Flotillins Interact with PSGL-1 in Neutrophils and, upon Stimulation, Rapidly Organize into Membrane Domains Subsequently Accumulating in the Uropod 英文参考文献.docVIP

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Flotillins Interact with PSGL-1 in Neutrophils and, upon Stimulation, Rapidly Organize into Membrane Domains Subsequently Accumulating in the Uropod 英文参考文献.doc

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Flotillins Interact with PSGL-1 in Neutrophils and, upon Stimulation, Rapidly Organize into Membrane Domains Subsequently Accumulating in the Uropod 英文参考文献

FlotillinsInteractwithPSGL-1inNeutrophilsand,upon Stimulation,RapidlyOrganizeintoMembraneDomains SubsequentlyAccumulatingintheUropod Je′re′mieRossy1,DominiqueSchlicht1¤,BrittaEngelhardt2,VerenaNiggli1* 1DepartmentofPathology,UniversityofBern,Bern,Switzerland,2TheodorKocherInstitute,UniversityofBern,Bern,Switzerland Abstract Background: Neutrophils polarize and migrate in response to chemokines. Different types of membrane microdomains (rafts)havebeenpostulatedtobepresentinrearandfrontofpolarizedleukocytesanddisruptionofraftsbycholesterol sequestration prevents leukocyte polarization. Reggie/flotillin-1 and -2 are two highly homologous proteins that are ubiquitouslyenrichedindetergentresistantmembranesandarethoughttoshapemembranemicrodomainsbyforming homo-andhetero-oligomers.Itwasthegoalofthisstudytoinvestigatedynamicmembranemicrodomainreorganization duringneutrophilactivation. Methodology/Principal Findings: We show now, using immunofluorescence staining and co-immunoprecipitation, that endogenous flotillin-1 and -2 colocalize and associate in resting spherical and polarized primary neutrophils. Flotillins redistributeveryearlyafterchemoattractantstimulation,andformdistinctcapsinmorethan90%oftheneutrophils.At later time points flotillins accumulate in the uropod of polarized cells. Chemotactic peptide-induced redistribution and cappingofflotillinsrequiresintegrityanddynamicsoftheactincytoskeleton,butdoesnotinvolveRho-kinasedependent signalingrelatedtoformationoftheuropod.Bothflotillinisoformsareinvolvedintheformationofthismembranedomain, asuropodlocationofexogenouslyexpressedflotillinsisdramaticallyenhancedbyco-overexpressionoftaggedflotillin-1 and-2indifferentiatedHL-60cellsascomparedtocellsexpressingonlyonetaggedisoform.Flotillin-1and-2associatewith P-selectin glycoprotein ligand 1 (PSGL-1) in resting and in stimulated neutrophils as shown by colocalization and co- immunoprecipitation. Neutrophils isolated from PSGL-1-deficient mice exhibit flotillin caps to the