Previously, we isolated the human AE2 (SLC4A2) gene, a member of the sodium-independent anion exchanger family, Rat ortholog of this gene was reported to drive alternative transcription yielding N-terminal variants of the AE2a message. We thus analyzed the human AE2 gene in this regard. Using HepG2 cells, two alternative first exons, each splicing to exon 3 in alternative transcripts, were found to be transcribed from overlapping sequences of intron 2. Exon 1b(1) corresponds to the rat variant "b" and encodes three initial residues (MTQ) in AE2b(1) isoform that replace the first 17 amino acids of AE2a protein, while the novel exon 1b(2) encodes eight initial residues (MDFLLRPQ) in AE2b(2) isoform, The relative abundance of AE2b(1) and AE2b(2) mRNAs was about 10% of AE2a mRNA each. Alternate promoter sequences have multiple potential binding motifs for liver-enriched factors, and dual-luciferase assays indicated that they possess the ability for driving transcription in transiently transfected HepG2 cells. Tissue survey showed that expression of human AE2b(1) and AE2b(2) transcripts is restricted to liver and kidney, while AE2a mRNA was encountered in all examined tissues, Our findings reveal a characteristic tissue-specific expression of two N-terminal variants of human AE2 from overlapping sequences within intron 2, one of which is a novel isoform,