Flotillins interact with PSGL-1 in neutrophils and, upon stimulation, rapidly organize into membrane domains subsequently accumulating in the uropod

Rossy, Jérémie; Schlicht, Dominique; Engelhardt, Britta; Niggli, Verena (2009). Flotillins interact with PSGL-1 in neutrophils and, upon stimulation, rapidly organize into membrane domains subsequently accumulating in the uropod. PLoS ONE, 4(4), e5403. Lawrence, Kans.: Public Library of Science 10.1371/journal.pone.0005403

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BACKGROUND: Neutrophils polarize and migrate in response to chemokines. Different types of membrane microdomains (rafts) have been postulated to be present in rear and front of polarized leukocytes and disruption of rafts by cholesterol sequestration prevents leukocyte polarization. Reggie/flotillin-1 and -2 are two highly homologous proteins that are ubiquitously enriched in detergent resistant membranes and are thought to shape membrane microdomains by forming homo- and hetero-oligomers. It was the goal of this study to investigate dynamic membrane microdomain reorganization during neutrophil activation. 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 redistribute very early after chemoattractant stimulation, and form distinct caps in more than 90% of the neutrophils. At later time points flotillins accumulate in the uropod of polarized cells. Chemotactic peptide-induced redistribution and capping of flotillins requires integrity and dynamics of the actin cytoskeleton, but does not involve Rho-kinase dependent signaling related to formation of the uropod. Both flotillin isoforms are involved in the formation of this membrane domain, as uropod location of exogenously expressed flotillins is dramatically enhanced by co-overexpression of tagged flotillin-1 and -2 in differentiated HL-60 cells as compared to cells expressing only one tagged isoform. Flotillin-1 and -2 associate with 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 same extent as cells isolated from wild type animals, implying that PSGL-1 is not required for the formation of the flotillin caps. Finally we show that stimulus-dependent redistribution of other uropod-located proteins, CD43 and ezrin/radixin/moesin, occurs much slower than that of flotillins and PSGL-1. CONCLUSIONS/SIGNIFICANCE: These results suggest that flotillin-rich actin-dependent membrane microdomains are importantly involved in neutrophil uropod formation and/or stabilization and organize uropod localization of PSGL-1.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Service Sector > Institute of Pathology
04 Faculty of Medicine > Service Sector > Institute of Pathology > Inflammatory Pathology
04 Faculty of Medicine > Pre-clinic Human Medicine > Theodor Kocher Institute

UniBE Contributor:

Rossy, Jérémie; Schlicht, Dominique Elisabeth; Engelhardt, Britta and Niggli, Verena

ISSN:

1932-6203

ISBN:

19404397

Publisher:

Public Library of Science

Language:

English

Submitter:

Factscience Import

Date Deposited:

04 Oct 2013 15:08

Last Modified:

16 Nov 2015 13:55

Publisher DOI:

10.1371/journal.pone.0005403

PubMed ID:

19404397

Web of Science ID:

000265614900014

BORIS DOI:

10.7892/boris.30007

URI:

https://boris.unibe.ch/id/eprint/30007 (FactScience: 165664)

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