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.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.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.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.