Analysis of the EBI/GeneBank database using nonhuman hair keratin associated protein (KAP) gene sequences as a query resulted in the identification of two human KAP gene domains on chromosome 21, one of which, located at 21q22.1, has recently been characterized. The second domain presented here, an approximately 90 kb domain on chromosome 21q23, harbored 16 KAP genes and two KAP pseudogenes. By comparison with known sheep and mouse KAP families, these genes could be assigned to two KAP families, KAP10 and KAP12, with the KAP10 family (12 members) being distinctly larger than the KAP12 family (four members). Systematic cDNA/3' rapid amplification of cDNA ends isolation studies using human scalp mRNA led to the identification of eight KAP10 and two KAP12 cDNA sequences. In situ hybridization analyses of human anagen hair follicles using specific 3'-noncoding sequences of the various KAP10/KAP12 genes revealed mRNA expression of nearly all KAP10 and KAP12 members exclusively in a narrow region of the middle portion of the hair fiber cuticle. Bioinformatic analyses of the promoter regions of the KAP10/KAP12 genes demonstrated several enhancer elements that were present in nearly all of the KAP genes. Primary among these were binding elements for the ETS, heat shock factor, AML, and HOX families of transcription factors.