Human CYP2C8 is post-transcriptionally regulated by microRNAs 103 and 107 in human liver.
The CYP2C genes are extensively regulated at the transcriptional stage. The present study shows for the first time that CYP2Cs are also regulated post-transcriptionally by microRNAs (miRNAs). By using online search engines, we found potential miRNA response elements (MREs) in the 3'-untranslated region (3'-UTR) of the CYP2C mRNAs. Among these were a MRE for the miRNAs miR-103 and miR-107 in the 3'-UTR of human CYP2C8. CYP2C8 protein levels (measured through immunoblot analyses) did not correlate with CYP2C8 mRNA levels (measured through quantitative polymerase chain reaction analyses) in human liver samples. The translation efficiency (protein/mRNA ratio) for CYP2C8 was inversely correlated with the expression of miR-103 and miR-107. When three copies of the putative MRE from CYP2C8 were inserted downstream from a luciferase expression reporter, transfection with precursors for miR-103 or miR-107 decreased luciferase activity in primary hepatocytes, whereas transfection with antisense oligonucleotides (AsOs) for miR-103/miR-107 increased luciferase activity. As expected, there was no effect of the precursors or AsOs when three copies of the putative MRE were inserted in the reverse orientation. When precursors for miR-103/miR-107 were transfected into primary human hepatocytes, CYP2C8 protein levels were decreased, whereas AsOs increased CYP2C8 protein levels. Neither precursors nor AsOs affected CYP2C8 mRNA levels, which indicated that the effect was post-transcriptional. Putative MRE motifs were also found in the 3'-UTRs of CYP2C9 and CYP2C19, which suggested that the same miRNAs could regulate translation of other members of the CYP2C family, although to a lesser degree than CYP2C8. These results clearly show that CYP2Cs are regulated post-transcriptionally by miR-103 and miR-107.
Laboratory of Toxicology and Pharmacology, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA.
Shu-Yun
SY
Zhang
Sailesh
S
Surapureddi
Sherry
S
Coulter
Stephen S
SS
Ferguson
Joyce A
JA
Goldstein
D020413:3' Untranslated Regions; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism*; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D065727:Cytochrome P-450 CYP2C8; D015971:Gene Expression Regulation, Enzymologic; D056945:Hep G2 Cells; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000201:enzymology / Q000378:metabolism / Q000502:physiology; D006801:Humans; D008099:Liver / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D008156:Luciferases / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000235:genetics; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism; D014162:Transfection / Q000379:methods
D020413:3' Untranslated Regions; C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; C549045:MIRN107 microRNA, human; D035683:MicroRNAs; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D008156:Luciferases; D001189:Aryl Hydrocarbon Hydroxylases; C450259:CYP2C8 protein, human; D065727:Cytochrome P-450 CYP2C8