Pallop KarnchanaphanurachRossen MirchevIonita GhiranJohn M. AsaraBrigitte Papahadjopoulos-SternbergAnne Nicholson-WellerDavid E. GolanHarvard UniversityHarvard Medical SchoolNanoAnalytical LaboratoryBrigham and Women's HospitalMahidol University2018-09-132018-09-132009-04-01Journal of Clinical Investigation. Vol.119, No.4 (2009), 788-80115588238002197382-s2.0-65249110965https://repository.li.mahidol.ac.th/handle/20.500.14594/28133Decay-accelerating factor (DAF, also known as CD55), a glycosylphosphatidylinositol-linked (GPI-linked) plasma membrane protein, protects autologous cells from complement-mediated damage by inhibiting complement component 3 (C3) activation. An important physical property of GPI-anchored complement regulatory proteins such as DAF is their ability to translate laterally in the plasma membrane. Here, we used singleparticle tracking and tether-pulling experiments to measure DAF lateral diffusion, lateral confinement, and membrane skeletal associations in human erythrocyte membranes. In native membranes, most DAF molecules exhibited Brownian lateral diffusion. Fluid-phase complement activation caused deposition of C3b, one of the products of C3 cleavage, onto erythrocyte glycophorin A (GPA). We then determined that DAF, C3b, GPA, and band 3 molecules were laterally immobilized in the membranes of complement-treated cells, and GPA was physically associated with the membrane skeleton. Mass spectrometry analysis further showed that band 3, α-spectrin, β-spectrin, and ankyrin were present in a complex with C3b and GPA in complement-treated cells. C3b deposition was also associated with a substantial increase in erythrocyte membrane stiffness and/or viscosity. We therefore suggest that complement activation stimulates the formation of a membrane skeleton-linked DAF-C3b-GPA-band 3 complex on the erythrocyte surface. This complex may promote the removal of senescent erythrocytes from the circulation.Mahidol UniversityMedicineArticleSCOPUS10.1172/JCI36088