Rigidity of Circulating Lymphocytes Is Primarily Conferred by Vimentin Intermediate Filaments

Brown, Martin J.; Hallam, John A.; Colucci‐Guyon, Emma; Shaw, Stephen

doi:10.4049/jimmunol.166.11.6640

Cited by 140 publications

(123 citation statements)

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“…As N-terminal serine phosphorylation of vimentin is responsible for vimentin fiber disassembly and cytoskeletal remodeling [27,30,33], the effects of the lack of PI3Kg on vimentin polymerization dynamics were investigated by immunofluorescence. Disassembly of vimentin filaments was evident, in WT BMDM, 5 min after stimulation with CCL5 (Fig.…”

Section: Chemokines Stimulate the Pi3kc-dependent Serine Phosphorylatmentioning

confidence: 99%

“…Phosphorylation of vimentin Ser6, as well as its surrounding serines, and the subsequent filament disassembly play a crucial role in transendothelial cell migration [27,30,31]; therefore, the function of PI3Kg in this process was analyzed. In the presence of a CCL5 gradient, transmigration through TNF-a-activated HUVECs of PI3Kg-null BMDM was significantly impaired as compared with WT controls (46.1 and 32.4% reduction at 30 min and 4 h, respectively; po0.01, two-tailed Student's t test, n 5 3).…”

Section: Pi3kc-mediated Vimentin Phosphorylation Affects Transendothementioning

confidence: 99%

“…Nonetheless, fibroblasts from vimentindeficient mice show mechanical instability and impaired ability to migrate and to contract a 3-D collagen network [29], thus providing a genetic evidence for the involvement of vimentin in cell plasticity and motility [30,31]. More recent results further confirm this view and indicate that control of vimentin phosphorylation is crucial for cell migration [19,27].…”

mentioning

confidence: 98%

“…Similar results were also obtained with PI3Kg KD/KD BMDM ( Fig. 4D), thus demonstrating the involvement of PI3Kg catalytic activity in the process leading to vimentin serine phosphorylation.Chemokines stimulate PI3Kc-dependent vimentin structure disassemblyAs N-terminal serine phosphorylation of vimentin is responsible for vimentin fiber disassembly and cytoskeletal remodeling [27,30,33], the effects of the lack of PI3Kg on vimentin polymerization dynamics were investigated by immunofluorescence. Disassembly of vimentin filaments was evident, in WT BMDM, 5 min after stimulation with CCL5 (Fig.…”

mentioning

confidence: 99%

“…This PKC activation might potentially be controlled by GPCR-induced PI3K signaling, since the PI3K inhibitor LY294002 blocks bradykinin-induced translocation of PKCd and PKCe [41]. However, bisindolylmaleimide I-mediated blockade of PKC activity did not alter vimentin Ser6 phosphorylation kinetics in chemokinestimulated BMDM (data not shown), thus advocating further work to characterize the precise vimentin kinase involved downstream PI3Kg.Circulating lymphocytes show vimentin intermediate filaments forming a spherical cage that confers resistance to global cell deformation [30]. Collapse of this vimentin filament network is necessary to achieve the cell deformability required for transendothelial migration [30], thus suggesting that alteration of this process can reduce the efficiency of leukocyte extravasation.…”

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confidence: 99%

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

et al. 2009

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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...

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Section: Chemokines Stimulate the Pi3kc-dependent Serine Phosphorylatmentioning

confidence: 99%

Section: Pi3kc-mediated Vimentin Phosphorylation Affects Transendothementioning

confidence: 99%

mentioning

confidence: 98%

mentioning

confidence: 99%

mentioning

confidence: 99%

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

et al. 2009

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Identification of specific proteins in different lymphocyte populations by proteomic tools

Vuadens

Gasparini

Déon³

et al. 2002

PROTEOMICS

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The solubilized proteins of purified CD19(+) (B), CD8(+) (T) as well as CD4(+) (T) lymphocytes were separated by high resolution two-dimensional polyacrylamide gel electrophoresis, and the gels were analyzed using Melanie 3.0. Nine gels were studied, three for each lymphocyte population. After image analysis, 1411 +/- 73 spots (mean + SD) were detected. The protein pattern of B lymphocytes segregated from the one of T lymphocytes by ascendant heuristic clustering analysis. In addition, computer analysis separated CD8(+) from CD4(+) lymphocytes. When a search was performed in order to detect subsets of specific spots (presence vs. absence), a group of three spots, detected in the area of the protein maps corresponding to isoelectric point (pI) of 5.2 to 5.4 and molecular weight (M(r)) of 50 to 51 kDa, were found in both CD8(+) and CD4(+) cells, but not in CD19(+) cells. Mass spectrometry analysis revealed that these spots were associated with several proteins such as vimentin, tubulin, desmin and cytokeratin. Two spots, located in the area of the gel corresponding to pI of about 5.0 and a M(r) of 30 kDa, appeared as CD8(+) cell associated. Mass spectrometry analysis showed that the two spots were related to the same non-identified protein. Moreover internal peptides sequences matched with two human expressed sequence tags: gi/9759776, gi/12798420. No spots were found as only B cell associated.

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A Systematic Study of Size Correlation and Young's Modulus Sensitivity for Cellular Mechanical Phenotyping by Microfluidic Approaches

Liang

Zhong

2022

Adv Healthcare Materials

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Cellular mechanical properties are a class of intrinsic biophysical markers for cell state and health. Microfluidic mechanical phenotyping methods have emerged as promising tools to overcome the challenges of low throughput and high demand for manual skills in conventional approaches. In this work, two types of microfluidic cellular mechanical phenotyping methods, contactless hydro‐stretching deformability cytometry (lh‐DC) and contact constriction deformability cytometry (cc‐DC) are comprehensively studied and compared. Polymerized hydrogel beads with defined sizes are used to characterize a strong negative correlation between size and deformability in cc‐DC (r = −0.95), while lh‐DC presents a weak positive correlation (r = 0.13). Young's modulus sensitivity in cc‐DC is size‐dependent while it is a constant in lh‐DC. Moreover, the deformability assessment for human breast cell line mixture suggests the lh‐DC exhibits better differentiation capability of cells with different size distributions, while cc‐DC provides higher sensitivity to identify cellular mechanical changes within a single cell line. This work is the first to present a quantitative study and comparison of size correlation and Young's modulus sensitivity of contactless and contact microfluidic mechanical phenotyping methods, which provides guidance to choose the most suitable cellular mechanical phenotyping platform for specific cell analysis applications.

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Rigidity of Circulating Lymphocytes Is Primarily Conferred by Vimentin Intermediate Filaments

Cited by 140 publications

References 35 publications

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

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

Identification of specific proteins in different lymphocyte populations by proteomic tools

A Systematic Study of Size Correlation and Young's Modulus Sensitivity for Cellular Mechanical Phenotyping by Microfluidic Approaches

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