein lipase and cAMP-binding ectoprotein. The identification of inositol phosphate after deamination of these proteins with nitrous acid suggests cleavage of their GPI membrane anchor by a GPI-specific phospholipase C. However, after longer incubation with glimepiride the amount of soluble versions of the GPI-proteins lacking inositol and anti-CRD immunoreactivity increases, which may be caused by additional drug-stimulated hydrolytic events within their GPI structure or C-termini. Since insulin also stimulates membrane release of these GPI-modified proteins, and in combination with glimepiride in a synergistic manner, sulphonylurea drugs may exert their peripheral actions in adipose tissue by using (part of) the insulin postreceptor signalling cascade at the step of activation of a GPI-specific phospholipase C.

Section: Discussionsupporting

confidence: 53%

The sulphonylurea drug, glimepiride, stimulates release of glycosylphosphatidylinositol-anchored plasma-membrane proteins from 3T3 adipocytes

Dearey

Pünter

1993

Biochemical Journal

“…A portion of at least one GPI-anchored protein, the ecto-receptor protein for cAMP, is released from the membrane through the activation of an endogenous PLC. This anchor cleavage following a nutritional upshift qualitatively resembles the lipolytic membrane release of some GPIanchored proteins (e.g., 5'-nucleotidase, alkaline phosphatase, lipoprotein lipase, and heparan sulfate proteoglycan) observed with cultured cells and tissues from vertebrates in response to glucose and insulin (Ishihara et al, 1987;Romero et al, 1988;Chan et al, 1988;Klip et al, 1988;Lisanti et al, 1989). In both cases, yeast as shown here andas far as is known-vertebrate cells, a phospholipase is activated in response to extrinsic signals.…”

Section: Discussionsupporting

confidence: 65%

Glucose induces lipolytic cleavage of a glycolipidic plasma membrane anchor in yeast

The Journal of Cell Biology

Bandlow

1993

Abstract. In the yeast Saccharomyces cerevisiae an amphiphilic cAMP-binding protein has been found recently to be anchored to plasma membranes by virtue of a glycolipid structure Bandlow, 1991a, 1992). The cAMP-binding parameters of this protein are affected by the lipolytic removal of the glycosylphosphatidylinositol (GPI) membrane anchor by exogenous (G)PI-specific phospholipases C or D (PLC or PLD) (Miiller and Bandlow, 1993) suggesting a regulatory role of glycolipidic membrane anchorage. Here we report that transfer of yeast cells from lactate to glucose medium results in the conversion of the amphiphilic form of the cAMP receptor protein into a hydrophilic version accompanied by the rapid loss of fatty acids from the GPI anchor of the [14C]palmitic acid-labeled protein. Analysis of the cleavage site identifies [~4C]inositol phosphate as the major product after treatment of the soluble, [t4C]inositol-labeled protein with nitrous acid which destroys the glucosamine constituent of the anchor. Together with the observed cross-reactivity of the hydrophilic fragment with antibodies directed against the cross-reacting determinant of soluble trypanosomal variable surface glycoproteins (i.e., myo-inositol-l,2-cyclic phosphate) this demonstrates that, in membrane release, the initial cleavage event is catalyzed by an intrinsic GPI-PLC activated upon transfer of cells to glucose medium. Release from the plasma membrane in soluble form requires, in addition, the presence of high salt or or-methyl mannopyranoside, or the removal of the carbohydrate moieties, because otherwise the protein remains associated with the membrane presumably at least in part via its N-glycosidic carbohydrate side chains. The data point to the possibility that cleavage of the anchor could play a role in the transfer of the signal for the nutritional situation to the interior of the cell.

“…In this context, it appears important that in yeast cells, the anchor of a GPI-anchored protein, Gce1p, is lipolytically cleaved in response to a nutritional signal, i.e., the transfer of the cells to glucose (37). Particularly compelling is the notion that anchor cleavage following nutritional upshift qualitatively resembles the lipolytic membrane release in response to insulin or to sulfonylurea drugs (in the presence of glucose) reported for some GPI-anchored proteins (e.g., 5Ј-nucleotidase, alkaline phosphatase, lipoprotein lipase, and heparan sulfate proteoglycan) in cultured BC 3 H1 myocytes, 3T3-L1 adipocytes, rat hepatocytes, and the isolated rat diaphragm (8,20,21,26,37,48). It has been proven recently that in yeast cells, Gce1p is processed by a GPI-PL of type C. In mammals, GPI-PLs of types C and D could be identified and partially purified or isolated (10,18,32).…”

Section: Discussionmentioning

confidence: 94%

Glucose-Induced Sequential Processing of a Glycosyl-Phosphatidylinositol-Anchored Ectoprotein in Saccharomyces cerevisiae

Molecular and Cellular Biology

Gross

Wied

et al. 1996

An increasing number of surface proteins are found anchored at the plasma membrane of various tissues and cell types by virtue of a glycosyl-phosphatidylinositol (GPI) structure instead of a transmembrane protein domain. This kind of membrane anchor typically contains, in addition to the myoinositol-harboring phospholipid, a carbohydrate core consisting of one nonacetylated glucosamine and three mannose residues in characteristic linkage. The latter may be branched and carry additional mannose, galactose, and/or phosphoethanolamine moieties. The preformed anchor structure is linked by a phosphodiester and an ethanolamine bridge to the carboxy terminus of the respective protein, thereby replacing a proteinaceous transmembrane domain present in the newly synthesized protein precursor (for reviews, see references 14, 17, 31, 52, and 56). The physiological significance of membrane anchorage by a glycolipidic structure has remained greatly obscure. Since GPI-anchored proteins have been detected recently also in lower eucaryotes, including the yeast Saccharomyces cerevisiae (9,35,43,54), the evolutionary conservation from S. cerevisiae to humans of the GPI anchor argues that it confers properties on the respective proteins that are not achieved by a proteinaceous transmembrane domain.It has been proposed that the spatial requirements, clustering behavior, and diffusion properties of GPI-anchored proteins in the outer leaflet of plasma membranes are the parameters critical for the use of either a transmembrane domain or a GPI moiety for membrane anchorage of a protein (7,13,61,62). In addition, it has been proposed, on the basis that, in situ, GPI anchors may be cleaved upon activation of endogenous (glycosyl)-phosphatidylinositol-specific phospholipases ([G]PIPLs), that GPI anchorage plays a role in regulating the concentration of certain proteins at the cell surface or in controlling protein localization and topology: in HeLa cells and in cultured bone marrow stromal cells, GPI-anchored decay-accelerating factor and heparan sulfate proteoglycan, respectively, are apparently removed constitutively from the plasma membrane and released into the culture medium by the action of an endogenous of type D (G)PI-PL ([G]PI-PLD) (32). In the yeast S. cerevisiae, expression of ␣-agglutinin at the cell wall depends on its synthesis as a GPI-anchored protein and occurs via an intermediate which lacks the myo-inositol moiety of the GPI anchor glycan (28, 59). However, other than (G)PI-PLs, which leave the myo-inositol residue attached to the glycan structure, processing enzymes of GPI anchors have not been identified. In addition to these cases of constitutive GPI anchor processing, in some vertebrate cells certain external signals, provided by serum factors like insulin and growth factors or by drugs, have been found to activate phospholipases which are capable of cleaving GPI anchors (20,26,40,48). In the case of the insulin-or sulfonylurea-induced lipolytic cleavage of the GPI anchors of lipoprotein lipase, 5Ј-nucleotidase, and ...