CD59, an 18-20-kD complement inhibitor anchored to the membrane via glycosyl phosphatidylinositol (GPI), can induce activation of T cells and neutrophils upon cross-linking with antibody. GPI-anchored molecules cocluster in high mol wt detergent-resistant complexes containing tyrosine kinases that are implicated in the signaling pathway. Exogenous, incorporated GPI-anchored molecules are initially unable to induce activation, presumably because they are not associated with kinases. Here we demonstrate that erythrocyte-derived CD59 incorporated in a CD59-negative cell line acquires signaling capacity in a time-dependent manner. Confocal microscopy revealed an initial diffuse distribution of CD59 that became clustered within 2 h to give a pattern similar to endogenous GPI-anchored molecules. Gel filtration of detergent-solubilized cells immediately after incorporation revealed that CD59 was mainly monomeric, but after 3 h incubation all was in high mol wt complexes and had become associated with protein kinases. Newly incorporated CD59 did not deliver a Ca2+ signal upon cross-linking, but at a time when it had become clustered and associated with kinase activity, cross-linking induced a large calcium transient, indicating that CD59 had incorporated in a specialized microenvironment that allowed it to function fully as a signal-transducing molecule.
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1 November 1995
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November 01 1995
Exogenous glycosyl phosphatidylinositol-anchored CD59 associates with kinases in membrane clusters on U937 cells and becomes Ca(2+)-signaling competent.
C W van den Berg,
C W van den Berg
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
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T Cinek,
T Cinek
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
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M B Hallett,
M B Hallett
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
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V Horejsi,
V Horejsi
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
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B P Morgan
B P Morgan
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
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C W van den Berg
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
T Cinek
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
M B Hallett
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
V Horejsi
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
B P Morgan
Department of Medical Biochemistry, University of Wales College of Medicine, Cardiff, United Kingdom.
Online ISSN: 1540-8140
Print ISSN: 0021-9525
J Cell Biol (1995) 131 (3): 669–677.
Citation
C W van den Berg, T Cinek, M B Hallett, V Horejsi, B P Morgan; Exogenous glycosyl phosphatidylinositol-anchored CD59 associates with kinases in membrane clusters on U937 cells and becomes Ca(2+)-signaling competent.. J Cell Biol 1 November 1995; 131 (3): 669–677. doi: https://doi.org/10.1083/jcb.131.3.669
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