title: Erythrocyte scaffolding protein p55/MPP1 functions as an essential regulator of neutrophil polarity.
abstract: As mediators of innate immunity, neutrophils respond to chemoattractants by adopting a highly polarized morphology. Efficient chemotaxis requires the formation of one prominent pseudopod at the cell front characterized by actin polymerization, while local inhibition suppresses the formation of rear and lateral protrusions. This asymmetric control of signaling pathways is required for directional migration along a chemotactic gradient. Here, we identify the MAGUK protein p55/MPP1 as a mediator of the frontness signal required for neutrophil polarization. We developed a p55 knockout (p55(-/-)) mouse model, and demonstrate that p55(-/-) neutrophils form multiple transient pseudopods upon chemotactic stimulation, and do not migrate efficiently in vitro. Upon agonist stimulation, p55 is rapidly recruited to the leading edge of neutrophils in mice and humans. Total F-actin polymerization, along with Rac1 and RhoA activation, appear to be normal in p55(-/-) neutrophils. Importantly, phosphorylation of Akt is significantly decreased in p55(-/-) neutrophils upon chemotactic stimulation. The activity of immunoprecipitated phosphatidylinositol 3-kinase gamma (PI3Kgamma), responsible for chemoattractant-induced synthesis of PIP(3) and Akt phosphorylation, is unperturbed in p55(-/-) neutrophils. Although the total amount of PIP(3) is normal in p55(-/-) neutrophils, PIP(3) is diffusely localized and forms punctate aggregates in activated p55(-/-) neutrophils, as compared to its accumulation at the leading edge membrane in the wild type neutrophils. Together, these results show that p55 is required for neutrophil polarization by regulating Akt phosphorylation through a mechanism that is independent of PI3Kgamma activity.
authors.affiliation: Department of Pharmacology, University of Illinois College of Medicine, Chicago, IL 60612, USA.
authors.forename: Brendan J
authors.initials: BJ
authors.lastname: Quinn
authors.forename: Emily J
authors.initials: EJ
authors.lastname: Welch
authors.forename: Anthony C
authors.initials: AC
authors.lastname: Kim
authors.forename: Mary A
authors.initials: MA
authors.lastname: Lokuta
authors.forename: Anna
authors.initials: A
authors.lastname: Huttenlocher
authors.forename: Anwar A
authors.initials: AA
authors.lastname: Khan
authors.forename: Shafi M
authors.initials: SM
authors.lastname: Kuchay
authors.forename: Athar H
authors.initials: AH
authors.lastname: Chishti
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D016764:Cell Polarity; D002634:Chemotaxis, Leukocyte; D058544:Class Ib Phosphatidylinositol 3-Kinase; D053595:Embryonic Stem Cells / Q000166:cytology / Q000502:physiology; D004789:Enzyme Activation; D005260:Female; D020558:GTP Phosphohydrolases / Q000378:metabolism; D051528:Guanylate Kinases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009479:Neuropeptides / Q000378:metabolism; D009504:Neutrophils / Q000166:cytology / Q000378:metabolism; D010641:Phenotype; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011247:Pregnancy; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D033581:Stem Cell Transplantation; D020744:rac GTP-Binding Proteins / Q000378:metabolism; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: D000199:Actins; D007527:Isoenzymes; D009479:Neuropeptides; C484602:Rac1 protein, mouse; D015544:Inositol 1,4,5-Trisphosphate; D019869:Phosphatidylinositol 3-Kinases; D058544:Class Ib Phosphatidylinositol 3-Kinase; C546760:PIK3CG protein, human; C546764:Pik3cg protein, mouse; D051057:Proto-Oncogene Proteins c-akt; D051528:Guanylate Kinases; D020558:GTP Phosphohydrolases; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein