GATA factors constitute a family of transcriptional regulatory proteins expressed with distinct developmental and tissue-specific profiles and thought to regulate cell-restricted programs of gene expression. Here we describe the molecular cloning, chromosomal location, and transcription of the human GATA-6 gene. The GATA-6 cDNA encodes a predicted 449-amino-acid protein, which is highly conserved among vertebrates, and includes the two adjacent zinc-finger/basic domains characteristic of the GATA factor family. GATA-6 maps to human chromosome 18q11.1-q11.2 by fluorescence in situ hybridization. The gene is transcribed in a pattern overlapping that of GATA-4. Transcripts for both of these genes are prominent in heart, pancreas, and ovary, but only GATA-6 mRNA is found in lung and liver. GATA-6 transcripts are also detected in cultures of human and rat vascular smooth muscle cells (VSMCs). In VSMCs, GATA-6 transcripts are down- regulated when quiescent cultures are stimulated to proliferate in response to mitogen activation. These data demonstrate that GATA-6 is subject to both tissue-specific and mitogen-responsive regulatory signals. GATA-6 is a prime candidate for a gene that might regulate the differentiative state of VSMCs.