We performed high-throughput in situ hybridization screening of sections of mouse epidermis using an equalized skin cDNA library as probes and identified a novel gene giving rise to two splicing variants, both of which are expressed in the spinous layer. This gene was mapped between two genes encoding keratinocyte-related peptides, suprabasin and keratinocyte differentiation-associated protein (Kdap), on human chromosome 19q13.1. These gene products appeared to carry functional signal sequences. We then designated these two splicing variants as dermokine-α and -β. Northern blotting and quantitative RT-PCR revealed that dermokine-α/-β, suprabasin, and Kdap were highly expressed in stratified epithelia. In mouse embryonic development, dermokine-α/-β began to be expressed during the period of stratification. Also, in differentiating primary cultured human keratinocytes, transcription of dermokine-α/-β, suprabasin, and Kdap was induced. These findings indicated that dermokine-α/-β, suprabasin, and Kdap are secreted from the spinous layer of the stratified epithelia and that these genes form a novel gene complex on the chromosome.