Leukocyte transmigration is modulated by chemokine‐mediated PI3Kγ‐dependent phosphorylation of vimentin

Abstract

Phosphoinositide 3-kinase c (PI3Kc) plays a fundamental role in mediating leukocyte migration to inflammation sites. However, the downstream cytoplasmic events triggered by its signaling activity are still largely obscure. To address this issue, tyrosine and serine/threonine phosphorylated proteins of chemokine-stimulated WT or PI3Kc-null macrophages were investigated. Among the proteins analyzed, the intermediate filament vimentin was found as a downstream effector of the PI3Kc signaling pathway. Specific analysis of the phosphorylation state of vimentin in macrophages showed that this protein becomes rapidly phosphorylated in both tyrosine and serine residues upon chemokine stimulation. In the absence of PI3Kc or the kinase activity of PI3Kc (PI3Kc KD/KD ), phosphorylation of vimentin was reduced. PI3Kc-null macrophages displayed impaired chemokine-driven vimentin fiber disassembly as well as reduced ability to transmigrate across endothelial cells. While WT macrophages infected with a vimentin mutant resistant to N-terminal serine phosphorylation showed a reduction in transendothelial migration, infection of PI3Kc-null macrophages with a vimentin mutant mimicking serine phosphorylation of N-terminal residues rescued the transendothelial migration defect. These results define vimentin N-terminal phosphorylation and fiber reorganization as a target of chemokine-dependent PI3Kc signaling in leukocytes.Key words: Cell migration . Intermediate filaments . Phosphoinositide 3-kinases .Signal transduction IntroductionPhosphoinositide 3-kinases (PI3K) are a family of ubiquitously expressed lipid and protein kinases. They are activated downstream transmembrane receptors [1] and are involved in several cellular responses such as metabolic regulation, survival, proliferation, cell migration and adhesion. PI3K convert PtdIns(4,5)P 2 into PtdIns(3,4,5)P 3 , a second lipid messenger product forming a docking site for various proteins harboring lipid binding modules such as the pleckstrin homology domain [2].Ã These authors contributed equally to this work. 1136Class I PI3K are heterodimeric proteins composed of a p110 catalytic subunit (a, b, g and d) and a regulatory adaptor subunit. According to their mechanism of activation, these enzymes can be further classified into two subclasses: while class IA PI3K (PI3Ka, b and d) include an adaptor protein of the p85 family and are activated by tyrosine kinase receptors, the sole known element of class IB, PI3Kg, is specifically activated by G protein-coupled receptors (GPCR) [3]. This effect is regulated by the interaction of PI3Kg with Gbg subunits of active G proteins through the involvement of either the p101 adaptor or its homologue p84/p87 [4]. Among mammalian tissues, the highest level of PI3Kg expression is found in leukocytes where this enzyme directs the chemotactic response to GPCR agonists [3]. Indeed, PI3Kg-null granulocytes show a significant reduction in chemotaxis toward chemokines and severely impaired bacteria-elicited peritoneal recruitment [5,6]. This migrati...

The Role of Vimentin Intermediate Filaments in the Progression of Lung Cancer

“…More recently, IFs are being recognized as essential signaling proteins involved in key cancer biological functions. These include epithelial-to-mesenchymal transition (EMT), regulation of major signal transduction pathways, cell migration and invasion, the positioning and anchorage of organelles, such as mitochondria (1), and cell-cell and cell-substrate adhesion (2). The focus of this Review is the role played by vimentin, a type III IF protein, in the progression of lung cancer.…”

“…Moreover, vimentin phosphorylation was shown to enhance tumor and metastasis growth in vivo (20). Vimentin has also been reported to be a downstream target of PI3Kg signaling, the activation of which results in increased vimentin phosphorylation that is required for efficient cellular migration (1). Furthermore, protein kinase C z was shown to promote the interaction of IFs with 14-3-3 (21).…”

Sphingolipids inhibit vimentin-dependent cell migration

“…1E,F ;Nakamura et al, 2000). S6 has a role in motility and was also included (Barberis et al, 2009;Ivaska et al, 2005;Kim et al, 2010), although mass spectrometry did not identify it. These results demonstrated that S71 is the primary vimentin phosphorylation site induced by S1P and SPC.…”

“…In fact, phosphorylation-mediated disassembly of the vimentin network at the cell periphery is a prerequisite for actin-dependent lamellipodia formation and is necessary for lamellipodia stability (Helfand et al, 2011). Vimentin phosphorylation is also important for integrin recycling and cell motility (Barberis et al, 2009;Ivaska et al, 2005) and might also regulate focal adhesion turnover (Svitkina et al, 1996) and integrin surface expression (Nieminen et al, 2006). Vimentin also functions as a signalling organiser.…”

Novel Phosphoinositide 3-Kinaseδ,γInhibitor: Potent Anti-Inflammatory Effects and Joint Protection in Models of Rheumatoid Arthritis

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