Signaling of chemotactic-receptor stimulated F-actin polymerization in the human neutrophil

The motility responses of the human neutrophil are essential in the body's defense against pathogens. Neutrophil motility is critically dependent on its ability to sense its environment and dynamically control its cellular actin cytoskeleton accordingly. The rearrangement of the cytoskeleton is associated with the temporary extension and stabilization of lamellipodia in the direction of movement. Lamella extension, and other migratory responses of the cell, is initiated by different membrane associated receptors that signal a variety of molecules grouped in signaling pathways. The receptor-generated signals eventually converge in the polymerization of F-actin, which is manifested as lamella formation and an overall rearrangement of the actin cytoskeleton. The precise mechanisms of the coupling and modulation of the signals generated from the different membrane receptors to the processes controlling cell migration are not yet entirely elucidated. However, it is known that the migration of the neutrophil is dependent on the p110 catalytic subunit of phosphoinositide 3-kinase γ. Here we present the first direct evidence for the existence of two distinct signaling pathways leading to F-actin polymerization in the pseudopod region: one pathway is dependent on the catalytic activity of the p110 subunit of the phosphoinositide 3-kinases and the other involves the regulatory p85α subunit. The p110-dependent signaling of F-actin polymerization also involves the activation of protein kinase C ζ and protein kinase B. The p85α-dependent pathway also includes the small GTPase RhoA, ROCK, Src family tyrosine kinases, NADPH and protein kinase A. These distinct pathways are chemotactic-receptor specific and utilize different isoforms of the phosphoinositide 3-kinases, some of which are differentially regulated by the growth factor, granulocyte-macrophage colony-stimulating factor, and insulin. Furthermore, in certain physiological conditions the p85α-dependent pathway may play a leading role in the polymerization of F-actin and overall cell migration. The results presented here were accomplished utilizing both single cell micropipet manipulation techniques and bulk biochemistry methods to characterize the motility responses of neutrophils, establishing links between the biochemical environment of the cell and the underlying signaling responses of activated neutrophils.