The aquaporin-4 (AQP4) gene encodes proteins of approximately 31 and 34 kDa with distinct N-terminal sequences. Both protein isoforms are expressed in brain, whereas mainly the smaller isoform is found in other tissues. We isolated the promoter sequences upstream of exon 0 (encoding the longer isoform) and exon 1 (encoding the shorter isoform). The exon 0 promoter region contained multiple TATA and CCAAT boxes and Sp1, AP1, and E-box elements; the exon 1 promoter lacked a CCAAT box but contained a TATA box and AP1, AP2, and E-box elements. Utilizing the CAT reporter assay, promoter activities were 7- to 9-fold greater for fragments from exon 1 vs exon 0 in human glioblastoma cells (SF-126, U-87 MG) and 11- to 13-fold greater in kidney cells (MDCK and LLC-PK1). Promoter deletion analysis indicated a strong suppressor element located just upstream of nucleotide -428 in the exon 0 promoter. RT-PCR analysis of human brain and kidney cDNAs confirmed much higher relative expression of transcript containing exon 0 vs exon 1 in brain. These results indicate differential transcriptional regulation of two AQP4 isoforms and suggest that tissue-specific factors regulate their relative expression.