Human potassium chloride cotransporter 1 (SLC12A4) promoter is regulated by AP-2 and contains a functional downstream promoter element

• Published in: Blood 2004-06, Vol.103 (11), p.4302-4309
• Main Authors: Zhou, Guo-Ping, Wong, Clara, Su, Robert, Crable, Scott C., Anderson, Kathleen P., Gallagher, Patrick G.
• Format: Article
• Language: English
• Subjects: 5' Untranslated Regions - genetics; Acetylation; Anemias. Hemoglobinopathies; Base Sequence; Biological and medical sciences; Carcinoma, Hepatocellular; Cell physiology; Chromatin; Cloning, Molecular; Diseases of red blood cells; DNA-Binding Proteins - metabolism; Erythroid Cells; Fundamental and applied biological sciences. Psychology; HeLa Cells; Hematologic and hematopoietic diseases; Humans; K Cl- Cotransporters; K562 Cells; Medical sciences; Membrane and intracellular transports; Molecular and cellular biology; Molecular Sequence Data; Precipitin Tests; Promoter Regions, Genetic - genetics; Sp1 Transcription Factor - metabolism; Symporters - genetics; Symporters - metabolism; Transcription Factor AP-2; Transcription Factors - metabolism; Transcription Initiation Site
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2003-01-0107

Most K-Cl cotransport in the erythrocyte is attributed to potassium chloride cotransporter 1 (KCC1). K-Cl cotransport is elevated in sickle erythrocytes, and the KCC1 gene has been proposed as a modifier gene in sickle cell disease. To provide insight into our understanding of the regulation of the human KCC1 gene, we mapped the 5′ end of the KCC1 cDNA, cloned the corresponding genomic DNA, and identified the KCC1 gene promoter. The core promoter lacks a TATA box and is composed of an initiator element (InR) and a downstream promoter element (DPE), a combination found primarily in Drosophila gene promoters and rarely observed in mammalian gene promoters. Mutational analyses demonstrated that both the InR and DPE sites were critical for full promoter activity. In vitro DNase I footprinting, electrophoretic mobility shift assays, and reporter gene assays identified functional AP-2 and Sp1 sites in this region. The KCC1 promoter was transactivated by forced expression of AP-2 in heterologous cells. Sequences encoding the InR, DPE, AP-2, and Sp1 sites were 100% conserved between human and murine KCC1 genes. In vivo studies using chromatin immunoprecipitation assays with antihistone H3 and antihistone H4 antibodies demonstrated hyperacetylation of this core promoter region. (Blood. 2004;103:4302-4309)