Vav1 and PI3K are required for phagocytosis of beta-glucan and subsequent superoxide generation by microglia.
ABSTRACT Microglia are the resident innate immune cells that are critical for innate and adaptive immune responses within the CNS. They recognize and are activated by pathogen-associated molecular patterns (PAMPs) present on the surface of pathogens. beta-glucans, the major PAMP present within fungal cell walls, are recognized by Dectin-1, which mediates numerous intracellular events invoked by beta-glucans in various immune cells. Previously, we showed that Dectin-1 mediates phagocytosis of beta-glucan and subsequent superoxide production in microglia. Here, we report that the guanine nucleotide exchange factor Vav1 as well as phosphoinositide-3 kinase (PI3K) are downstream mediators of what is now recognized as the Dectin-1 signaling pathway. Both Vav1 and PI3K are activated upon stimulation of microglia with beta-glucans, and the two proteins are required for phagocytosis of the glucan particles and for subsequent superoxide production. We also show that Vav1 functions upstream of PI3K and is required for activation of PI3K. Together, our results provide an important insight into the mechanistic aspects of microglial activation in response to beta-glucans.
SourceAvailable from: Karin Strijbis[Show abstract] [Hide abstract]
ABSTRACT: Phagocytosis of the opportunistic fungal pathogen Candida albicans by cells of the innate immune system is vital to prevent infection. Dectin-1 is the major phagocytic receptor involved in anti-fungal immunity. We identify two new interacting proteins of Dectin-1 in macrophages, Bruton's Tyrosine Kinase (BTK) and Vav1. BTK and Vav1 are recruited to phagocytic cups containing C. albicans yeasts or hyphae but are absent from mature phagosomes. BTK and Vav1 localize to cuff regions surrounding the hyphae, while Dectin-1 lines the full length of the phagosome. BTK and Vav1 colocalize with the lipid PI(3,4,5)P3 and F-actin at the phagocytic cup, but not with diacylglycerol (DAG) which marks more mature phagosomal membranes. Using a selective BTK inhibitor, we show that BTK contributes to DAG synthesis at the phagocytic cup and the subsequent recruitment of PKCε. BTK- or Vav1-deficient peritoneal macrophages display a defect in both zymosan and C. albicans phagocytosis. Bone marrow-derived macrophages that lack BTK or Vav1 show reduced uptake of C. albicans, comparable to Dectin1-deficient cells. BTK- or Vav1-deficient mice are more susceptible to systemic C. albicans infection than wild type mice. This work identifies an important role for BTK and Vav1 in immune responses against C. albicans.PLoS Pathogens 06/2013; 9(6):e1003446. DOI:10.1371/journal.ppat.1003446 · 8.06 Impact Factor
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ABSTRACT: β-glucans possess broad immunomodulatory properties, including activation of innate immune functions such as oxidative burst activity. The differential roles of complement receptor type 3 and Dectin-1, the known β-glucan receptors and their associated signaling pathways in the generation of oxidative burst induced by different physical forms of Saccharomyces cerevisiae-derived β-glucan were examined in human peripheral blood mononuclear cells. In this study whole glucan particle or immobilized soluble β-glucan was used to represent the phagocytizable or the non-phagocytizable form of a fungus, respectively. Oxidative burst as measured by the formation of superoxide was detected in peripheral blood mononuclear cells in response to whole glucan particle and immobilized soluble β-glucan. Superoxide induction with whole glucan particle was concluded to be Dectin-1-mediated, and required Src family kinases, phosphatidylinositol-3 kinase and protein kinase B/Akt. In contrast, the superoxide induction generated by immobilized soluble β-glucan was complement receptor type 3-mediated and required focal adhesion kinase, spleen tyrosine kinase, phosphatidylinositol-3 kinase, Akt, p38 mitogen activated protein kinase, phospholipase C and protein kinase C. The study results support the hypothesis that human peripheral blood mononuclear cells, specifically monocytes, utilize distinct receptors and overlapping, but distinct, signaling pathways for the oxidative burst in response to challenge by different physical forms of β-glucan.Glycobiology 01/2014; DOI:10.1093/glycob/cwu005 · 3.75 Impact Factor