Platelet glycocalicin. Its membrane association and solubilization in aqueous media

Solum, Nils Olav; Hagen, Inger; Filion-Myklebust, C.; Stabæk, T.

doi:10.1016/0005-2736(80)90102-9

Cited by 98 publications

(44 citation statements)

References 17 publications

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“…The platelet GPIb-IX-V complex is the key platelet receptor for VWF (17)(18)(19). Human platelet GPIbα has abundant Oglycans, including sialylated versions of core 1 and core 2 O-glycans (20,21), and a majority of sialic acid on the platelet surface appears to be associated with GPIbα (22). Although the O-glycan structures in murine GPIbα are unknown, there are 73 potential glycosylation sites in murine GPIbα in comparison with the 42 potential O-glycosylation sites in human GPIbα.…”

Section: Discussionmentioning

confidence: 99%

Platelet biogenesis and functions require correct protein O-glycosylation

Wang¹,

Jobe

Ding³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Platelets express a variety of membrane and secreted glycoproteins, but the importance of glycosylation to platelet functions is poorly understood. To explore the importance of O-glycosylation, we generated mice with a targeted deletion of Cosmc in murine endothelial/hematopoietic cells (EHC) (EHC Cosmc −/ y ). X-linked Cosmc encodes an essential chaperone that regulates protein O-glycosylation. This targeted mutation resulted in lethal perinatal hemorrhage in the majority of mice, and the surviving mice displayed severely prolonged tail-bleeding times and macrothrombocytopenia. EHC Cosmc − /y platelets exhibited a marked decrease in GPIb-IX-V function and agonist-mediated integrin αIIbβ3 activation, associated with loss of interactions with von Willebrand factor and fibrinogen, respectively. Significantly, three O-glycosylated glycoproteins, GPIbα, αIIb, and GPVI normally on platelet surfaces that play essential roles in platelet functions, were partially proteolyzed in EHC Cosmc − /y platelets. These results demonstrate that extended O-glycans are required for normal biogenesis of the platelets as well as the expression and functions of their essential glycoproteins, and that variations in O-glycosylation may contribute to altered hemostasis.

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Section: Discussionmentioning

confidence: 99%

Platelet biogenesis and functions require correct protein O-glycosylation

Wang¹,

Jobe

Ding³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…The modifications took approximately 160 min to complete because wheat germ agglutinin could only precipitate GP Ib␣ after a 150-min chase. We chose wheat germ agglutinin because it recognizes the terminal sialic acids on O-linked oligosaccharides of GP Ib␣ (32,41,42). Sialylation is the last modification of O-linked carbohydrate chains to occur before glycoproteins are either secreted or expressed on the plasma membrane.…”

Section: Discussionmentioning

confidence: 99%

Synthesis, Assembly, and Intracellular Transport of the Platelet Glycoprotein Ib-IX-V Complex

Dong¹,

Gao²,

López³

1998

Journal of Biological Chemistry

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The platelet glycoprotein Ib-IX-V complex plays critical roles in adhering platelets to sites of blood vessel injury and in platelet aggregation under high fluid shear stress. The complex is composed of four membrane-spanning polypeptides: glycoprotein (GP) Ib␣, GP Ib␤, GP IX, and GP V. Glycoprotein Ib␣ contains a binding site for von Willebrand factor through which it mediates platelet adhesion; GP V is required for the complex to bind thrombin with high affinity; and both GP Ib␤ and GP IX are necessary for efficient plasma membrane expression of the complex. To further define the roles of the individual polypeptide subunits in the biosynthesis and intracellular transport of the GP Ib-IX-V complex, we studied full and partial complexes expressed in heterologous mammalian cells. We found that the full complex was formed within minutes in the endoplasmic reticulum before being transported into the Golgi cisternae. Approximately 160 min were required for the complex to be fully processed and to appear on the plasma membrane. About 25% of GP Ib␣ expressed as part of either a GP Ib-IX complex or a GP Ib-IX-V complex was degraded through a nonlysosomal pathway. Over 60% of GP Ib␣, however, was degraded when it was expressed in partial complexes with only GP Ib␤ or GP IX. The increased degradation was blocked by treating cells either with brefeldin A to prevent the transport of proteins from the endoplasmic reticulum to the Golgi or with lysosomal inhibitors, indicating that GP Ib␣ expressed in partial complexes was targeted to the lysosomes for degradation. These results indicate that the presence of both GP Ib␤ and GP IX, but not the presence of GP V, is required for efficient processing and targeting of GP Ib␣ to the plasma membrane. Absence of either GP Ib␤ or GP IX increased the rate of GP Ib␣ degradation, providing an explanation for why mutation of their genes leads to deficient GP Ib␣ expression and platelet adhesion in Bernard-Soulier syndrome, the deficiency disorder of the complex.When the blood vessel is injured, platelets adhere to the subendothelial matrix exposed at the site of injury. This adhesion is initiated by an interaction between von Willebrand factor exposed on the subendothelium and the glycoprotein (GP)1 Ib-IX-V complex on the platelet surface (1, 2). The same interaction also precipitates pathological platelet aggregation induced by high shear stresses, which occurs at sites of arterial stenosis (3-5).The GP Ib-IX-V complex is composed of four polypeptide subunits, GP Ib␣, GP Ib␤, GP IX, and GP V (1, 6). Glycoprotein Ib␣ is disulfide-linked to GP Ib␤; GP IX and GP V associate with the complex by noncovalent means (7-9). Although the four polypeptides are encoded by different genes (10 -13), they are homologous to each other, all belonging to a phylogenetically widespread protein family defined by the presence of a motif containing tandemly repeated leucine-rich sequences (1,14). Among the four subunits, GP Ib␣ is the largest and so far the only subunit shown to bind von Willebrand factor and...

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“…The calciumactivated protease releases a 150-kDa piece from GPIb that is termed glycocalicin [l, 5, 61. Chymotrypsin and trypsin have been reported to cleave a 'macroglycopeptide' of about 120 kDa from GPIb [7,8], implying that this was the external part of glycocalicin. However, with short digestion times both chymotrypsin and trypsin gave a glycocalicin-like fragment in the supernatant as well as the 'macroglycopeptide' and 45-kDa piece (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…8B). Trypsin is known to cleave glycocalicin into fragments of 45 kDa and 120 kDa [7] and glycocalicin treated with elastase gave two similar fragments. In the gel electrophoresis system used here, the mass of the highly glycosylated fragment was 90 kDa rather than 120 kDa but masses of 108 kDa and 60 kDa have also been reported for this fragment using various methods [47].…”

Section: Discussionmentioning

confidence: 99%

“…Glycocalicin [5] is a ma.jor proteolytic cleavage product of GPIb, produced by the action of the endogenous platelet calcium-activated protease [I, 61. I t can itself be split by trypsin into two initial fragments [7], one of apparent mass 120 kDa which is heavily glycosylated and does not contain the von Willebrand factor binding site [8], and the other of 45 kDa which is less glycosylated, contains at least one intramolecular disulfide bond [6, 91 and a binding site for thrombin [lo]. Antibodies to GPIb, both monoclonal [ l l , 121 and polyclonal [13], have been shown to inhibit the platelet response to thrombin.…”

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confidence: 99%

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Structure and function of platelet membrane glycoproteins Ib and V

Wicki

Clemetson

1985

European Journal of Biochemistry

143

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Treatment of platelets with human leukocyte elastase causes a rapid loss in response to von Willcbrand factor and a biphasic loss in rcsponse to thrombin, first rapid then more slowly. The rapid phase corresponds to cleavage of a 45-kDa glycopeptide from the extracellular end of membrane glycoprotein GPIb. Longer treatment removes 80-kDa and 90-kDa glycopcptides and a glycopeptide corresponding to the major part of GPV. The 45-kDa and 90-kDa species could be obtained by elastase treatment of glycocalicin, the major proteolytic cleavage product of GPIb. Polyclonal rabbit antibodies against the purified 45-kDa glycopeptide had the same effect on the action of von Willebrand factor and thrombin on platelets as cleavage of GPIb by elastase. These results indicate that both the von Willebrand factor and thrombin binding sites on GPIb are located in the samc region on thc outside of the molecule. Thrombin activation of platelets involves at least two receptors, one on the 45-kDa glycopeptide, which when occupied causes an increase in the speed of response of the platelets to the cleavage of the second. GPV, a candidate for the second receptor, is a hydrophobic glycoprotein that is cleaved from the platelet membrane by several proteases. Proteases that do not activate platelets but degrade the second receptor remove larger fragments from GPV than do proteases such as thrombin or trypsin which activate platelets.

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Platelet glycocalicin. Its membrane association and solubilization in aqueous media

Cited by 98 publications

References 17 publications

Platelet biogenesis and functions require correct protein O-glycosylation

Platelet biogenesis and functions require correct protein O-glycosylation

Synthesis, Assembly, and Intracellular Transport of the Platelet Glycoprotein Ib-IX-V Complex

Structure and function of platelet membrane glycoproteins Ib and V

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