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Objective: The KCl cotransporter (KCC) plays an important role in cellular cation and volume regulation and contributes to the process of volume reduction that accompanies reticulocyte maturation. In human red cells containing sickle hemoglobin, KCl cotransporter activity is high compared to normal cells, and contributes to the deleterious dehydration of sickle reticulocytes. To date, genes for four KCC isoforms have been identified. As a step toward determining which isoform(s) is responsible for the Cl-dependent K fluxes in reticulocytes, human erythroid cells were examined for the presence of various KCC isoform transcripts., Methods: In vitro differentiated erythroid precursors, and reticulocytes isolated from normal individuals and sickle patients, were examined by reverse-transcriptase PCR for the expression of KCC isoforms. Transient transfection experiments were subsequently performed to characterize a novel KCC1 promoter., Results: Expression of multiple isoforms was detected, with transcripts for KCC1, 3, and 4 detected in all samples of erythroid cells. Two N-terminal splicing variants were detected for both KCC1 and 3. Sickle hemoglobin containing reticulocytes demonstrated KCC isoform expression patterns similar to wild-type cells, except for a consistent difference in the relative abundance of one KCC1 splice variant. This N-terminal variant initiates from a newly described promoter in the KCC1 gene., Conclusion: Three KCC genes are expressed in human red cells. Splicing variants arising from the KCC1 and 3 genes are also evident. Structure/function studies of mouse KCC1 suggest that these natural variants could profoundly affect overall cotransporter activity in the red cell.