title: Directed migration of mouse macrophages in vitro involves myristoylated alanine-rich C-kinase substrate (MARCKS) protein.
abstract: A role for MARCKS protein in directed migration of macrophages toward a chemoattractant was investigated. A peptide identical to the N-terminus of MARCKS (the MANS peptide), shown previously to inhibit the function of MARCKS in various cell types, was used. We investigated whether this MARCKS-related peptide could affect migration of macrophages, using the mouse macrophage-like J774A.1 cell line and primary murine macrophages. Both of these cell types migrated in response to the chemoattractants macrophage/MCPs, MCP-1 (25-100 ng/ml) or C5a (5-20 ng/ml). Cells were preincubated (15 min) with MANS or a mis-sense control peptide (RNS), both at 50 μM, and effects on migration determined 3 h after addition of chemoattractants. The movement and interactions of MARCKS and actin also were followed visually via confocal microscopy using a fluorescently labeled antibody to MARCKS and fluorescently tagged phalloidin to identify actin. MANS, but not RNS, attenuated migration of J774A.1 cells and primary macrophages in response to MCP-1 or C5a, implicating MARCKS in the cellular mechanism of directed migration. Exposure of cells to MCP-1 resulted in rapid phosphorylation and translocation of MARCKS from plasma membrane to cytosol, whereas actin appeared to spread through the cell and into cell protrusions; there was visual and biochemical evidence of a transient interaction between MARCKS and actin during the process of migration. These results suggest that MARCKS is involved in directed migration of macrophages via a process involving its phosphorylation, cytoplasmic translocation, and interaction with actin.
authors.affiliation: Department of Molecular Biomedical Sciences, North Carolina State University, College of Veterinary Medicine, Raleigh, NC, USA.
authors.forename: Teresa D
authors.initials: TD
authors.lastname: Green
authors.forename: Joungjoa
authors.initials: J
authors.lastname: Park
authors.forename: Qi
authors.initials: Q
authors.lastname: Yin
authors.forename: Shijing
authors.initials: S
authors.lastname: Fang
authors.forename: Anne L
authors.initials: AL
authors.lastname: Crews
authors.forename: Samuel L
authors.initials: SL
authors.lastname: Jones
authors.forename: Kenneth B
authors.initials: KB
authors.lastname: Adler
mesh_terms: D008841:Actin Cytoskeleton / Q000276:immunology / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002634:Chemotaxis, Leukocyte / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins / Q000276:immunology / Q000378:metabolism*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D008565:Membrane Proteins / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D021381:Protein Transport / Q000502:physiology
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; C000619711:Marcks protein, mouse; D008565:Membrane Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate