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Am J Physiol Gastrointest Liver Physiol 286: G23-G30, 2004. First published August 14, 2003; doi:10.1152/ajpgi.00203.2003
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MUCOSAL BIOLOGY

Enterocyte differentiation marker intestinal alkaline phosphatase is a target gene of the gut-enriched Krüppel-like factor

Brian F. Hinnebusch,1,* Aleem Siddique,1,* J. Welles Henderson,1 Madhu S. Malo,1 Wenying Zhang,1 Christopher P. Athaide,1 Mario A. Abedrapo,1 Xinming Chen,2 Vincent W. Yang,2 and Richard A. Hodin1

1Department of Surgery, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts 02114; and 2Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322

Submitted 5 May 2003 ; accepted in final form 9 August 2003

We have examined the role that the transcription factor gut-enriched Krüppel-like factor (KLF4 or GKLF) plays in activating the enterocyte differentiation marker gene intestinal alkaline phosphatase (IAP). A yeast one-hybrid screen was used to identify proteins interacting with a previously identified cis-element (IF-III) located within the human IAP gene promoter. DNA-protein interactions were determined by using EMSA. Northern blot analysis was used to study RNA expression in human colon cancer RKO cells engineered to overexpress KLF4. Transient transfections with IAP-luciferase reporter constructs were used to characterize the mechanisms by which KLF4 activates IAP transcription. The yeast one-hybrid screen and EMSA identified KLF4 as binding to IF-III. RKO cells induced to overexpress KLF4 demonstrated a corresponding dose-dependent increase in IAP expression, and EMSA with nuclear extract from these cells confirmed that KLF4 binds to the IF-III element. Transient transfections revealed that KLF4 transactivated the IAP gene largely via a critical segment in the IAP promoter that includes the IF-III cis-element. Mutant KLF4 constructs failed to fully activate IAP. We have identified the enterocyte differentiation marker IAP as a KLF4 target gene. IAP transactivation by KLF4 is likely mediated through a critical region located within the proximal IAP promoter region.

intestine; transcription factor; gene regulation


Address for reprint requests and other correspondence: R. A. Hodin, Massachusetts General Hospital, Dept. of Surgery, Gray-Bigelow 504, 55 Fruit St., Boston, MA 02114 (E-mail: rhodin{at}partners.org).




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