The interaction between keratinocytes (KC) and skin-resident immune cells plays an important role in induction of contact hypersensitivity (CHS). A specific subset of γδ T cells termed dendritic epidermal T cells (DETC) are located in mouse epidermis, and we have recently shown that DETC become activated and produce IL-17 in an IL-1β-dependent manner during CHS. Various receptors on DETC, including NKG2D, are involved in DETC responses against tumors and during wound healing. The ligands for NKG2D (NKG2DL) are stress-induced proteins such as Mult-1, H60, Rae-1 in mice and MICA, MICB and ULBP in humans. Here, we show that allergens up-regulate expression of the NKG2DL Mult-1, H60 and Rae-1 in cultured mouse KC and of MICA in primary human KC. We demonstrate that Mult-1 is expressed in mouse skin exposed to allergen. Furthermore, we find that the vast majority of DETC in murine epidermis and skin-homing cutaneous lymphocyte-associated antigen (CLA) positive γδ T cells in humans express NKG2D. Finally, we demonstrate that blocking of NKG2D partially inhibits allergen-induced DETC activation. These findings demonstrate that NKG2D and NKG2DL are involved in allergen-induced activation of DETC and indicate that the NKG2D/NKG2DL pathway might be a potential target for treatment of CHS.Journal of Investigative Dermatology accepted article preview online, 29 January 2015. doi:10.1038/jid.2015.23.