Kinesin 5B Is Necessary for Delivery of Membrane and Receptors during FcγR-Mediated Phagocytosis

Abstract

FcγR-mediated phagocytosis is a cellular event that is evolutionary conserved to digest IgG-opsonized pathogens. Pseudopod formation during phagocytosis is a limiting step in managing the uptake of particles, and in this paper, we show that the conventional kinesin is involved in both receptor and membrane delivery to the phagocytic cup. Expression of a mutant kinesin isoform (GFP dominant negative mutant of kinesin H chain [EGFP-Kif5B-DN]) in RAW264.7 cells significantly reduced binding of IgG–sheep RBCs when macrophages were faced with multiple encounters with opsonized particles. Scanning electron microscopy analysis of EGFP-Kif5B-DN–expressing cells challenged with two rounds of IgG–sheep RBCs showed sparse, extremely thin pseudopods. We saw disrupted Rab11 trafficking to the phagocytic cup in EGFP-Kif5B-DN–transfected cells. Our particle overload assays also implicated phagosome membrane recycling in pseudopod formation. We observed reduced phagosome fission and trafficking in mutant kinesin-expressing cells, as well as reduced cell surface expression of FcγRs and Mac-1 receptors. In conclusion, anterograde trafficking via kinesin is essential for both receptor recycling from the phagosome and delivery of Rab11-containing membrane stores to effect broad and functional pseudopods during FcγR-mediated phagocytosis.

Classically Activated Macrophages Use Stable Microtubules for Matrix Metalloproteinase-9 (MMP-9) Secretion

“…More recently, we have implicated kinesin 5B in mediating vesicle targeting to active plasma membrane regions during the process of phagocytosis in macrophages (60). In the present study, we verified the enhanced MT stabilization and association of kinesin with MTs after macrophage activation and assessed their involvement in mediating the extensive trafficking of newly produced MMP-9 vesicles, required for macrophage migration (23, 28 -30).…”

“…10, A and B). Recently, we have shown that kinesin 5B is the major MT motor involved in vesicle targeting to active plasma membrane regions during the process of phagocytosis in macrophages (60). Having demonstrated a functional role for MTs in MMP-9 trafficking and an enhanced association of kinesin 5B FIGURE 8.…”

14-3-3γ meditated transport of plakoglobin to the cell border is required for the initiation of desmosome assembly in vitro and in vivo

“…Cells that had been transfected with either of the KIF3A constructs showed border staining for plakoglobin. Cells that had been transfected with either of the dominant-negative constructs had round edges and showed a morphology that was different from that of cells that had been transfected with either of the wild-type constructs, presumably because overexpression of the dominant-negative constructs affects other cellular processes, such as microtubule organization (Silver and Harrison, 2011). However, despite the change in morphology, only the cells that had been transfected with the dominant-negative KIF5B construct, and not those transfected with the dominant-negative KIF3A construct, showed a disruption of plakoglobin localization (Fig.…”

Rab31 and APPL2 enhance FcγR-mediated phagocytosis through PI3K/Akt signaling in macrophages

“…Rab35 recruits Arf6, Rac1, and Cdc42 for early actin assembly and phagocytic cup formation (Shim et al ., 2010; Egami et al ., 2011). Rab5, Rab11, and Rab7 mediate the fusion of early, recycling, and late endosomes, respectively, during phagosome maturation (Cox et al ., 2000; Murray et al ., 2005a; Silver and Harrison, 2011). Rab5 isoforms in particular have multiple roles, most notably in phagosome maturation and signaling by recruiting a series of effectors including VPS34, Hrs, EEA1, and phosphoinositide kinases and phosphatases (Fratti et al ., 2001; Vieira et al ., 2003; Kinchen et al ., 2008; Bohdanowicz et al ., 2012).…”