Regulation of aquaporin 3 expression by magnesium ion.
For understanding the actions of magnesium formulations, magnesium oxide and magnesium sulfate as a constituent of antacid, in the gastrointestinal tract, the effect of magnesium ion on the water channel aquaporin 3 (AQP3) known to be permeable mainly to water and glycerol was investigated in Caco-2 cells. The mRNA and protein of aquaporin 3 were detected by real-time RT-PCR and Western blotting, respectively, and found to increase significantly after treatment with magnesium acetate. Inhibitors for signal transducers, MDL-12330A, H-89, U0126, and Ro 31-8220, were shown to repress the increase in expression of the mRNA. A luciferase reporter vector containing bp -1382 to -12 of the 5'-flanking region of the aquaporin 3 gene was constructed for a reporter gene assay. The luciferase activity in transfectants increased on treatment with magnesium acetate. Serial deletion constructs revealed two regions responsible for the magnesium ion-mediated activation, one between bps -404 and -190, and the other between bps -190 and -82. siRNA for the cAMP response element-binding protein (CREB) sequence located between bp -404 and -190 counteracted the magnesium ion-mediated activation of aquaporin 3 transcription. These results suggest that signal transducers, adenylyl cyclase, protein kinase A (PKA), mitogen-activated protein kinase 1/2 (MEK1/2), and mitogen- and stress-activated protein kinase 1 (MSK1), were involved in the signaling pathway for regulating transcription of the aquaporin 3 gene and CREB is one of the transcriptional regulators for aquaporin 3 gene expression mediated by magnesium ion.
Okahira
Masasi
M
Laboratory of Pharmaceutics, Department of Clinical Pharmaceutics, Gifu Pharmaceutical University, 5-6-1 Mitahora-higashi, Gifu, Gifu 502-8585, Japan.
Kubota
Masafumi
M
Iguchi
Kazuhiro
K
Usui
Shigeyuki
S
Hirano
Kazuyuki
K
D051400:Aquaporin 3 / Q000096:biosynthesis*; D015153:Blotting, Western; D018938:Caco-2 Cells; D002626:Chemistry, Pharmaceutical; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D005822:Genetic Vectors; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D017616:Magnesium Compounds / Q000494:pharmacology*; D016297:Mutagenesis, Site-Directed; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects
D017616:Magnesium Compounds; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D051400:Aquaporin 3; D008156:Luciferases; D008274:Magnesium