Homeobox genes code for transcription factors and are arranged in clusters, named A, B, C and D, found on four separate chromosomes in vertebrates. They contain a homeobox DNA sequence which codes for the homeodomain, a region of amino acids responsible for the DNA binding exhibited by these proteins. During embryonic development, the homeobox genes are both spatially and temporally regulated. In teratocarcinoma cell cultures, homeobox genes are regulated by retinoic acid (RA). The cDNAs from the first gene in the human HOX A cluster, HOX A1 (1.6), were cloned and the nucleotide sequence of a full-length cDNA was determined. It is highly homologous to its murine counterpart. Another HOX A1 cDNA was cloned, corresponding to an alternatively spliced form. In vitro translation of the full-length cDNA clone gave rise to a protein of 36 kDa. In PA-1 human teratocarcinoma cells HOX A1 is the earliest HOX A gene to be expressed after treatment with RA. To test whether HOX A1 could function as a early regulator of other HOX A cluster genes, we cotransfected into PA-1 human teratocarcinoma cells sense and antisense HOX A1 cDNAs expressed from an SV40 promoter with a 5.4-kb RA-sensitive HOX A4 (1.4) promoter-cat reporter. We found no effect of HOX A1 on the HOX A4 promoter. However, cotransfection of HOX A5 (1.3) was able to inhibit the HOX A4 promoter activity.