Summary

GPIbβ is disulfide-linked to GPIbα to form GPIb, a platelet receptorfor von Willebrand factor (vWF). GPIb is in turn non covalently linkedto GPIX and GPV to form the GPIb/V/IX complex. Apart from its contributionto controlling surface expression of the complex, the exactfunction of GPIbβ is not well established due to a lack of suitable ligandsor antibodies. The present report describes a monoclonal antibody(RAM.1) that labeled the 26 kDa GPIbβsubunit on western blotsand coprecipitated the three subunits of the GPIb/IX complex from lysatesof platelets and transfected CHO and K562 cells. RAM.1 bound toGPIbβ deleted of its intracellular domain whereas Gi27, directedagainst intracellular GPIbβ, did not. Using synthetic peptides, theRAM.1 epitope was mapped to a putative cysteine loop within theCOOH-terminal leucine-rich flanking region. In functional assays,RAM.1 had no effect on platelet aggregation induced by ADP, collagenor thrombin, but inhibited ristocetin induced platelet agglutination andbotrocetin induced vWF binding. RAM.1 inhibited adhesion ofGPIb/V/IX transfected K562 cells to a vWF matrix under flow, increasedtheir rolling velocity and decreased the resistance of cells to detachmentat high shear. This study suggests a role of GPIbβ in modulatingthe adhesive properties of GPIb/V/IX and describes a useful tool to analyzethe exact functions of GPIbβ.