Glycosylphosphatidyl inositol-anchored proteins and fyn kinase assemble in noncaveolar plasma membrane microdomains defined by reggie-1 and -2.
Using confocal laser scanning and double immunogold electron microscopy, we demonstrate that reggie-1 and -2 are colocalized in < or =0.1-microm plasma membrane microdomains of neurons and astrocytes. In astrocytes, reggie-1 and -2 do not occur in caveolae but clearly outside these structures. Microscopy and coimmunoprecipitation show that reggie-1 and -2 are associated with fyn kinase and with the glycosylphosphatidyl inositol-anchored proteins Thy-1 and F3 that, when activated by antibody cross-linking, selectively copatch with reggie. Jurkat cells, after cross-linking of Thy-1 or GM1 (with the use of cholera toxin), exhibit substantial colocalization of reggie-1 and -2 with Thy-1, GM1, the T-cell receptor complex and fyn. This, and the accumulation of reggie proteins in detergent-resistant membrane fractions containing F3, Thy-1, and fyn imparts to reggie-1 and -2 properties of raft-associated proteins. It also suggests that reggie-1 and -2 participate in the formation of signal transduction centers. In addition, we find reggie-1 and -2 in endolysosomes. In Jurkat cells, reggie-1 and -2 together with fyn and Thy-1 increase in endolysosomes concurrent with a decrease at the plasma membrane. Thus, reggie-1 and -2 define raft-related microdomain signaling centers in neurons and T cells, and the protein complex involved in signaling becomes subject to degradation.
Stuermer
C A
CA
Department of Biology, University of Konstanz, 78467 Konstanz, Germany. claudia.stuermer@uni.konstanz.de
Lang
D M
DM
Kirsch
F
F
Wiechers
M
M
Deininger
S O
SO
Plattner
H
H
D000818:Animals; D001253:Astrocytes / Q000378:metabolism; D001921:Brain / Q000378:metabolism; D021941:Caveolae / Q000378:metabolism / Q000648:ultrastructure; D022461:Caveolins / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D029941:Fish Proteins; D005677:G(M1) Ganglioside / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D019169:Jurkat Cells / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism* / Q000648:ultrastructure; D018613:Microscopy, Confocal; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism* / Q000648:ultrastructure; D016716:PC12 Cells / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction / Q000502:physiology; D018800:Thy-1 Antigens / Q000378:metabolism*; D019061:src-Family Kinases
D022461:Caveolins; D015815:Cell Adhesion Molecules; D029941:Fish Proteins; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D018800:Thy-1 Antigens; C106354:flotillins; C104436:reggie-2 protein, Carassius auratus; D005677:G(M1) Ganglioside; C490511:Frk protein, rat; D011505:Protein-Tyrosine Kinases; C092830:FRK protein, human; D019061:src-Family Kinases