Differential suppression of the aryl hydrocarbon receptor nuclear translocator-dependent function by an aryl hydrocarbon receptor PAS-A-derived inhibitory molecule.
The aryl hydrocarbon receptor (AhR) heterodimerizes with the aryl hydrocarbon receptor nuclear translocator (Arnt) for transcriptional regulation. We generated three N-terminal deletion constructs of the human AhR of 12-24 kDa in size--namely D1, D2, and D3--to suppress the Arnt function. We observed that all three deletions interact with the human Arnt with similar affinities. D2, which contains part of the AhR PAS-A domain and interacts with the PAS-A domain of Arnt, inhibits the formation of the AhR gel shift complex. D2 suppresses the 3-methylcholanthrene-induced, dioxin response element (DRE)-driven luciferase activity in Hep3B cells and exogenous Arnt reverses this D2 suppression. D2 suppresses the induction of CYP1A1 at both the message and protein levels in Hep3B cells; however, the CYP1B1 induction is not affected. D2 suppresses the recruitment of Arnt to the cyp1a1 promoter but not to the cyp1b1 promoter, partly because the AhR/Arnt heterodimer binds better to the cyp1b1 DRE than to the cyp1a1 DRE. Interestingly, D2 has no effect on the cobalt chloride-induced, hypoxia inducible factor-1 (HIF-1)-dependent expression of vegf, aldolase c, and ldh-a messages. Our data reveal that the flanking sequences of the DRE contribute to the binding affinity of the AhR/Arnt heterodimer to its endogenous enhancers and the function of AhR and HIF-1 can be differentially suppressed by the D2 inhibitory molecule.
Department of Pharmaceutics and Medicinal Chemistry, Thomas J. Long School of Pharmacy and Health Sciences, University of the Pacific, Stockton, CA 95211, USA.
Jinghang
J
Xie
Department of Pharmacy, Qianfoshan Hospital of Shandong University, Jinan, Shandong, 250014, China.
Xin
X
Huang
Department of Pharmaceutics and Medicinal Chemistry, Thomas J. Long School of Pharmacy and Health Sciences, University of the Pacific, Stockton, CA 95211, USA.
Miki S
MS
Park
Department of Pharmaceutics and Medicinal Chemistry, Thomas J. Long School of Pharmacy and Health Sciences, University of the Pacific, Stockton, CA 95211, USA.
Hang M
HM
Pham
Department of Pharmaceutics and Medicinal Chemistry, Thomas J. Long School of Pharmacy and Health Sciences, University of the Pacific, Stockton, CA 95211, USA. Electronic address: wchan@pacific.edu.
William K
WK
Chan
D000595:Amino Acid Sequence; D000818:Animals; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator / Q000037:antagonists & inhibitors* / Q000502:physiology*; D045744:Cell Line, Tumor; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics* / Q000378:metabolism*
D018336:Receptors, Aryl Hydrocarbon; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; C116037:PAS domain kinases; D017346:Protein Serine-Threonine Kinases
AhR; AhR deletion constructs; Arnt; DRE binding; HIF-1