Regulation of the Dlx3 homeobox gene upon differentiation of mouse keratinocytes.

The Distal-less Dlx3 homeodomain gene is expressed in terminally differentiated murine epidermal cells, and there is evidence to support an essential role as a transcriptional regulator of the terminal differentiation process in these cells. In an attempt to determine the factors that induce Dlx3 gene expression, we have cloned the 1.2-kilobase pair proximal region of murine gene and analyzed its cis-regulatory elements and potential trans-acting factors. The proximal region of the Dlx3 gene has a canonical TATA box and CCAAT box, and the transcription start site was located 205 base pairs upstream from the initiation of translation site. Serial deletion analysis showed that the region between -84 and -34 confers the maximal promoter activity both in undifferentiated and differentiated primary mouse keratinocytes. Gel retardation assays and mutational analysis demonstrated that the transcriptional regulator NF-Y (also referred to as CBF) binds to a CCAAT box motif within this region and is responsible for the majority of the Dlx3 promoter activity. In addition, an Sp1-binding site was located immediately upstream of transcription start site that acts as a positive regulatory element of the Dlx3 promoter, independent of the CCAAT box motif. Importantly, elements residing between +30 to +60 of the Dlx3 gene are responsible for the Ca(2+)-dependent induction of Dlx3 during keratinocyte differentiation.