Proteomic analysis of murine skin has shown that a variety of heat shock proteins (HSPs) are constitutively expressed in the skin. Using murine allergic contact hypersensitivity as a model, we investigated the role of two heat shock proteins, HSP27 and HSP70, in the induction of cutaneous cell-mediated immune responses. Immunohistochemical examination of skin specimens showed that HSP27 was present in the epidermis and HSP70 was present in both the epidermis and dermis. Inhibition of HSP27 and HSP70 produced a reduction in the 1-fluoro-2,4-dinitrobenzene contact hypersensitivity response and resulted in the induction of Agspecific unresponsiveness. Treatment of dendritic cell cultures with recombinant HSP27 caused in the up-regulation of IL-1β, TNF-α, IL-6, IL-12p70, and IL-12p40 but not IL-23p19, which was inhibited when Abs to HSP27 were added. The 1-fluoro-2,4-dinitrobemzene-conjugated dendritic cells that had been treated with HSP27 had an increased capacity to initiate contact hypersensitivity responses compared with control dendritic cells. This augmented capacity required TLR4 signaling because neither cytokine production by dendritic cells nor the increased induction of contact hypersensitivity responses occurred in TLR4-deficient C3H/HeJ mice. Our findings indicate that a cascade of events occurs following initial interaction of hapten with the skin that includes increased activity of HSPs, their interaction with TLR4, and, in turn, increased production of cytokines that are known to enhance Ag presentation by T cells. The results suggest that HSPs form a link between adaptive and innate immunity during the early stages of contact hypersensitivity. Copyright © 2008 by The American Association of Immunologists, Inc.