Title

Regulation of directionality in fMLP-induced neutrophil chemotaxis

Date of Completion

January 2003

Keywords

Health Sciences, Immunology

Degree

Ph.D.

Abstract

Neutrophils play important roles in the innate immune system. Chemotaxis, which is a directional cellular movement up a chemoattractant gradient, is one of the major functions of neutrophils. The PI3 kinase signaling pathway and the downstream F-actin polarized polymerization were already known to control chemotaxis directionality. Recent findings suggested that p38 MAPK-MAPKAPK2 (abbreviated as “MK2”)-Lsp1 signaling pathway was important for controlling directionality, because p38 MAPK inhibitor SB203580 inhibited fMLP-induced neutrophil chemotaxis and MK2−/− mouse neutrophils showed a loss of directionality in Zigmond chambers containing the fMLP gradients. The major substrate of MK2 in neutrophils is Lsp1, which is an F-actin binding protein. Lsp1−/− mouse neutrophils also showed impairments in directionality control during chemotaxis. ^ In this study, the molecular mechanisms underlying how MK2 controls chemotaxis directionality were examined. (1) MK2 mediated Lsp1 phosphorylation is required for maintaining the F-actin polarization and stability in fmLP-induced neutrophil chemotaxis. MK2 deficiency causes the defects in maintaining the F-actin polarization, which then induces the impaired uropod localization of PI3 kinase signaling negative regulator PTEN. Some PTEN are localized to the front of migrating neutrophils and down regulate the PI3 kinase signaling. (2) In quiescent neutrophils, the basal MK2 serine/threonine kinase activity was implicated in regulating but not totally controlling adenylyl cyclase (AC) activity. MK2 deficiency causes the enhanced AC activity and the ensuing elevated cAMP level. Increased intracellular cAMP levels probably render the neutrophils to be in desensitized state, and further induce the directionality loss by PTEN dislocalization. (3) MK2 deficiency causes the decreased microtubule stability in both quiescent and chemotaxing neutrophils, and further polarization loss of these microtubules in chemotaxing neutrophils. Increased cAMP level may contribute to the disruption of microtubule stability. ^ There is a link between diabetes phenotype and neutrophil chemotaxis defects. Therefore, it will be of a great importance to study neutrophil chemotaxis on a diabetic background. Diabetic NOD mice were shown to harbor a mutation −4 amino acids away from the Src kinase LynA autophosphorylation site. In this study, the mutated Lyn in NOD neutrophils was shown to confer the increased Lyn kinase activity, which may have a link with the decreased PTEN uropod localization observed in these neutrophils during chemotaxis. ^