We have two projects. 1. What is the signal transduction pathway link chemokine GPCR signaling to the reorganization of actin cytoskeleton during neutrophil chemotaxis? Chemotaxis requires precisely coordinated polymerization and depolymerization of the actin cytoskeleton at leading fronts of migrating cells. However, GPCR activation-controlled F-actin depolymerization remains largely elusive. Recently, we identify slingshot 2 (SSH2), a phosphatase of cofilin (actin depolymerization factor), as a target of PKD1 that regulates cofilin phosphorylation and remodeling of the actin cytoskeleton during neutrophil chemotaxis (Xu et al. MBoC, 2015) 2. What is the role of beta-arrestin in chemokine GPCR-mediated neutrophil chemotaxis? β-arrestins have emerged as key regulators of cytoskeletal re-arrangement that are required for directed cell migration. While it is known that β-arrestins are required for formyl-Met-Leu-Phe receptor (FPR) recycling, less is known about their role in regulating FPR mediated neutrophil chemotaxis. Here, we show that β-arrestin1 (ArrB1) co-localized with F-actin in the leading edge of neutrophil-like HL-60 cells during chemotaxis and its knockdown resulted in markedly reduced migration within fMLP gradients. The small GTPase Rap2 was found to bind ArrB1 under resting conditions but dissociated upon fMLP stimulation. The FPR-dependent activation of Rap2 required ArrB1 but was independent of Gi activity. Significantly, depletion of either ArrB1 or Rap2 resulted in reduced chemotaxis and defects in cellular re-polarization within fMLP gradients. These data strongly suggest a model in which FPR is able to direct ArrB1 and other bound proteins that are required for lamillopodial extension to the leading edge in migrating neutrophils, thereby orientating and directing cell migration (Gera et al, to be submitted).
