1451. Fungal Phagosome Maturation is Controlled by the -1,3-glucan receptor, Dectin-1
Session: Poster Abstract Session: Fungal Infections
Saturday, October 5, 2013
Room: The Moscone Center: Poster Hall C
Background: Elimination of fungal pathogens by phagocytes requires phagosome maturation, a process that involves the recruitment and fusion of intracellular proteins.  The role of Dectin-1, a ß-1,3-glucan receptor, critical for fungal recognition and triggering of Th17 responses, to phagosomal maturation has not been defined.

Methods: Phagosome maturation was tracked using live-cell confocal microscopy and phagosome isolation. A GFP-Dectin-1 fusion protein-expressing RAW macrophage cell line allowed visualization of Dectin-1 in different intracellular compartments.

Results: We show that GFP-Dectin-1 translocates to the fungal phagosome, but its signal decays after two hours. Inhibition of acidification results in retention of GFP-Dectin-1 to phagosome membranes highlighting the requirement for an acidic pH. Following ß-1,3-glucan recognition, GFP-Dectin-1 undergoes tyrosine phosphorylation by Src kinases with subsequent Syk activation. Our results demonstrate that Syk is activated independently of intraphagosomal pH. Inhibition of Src or Syk results in prolonged retention of GFP-Dectin-1 to the phagosome signifying a link between Syk and intraphagosomal pH. ß-1,3-glucan phagosomes expressing a signaling incompetent Dectin-1 failed to mature as demonstrated by prolonged Dectin-1 retention, presence of Rab5B, failure to acquire LAMP-1 and inability to acidify. Phagosomes containing C. albicans also require Dectin-1-dependent Syk activation for phagosomal maturation.

Conclusion: Taken together, these results support a model where Dectin-1 not only controls internalization of ß-1,3-glucan containing cargo and triggers proinflammatory cytokines, but also acts as a master regulator for subsequent phagolysosomal maturation through Syk activation.

Michael Mansour, M.D., Ph.D.1, Jenny Tam, Ph.D.2, Jennifer Reedy, MD, PhD2, Nida Khan, B.S.2, Srav Puranam2, Zeina Dagher, PhD2, David Sykes, MD, PhD3, Anna Sokolovska, PhD4, Christine Becker, PhD4, Antoine Tanne, PhD4, Lynda Stuart, MD, PhD4 and Jatin M. Vyas, M.D., Ph.D.2, (1)Massachusetts General Hospital, Division of Infectious Diseases, Boston, MA, (2)Massachsetts General Hospital, Division of Infectious Diseases, Boston, MA, (3)Division of Hematology and Oncology, Massachusetts General Hospital, Boston, MA, (4)Pediatrics, Massachusetts General Hosp, Boston, MA

Disclosures:

M. Mansour, None

J. Tam, None

J. Reedy, None

N. Khan, None

S. Puranam, None

Z. Dagher, None

D. Sykes, None

A. Sokolovska, None

C. Becker, None

A. Tanne, None

L. Stuart, None

J. M. Vyas, None

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