title: HSP70 enhances macrophage phagocytosis by interaction with lipid raft-associated TLR-7 and upregulating p38 MAPK and PI3K pathways.
abstract: BACKGROUND
  Exogenous Heat Shock Protein-70 (HSP70), a product of necrotic cell death, binds the lipid raft microdomains of macrophages and, within minutes, stimulates the phagocytosis and presentation of internalized antigens. The aim of this study was to identify (a) the receptor on the lipid raft microdomain that interacts with HSP70 and (b) the subsequent signaling pathways that mediate HSP70-enhanced phagocytosis.
  
  
  METHODS
  Cells including RAW264.7, bone-marrow-derived macrophages from TLR7-/- mice or controls and cells subjected to genetic methods reducing the mRNA expression of TLR7 were used to examine the interaction of HSP70 with TLR7. The effect of HSP70-TLR7 interaction on phagocytosis was assessed using phagocytosis assays described earlier.
  
  
  RESULTS
  HSP70 binds Toll-like receptor-7 (TLR7) on the lipid raft microdomain of macrophage plasma membrane. Subsequent signaling is mediated through phosphoinositide 3-kinase (PI3K) and the up-regulation of the p38 MAP kinase pathways, both known activators of the phagocytic mechanisms. Reduced expression of TLR7 either via short interfering RNA for TLR7 (siRNA-TLR7) or using bone-marrow derived macrophages from TLR7-/- mice show that, as macrophages lose expression of TLR7, their ability to mediate HSP70-induced phagocytosis undergoes a corresponding diminution. Similarly, disruption of lipid rafts or blocking HSP70-TLR7-interaction or treatment with wortmannin and SB203580, inhibitors of PI3K or p38 MAPK, respectively, abrogates HSP70-induced macrophage phagocytosis.
  
  
  CONCLUSIONS
  The interaction of HSP70 and LR-associated TLR7, two phylogenetically conserved molecules, activates a rapid, membrane-bound signaling pathway that enhances phagocytosis, a vital innate defense mechanism. This study elucidates critical mechanistic elements that mediate HSP70-enhanced phagocytosis by macrophages.
authors.affiliation: Center for Immunotherapy, University of Connecticut School of Medicine, Farmington, Connecticut 06030-1601, USA.
authors.forename: Ruibo
authors.initials: R
authors.lastname: Wang
authors.forename: Terrence
authors.initials: T
authors.lastname: Town
authors.forename: Varun
authors.initials: V
authors.lastname: Gokarn
authors.forename: Richard A
authors.initials: RA
authors.lastname: Flavell
authors.forename: Rajiv Y
authors.initials: RY
authors.lastname: Chandawarkar
mesh_terms: D000818:Animals; D002460:Cell Line; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism* / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008817:Mice, Mutant Strains; D010587:Phagocytosis / Q000187:drug effects / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051199:Toll-Like Receptor 7 / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
chemical_list: D018840:HSP70 Heat-Shock Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C477038:Tlr7 protein, mouse; D051199:Toll-Like Receptor 7; D012313:RNA; D019869:Phosphatidylinositol 3-Kinases; D048051:p38 Mitogen-Activated Protein Kinases