Kim J H, Meng S, Shei A, Hodin R A
Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School and Harvard Digestive Diseases Center, Boston, Massachusetts 02215, USA.
Am J Physiol. 1999 Apr;276(4):G800-7. doi: 10.1152/ajpgi.1999.276.4.G800.
We have used sodium butyrate-treated HT-29 cells as an in vitro model system to study the molecular mechanisms underlying intestinal alkaline phosphatase (IAP) gene activation. Transient transfection assays using human IAP-CAT reporter genes along with DNase I footprinting were used to localize a critical cis element (IF-III) corresponding to the sequence 5'-GACTGGGCGGGGTCAAGATGGA-3'. Deletion of the IF-III element resulted in a dramatic reduction in reporter gene activity, and IF-III was shown to function in the context of a heterologous (SV40) promoter in a cell type-specific manner, further supporting its functional role in IAP transactivation. Electrophoretic mobility shift assays revealed that IF-III binds Sp1 and Sp3, but these factors comprise only a portion of the total nuclear binding and appear to mediate only a small portion of its transcriptional activity. IF-III does not correspond to any previously characterized regulatory region from other intestine-specific genes. We have thus identified a novel, Sp1-related cis-regulatory element in the human IAP gene that appears to play a role in its transcriptional activation during differentiation in vitro.
