Lipid Raft Is Required for PSGL-1 Ligation Induced HL-60 Cell Adhesion on ICAM-1

Abstract

P-selectin glycoprotein ligand-1 (PSGL-1) and integrins are adhesion molecules that play critical roles in host defense and innate immunity. PSGL-1 mediates leukocyte rolling and primes leukocytes for integrin-mediated adhesion. However, the mechanism that PSGL-1 as a rolling receptor in regulating integrin activation has not been well characterized. Here, we investigate the function of lipid raft in regulating PSGL-1 induced β2 integrin-mediated HL-60 cells adhesion. PSGL-1 ligation with antibody enhances the β2 integrin activation and β2 integrin-dependent adhesion to ICAM-1. Importantly, with the treatment of methyl-β-cyclodextrin (MβCD), we confirm the role of lipid raft in regulating the activation of β2 integrin. Furthermore, we find that the protein level of PSGL-1 decreased in raft fractions in MβCD treated cells. PSGL-1 ligation induces the recruitment of spleen tyrosine kinase (Syk), a tyrosine kinase and Vav1 (the pivotal downstream effector of Syk signaling pathway involved in cytoskeleton regulation) to lipid raft. Inhibition of Syk activity with pharmacologic inhibitor strongly reduces HL-60 cells adhesion, implicating Syk is crucial for PSGL-1 mediated β2 integrin activation. Taken together, we report that ligation of PSGL-1 on HL-60 cells activates β2 integrin, for which lipid raft integrity and Syk activation are responsible. These findings have shed new light on the mechanisms that connect leukocyte initial rolling with subsequent adhesion.

RhoGDI2 positively regulates the Rho GTPases activation in response to the β2 outside-in signaling in T cells adhesion and migration on ICAM-1

“…Lipid rafts contain a variety of proteins and are a platform for signal transduction . As shown in our previous studies, the signals regulated by integrins also depend on lipid rafts . Due to the accumulation of RhoGDI2 at the membrane of T cells upon PSGL‐1 engagement or β2 integrin engagement, we presumed that the mechanisms regulating RhoGDI2 signaling rely on lipid rafts.…”

“…38 As shown in our previous studies, the signals regulated by integrins also depend on lipid rafts. 39 Due to the accumulation of RhoGDI2 at the membrane of T cells upon PSGL-1 engagement or 2 integrin engagement, we presumed that the mechanisms regulating RhoGDI2 signaling rely on lipid rafts. We disrupted the integrity of lipid rafts in Jurkat cells using M CD to verify this hypothesis.…”

Lipid Rafts Disruption Increases Ochratoxin A Cytotoxicity to Hepatocytes

“…In addition, lipid raft is reported to be closely related with cell adhesion. Lipid raft is required for HL‐60 cell adhesion . Wang et al.…”

“…In addition, lipid raft is reported to be closely related with cell adhesion. Lipid raft is required for HL-60 cell adhesion [16]. Wang et al reported that disrupting the integrity of lipid rafts inhibits focal adhesion disassembly, accompanied with alterations in cell morphology.…”

Mechanical regulation of calcium signaling of HL-60 on P-selectin under flow

“…Human leukaemic HL-60 cells, the neutrophilic promyelocytes, are widely used for studying their rolling behaviour on selectins [22, 24], signal transduction mechanisms [25, 26] and acute promyelocytic leukemia differentiation therapy [27, 28], because they can be easily got and modified by gene engineering [29]. In the present work, we examined intercellular calcium signaling of HL-60 on P-selectin with the parallel-plate flow chamber and fluorescence microscope techniques.…”