Abstract
Phagocytosis is a pivotal process by which macrophages eliminate microorganisms upon recognition by pathogen sensors. Surprisingly, the self-ligand cell surface receptor Slamf1 functions not only as a co-stimulatory molecule but also as a microbial sensor of several Gram-negative bacteria. Upon entering the phagosome of macrophages Slamf1 induces production of phosphatidylinositol 3-phosphate, which positively regulates the activity of the NOX2 enzyme and phagolysosomal maturation. Here, we report that in Escherichia coli-containing phagosomes of mouse macrophages, Slamf1 interacts with the class III PI3K Vps34 in a complex with Beclin-1 and UVRAG. Upon phagocytosis of bacteria the NOX2 activity was reduced in macrophages isolated from Beclin-1(+/-) mice compared with wild-type mice. This Slamf1/Beclin-1/Vps34/UVRAG protein complex is formed in intracellular membrane compartments as it is found without inducing phagocytosis in macrophages, human chronic lymphocytic leukemia cells, and transfectant HEK293 cells. Elimination of its cytoplasmic tail abolished the interaction of Slamf1 with the complex, but deletion or mutation of the two ITAM motifs did not. Both the BD and CCD domains of Beclin-1 were required for efficient binding to Slamf1. Because Slamf1 did not interact with Atg14L or Rubicon, which can also form a complex with Vps34 and Beclin-1, we conclude that Slamf1 recruits a subset of Vps34-associated proteins, which is involved in membrane fusion and NOX2 regulation.
Highlights
The receptor Slamf1 functions as a microbial sensor regulating phagocytosis
Because Slamf1 did not interact with Atg14L or Rubicon, which can form a complex with Vps34 and Beclin-1, we conclude that Slamf1 recruits a subset of Vps34-associated proteins, which is involved in membrane fusion and NOX2 regulation
Slamf1 Interacts with Vps34, Beclin-1, and UVRAG in Primary Murine Macrophages and E. coli-containing Phagosomes— As Slamf1 is involved in the regulation of phagocytosis by murine macrophages, to investigate this process in more detail, we assessed the interactions of Slamf1 with Vps34-associated proteins in these cells
Summary
The receptor Slamf functions as a microbial sensor regulating phagocytosis. Results: Slamf interacts with Vps34/Beclin-1/UVRAG complex in a wide range of cells. Because Slamf did not interact with Atg14L or Rubicon, which can form a complex with Vps and Beclin-1, we conclude that Slamf recruits a subset of Vps34-associated proteins, which is involved in membrane fusion and NOX2 regulation. Slamf Recruits a Beclin-1/Vps34/UVRAG Complex required for phagocytosis, no attempts have been made to assess the roles of the different Vps34-interacting proteins in this process It is interesting from cellular and evolutionary perspectives that autophagy and phagocytosis are ancient, highly conserved processes controlled by a similar mechanism. The results indicate that a complex containing Vps, Beclin-1, and UVRAG is important in Slamf1-modulated immune responses These results allow a better understanding of the functional specificity of the Vps34-associated proteins and suggest alternative innate immune regulation mechanisms (other than autophagy) for these proteins
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