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标题：O-glycans direct selectin ligands to lipid rafts on leukocytes
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作者：Bojing Shao ; Tadayuki Yago ; Hendra Setiadi 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：28
页码：8661-8666
DOI：10.1073/pnas.1507712112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceLeukocytes partition certain proteins into cholesterol- and sphingolipid-rich membrane regions (lipid rafts) that function as signaling platforms. Inflammatory stimuli cause leukocytes to elongate to form lamellipodia and uropods at opposite ends that facilitate migration. Many raft-associated proteins move to uropods. Proteins are typically thought to use their transmembrane and cytoplasmic domains to associate with rafts. Here, we found that some leukocyte adhesion proteins used carbohydrate modification (glycosylation) of their extracellular domains to associate with lipid rafts. These proteins required preassociation with rafts to transduce signals but, unexpectedly, not to move to uropods. These data define a mechanism for localizing proteins to critical membrane regions of leukocytes. Palmitoylated cysteines typically target transmembrane proteins to domains enriched in cholesterol and sphingolipids (lipid rafts). P-selectin glycoprotein ligand-1 (PSGL-1), CD43, and CD44 are O-glycosylated proteins on leukocytes that associate with lipid rafts. During inflammation, they transduce signals by engaging selectins as leukocytes roll in venules, and they move to the raft-enriched uropods of polarized cells upon chemokine stimulation. It is not known how these glycoproteins associate with lipid rafts or whether this association is required for signaling or for translocation to uropods. Here, we found that loss of core 1-derived O-glycans in murine C1galt1-/- neutrophils blocked raft targeting of PSGL-1, CD43, and CD44, but not of other glycosylated proteins, as measured by resistance to solubilization in nonionic detergent and by copatching with a raft-resident sphingolipid on intact cells. Neuraminidase removal of sialic acids from wild-type neutrophils also blocked raft targeting. C1galt1-/- neutrophils or neuraminidase-treated neutrophils failed to activate tyrosine kinases when plated on immobilized anti-PSGL-1 or anti-CD44 F(ab')2. Furthermore, C1galt1-/- neutrophils incubated with anti-PSGL-1 F(ab')2 did not generate microparticles. In marked contrast, PSGL-1, CD43, and CD44 moved normally to the uropods of chemokine-stimulated C1galt1-/- neutrophils. These data define a role for core 1-derived O-glycans and terminal sialic acids in targeting glycoprotein ligands for selectins to lipid rafts of leukocytes. Preassociation of these glycoproteins with rafts is required for signaling but not for movement to uropods.
关键词：cell adhesion ; cell signaling ; inflammation ; uropod ; neutrophil