title: LPS induces phosphorylation of actin-regulatory proteins leading to actin reassembly and macrophage motility.
abstract: Upon bacterial infection lipopolysaccharide (LPS) induces migration of monocytes/macrophages to the invaded region and production of pro-inflammatory mediators. We examined mechanisms of LPS-stimulated motility and found that LPS at 100 ng/ml induced rapid elongation and ruffling of macrophage-like J774 cells. A wound-healing assay revealed that LPS also activated directed cell movement that was followed by TNF-α production. The CD14 and TLR4 receptors of LPS translocated to the leading lamella of polarized cells, where they transiently colocalized triggering local accumulation of actin filaments and phosphatidylinositol 4,5-bisphosphate. Fractionation of Triton X-100 cell lysates revealed that LPS induced polymerization of cytoskeletal actin filaments by 50%, which coincided with the peak of cell motility. This microfilament population appeared at the expense of short filaments composing the plasma membrane skeleton of unstimulated cells and actin monomers consisting prior to the LPS stimulation about 60% of cellular actin. Simultaneously with actin polymerization, LPS stimulated phosphorylation of two actin-regulatory proteins, paxillin on tyrosine 118 by 80% and N-WASP on serine 484/485 by 20%, and these events preceded activation of NF-κB. LPS-induced protein phosphorylation and reorganization of the actin cytoskeleton were inhibited by PP2, a drug affecting activity of tyrosine kinases of the Src family. The data indicate that paxillin and N-WASP are involved in the reorganization of actin cytoskeleton driving motility of LPS-stimulated cells. Disturbances of actin organization induced by cytochalasin D did not inhibit TNF-α production suggesting that LPS-induced cell motility is not required for TNF-α release.
authors: Department of Cell Biology, Nencki Institute of Experimental Biology, 02-093 Warsaw, Poland.
authors: Galyna
authors: G
authors: Kleveta
authors: Kinga
authors: K
authors: Borzęcka
authors: Mykola
authors: M
authors: Zdioruk
authors: Maciej
authors: M
authors: Czerkies
authors: Hanna
authors: H
authors: Kuberczyk
authors: Natalia
authors: N
authors: Sybirna
authors: Andrzej
authors: A
authors: Sobota
authors: Katarzyna
authors: K
authors: Kwiatkowska
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D015638:Cytochalasin D / Q000494:pharmacology; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D051419:Paxillin / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000096:biosynthesis; D010766:Phosphorylation; D011743:Pyrimidines / Q000494:pharmacology; D015398:Signal Transduction / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal / Q000378:metabolism
chemical_list: C412373:AG 1879; D000199:Actins; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D016328:NF-kappa B; D051419:Paxillin; D019269:Phosphatidylinositol 4,5-Diphosphate; C496062:Pxn protein, mouse; D011743:Pyrimidines; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; C495772:Wasl protein, mouse; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D015638:Cytochalasin D