Cytoskeletal reorganization driven by Rho GTPases plays a crucial role in the migration of T cells, which are key regulators of immunity. The molecular mechanisms that control actin cytoskeleton remodeling during T cell movement have only partially been clarified as the function of many modulators has not been evaluated in these cells. Here, we report a new function of RhoGDI2 by showing that this protein positively regulates Rho GTPase activation during T cell adhesion and migration. RhoGDI2 knockdown significantly reduced T cell adhesion and migration. Furthermore, RhoGDI2 knockdown decreased the activation of Rac1 and Cdc42, 2 members of Rho GTPases, and the remodeling of the actin cytoskeleton. Upon P‐selectin glycoprotein ligand‐1 engagement, RhoGDI2 was phosphorylated at Y24 and Y153 by kinases related to β2 integrin outside‐in signaling, Src, c‐Abl, and Syk, resulting in the accumulation of RhoGDI2 at the cell membrane. Subsequent phosphorylation of S31 induced the opening of RhoGDI2 and the release of Rho GTPases, whereas phosphorylation of Y153 might promote the activation of Rho GTPases by recruiting Vav1. Moreover, the disruption of lipid rafts with methyl‐β‐cyclodextrin blocked the interaction between integrins and RhoGDI2, reducing the level of phosphorylated RhoGDI2 and the activation of downstream Rho GTPases. Based on these observations, RhoGDI2 is a target of intergrin outside‐in signaling that activates Rho GTPases during T cell adhesion and migration, and RhoGDI2‐mediated signal transduction is based on the lipid rafts integrity.

P-selectin glycoprotein ligand-1 (PSGL-1) is involved in the initial step of lymphocyte homing by interacting with P- or E-selectins expressed on activated endothelium cells. Besides, it also functions as a receptor to induce signals that increase integrin affinity to ligands and mediate cell adhesion to endothelium. Integrin is required for the second step of lymphocyte homing, whose activation has been reported tightly regulated by inside-out signals triggered by chemokines or the shear-stress generated during lymphocyte rolling on endothelium. However, the relationship between PSGL-1-triggered signals and integrin activation is not clear. In this study, we demonstrated that PSGL-1 ligation induces β1 integrin-mediated adhesion to fibronectin via regulation of both β1 subunit clustering and conformation changes. Phosphoinositide 3-kinase (PI3K) is required for PSGL-1-induced β1 integrin clustering which ultimately regulates β1 integrin-mediated Jurkat cell adhesion to fibronectin. However, PI3K is not involved in the conformation changes or increases in the total expression of β1 integrin. Taken together, we found a novel signal pathway, PSGL-1-PI3K-β1 integrin, demonstrating the cooperation between initial adhesion and subsequent arrest and stable adhesion.

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.

Lipid rafts, a liquid-ordered plasma membrane microdomain, are related to cell-surface receptor function. PSGL-1, a major surface receptor protein for leukocyte, also acts as a signaling receptor in leukocyte rolling. To investigate the role of lipid raft in PSGL-1 signaling in human neutrophils, we quantitatively analyzed lipid raft proteome of human promyelocytic leukemia cell line HL-60 cells and identified a lipid raft-associated protein β-adducin. PSGL-1 ligation induced dissociation of the raft-associated protein β-adducin from lipid rafts and actin, as well as phosphorylation of β-adducin, indicating a transient uncoupling of lipid rafts from the actin cytoskeleton. Knockdown of β-adducin greatly attenuated HL-60 cells rolling on P-selectin. We also showed that Src kinase is crucial for PSGL-1 ligation-induced β-adducin phosphorylation and relocation. Taken together, these results show that β-adducin is a pivotal lipid raft-associated protein in PSGL-1-mediated neutrophil rolling on P-selectin.

In this study, we determined the complete mitogenome of Sphaeroma terebrans (Crustacea, Isopod, Sphaeromatidae), which was the first one in the family Sphaeromatidae. The mitogenome of S. terebrans was 15,540 bp in length and contained 13 protein-coding genes, 21 tRNA genes, 2 rRNA genes, 1 control region, and 2 unknown fragments which is longer than 200 bp. The overall base composition was 30.1% A, 14.9% C, 21.3% G, and 33.7% T. Six start codons (ATA, ATC, ATG, ATT, ACG, GTG) and three stop codons (TAA, TAG, T-) were found in the protein-coding genes. The size of 21 tRNA genes ranged from 51 to 66 bp, tRNA-Gly was not found. All nodes strongly supported that S. terebrans was placed as sister to Sphaeroma serratum in the maximum likelihood tree.