We have previously demonstrated that a short sequence element (L7ATE) within the proximal promoter of a Purkinje cell-specific gene, pcp-2(L7), is required for the normal pattern of expression of the gene in the cerebellum of transgenic mice. The presence of a series of TAAT sequence motifs in this element suggested its interaction with homeodomain proteins. To extend these observations, degenerate oligonucleotides were used to clone by reverse-transcriptase polymerase chain reaction members of the mouse Hox gene family expressed in neonatal cerebellum but not forebrain. Two of these, HoxB7 and HoxA5, are continuously expressed from the neonatal period into adult stages in cerebellar Purkinje cells. These Hox proteins are shown to synergistically activate the L7 promoter by cotransfection assay in vitro. In contrast, another homeodomain protein that is normally expressed in Purkinje cells only during the embryonic period, En-2, has a negative effect on L7 gene expression. These data suggest a biphasic, combinatorial control mechanism for the Purkinje cell-specific expression of the pcp-2(L7) gene.