Circadian clock-controlled intestinal expression of the multidrug-resistance gene mdr1a in mice.
OBJECTIVE
P-glycoprotein, the product of the multidrug resistance (mdr) gene, functions as a xenobiotic transporter contributing to the intestinal barrier. Although intestinal expression of the mdr1a gene and its efflux pump function has been shown to exhibit 24-hour variation, the mechanism of the variations remains poorly understood. Here, we demonstrated that the molecular components of the circadian clock act as regulators to control 24-hour variation in the expression of the mdr1a gene.


METHODS
Luciferase reporter assay and gel mobility shift assay were used to study the mechanism of transcriptional regulation of the mdr1a gene by clock gene products. The messenger RNA levels and protein abundances in colon 26 cells and mouse intestine were measured by quantitative real-time polymerase chain reaction and Western blotting, respectively.


RESULTS
Hepatic leukemia factor (HLF) and E4 promoter binding protein-4 (E4BP4) regulated transcription of the mdr1a gene by competing with each other for the same DNA binding site. Molecular and biochemical analyses of HLF- and E4BP4-down-regulated colon 26 cells and the intestinal tract of Clock mutant mice suggested that these 2 proteins consisted of a reciprocating mechanism in which HLF activated the transcription of the mdr1a gene, whereas E4BP4 periodically suppressed transcription at the time of day when E4BP4 was abundant.


CONCLUSIONS
The intestinal expression of the mdr1a gene is influenced by the circadian organization of molecular clockwork. Our present findings provide a link between the circadian timekeeping system and xenobiotic detoxification.
Murakami
Yuichi
Y
Pharmaceutics, Division of Clinical Pharmacy, Department of Medico-Pharmaceutical Sciences, Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Higashi
Yuko
Y
Matsunaga
Naoya
N
Koyanagi
Satoru
S
Ohdo
Shigehiro
S
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000096:biosynthesis / Q000235:genetics*; D000230:Adenocarcinoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000818:Animals; D050976:Basic-Leucine Zipper Transcription Factors / Q000378:metabolism; D015153:Blotting, Western; D047369:Chromatin Immunoprecipitation; D002940:Circadian Rhythm / Q000502:physiology*; D003110:Colonic Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D018432:Drug Resistance, Multiple / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic; D007413:Intestinal Mucosa / Q000378:metabolism* / Q000473:pathology; D016350:Leucine Zippers; D051379:Mice; D008817:Mice, Mutant Strains; D016133:Polymerase Chain Reaction; D012334:RNA, Neoplasm / Q000235:genetics*
D018435:ATP Binding Cassette Transporter, Subfamily B; D050976:Basic-Leucine Zipper Transcription Factors; C490670:Hlf protein, mouse; C497441:Nfil3 protein, mouse; D012334:RNA, Neoplasm; C095122:multidrug resistance protein 3