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MicroRNA regulates the expression of human cytochrome P450 1B1
Yuki Tsuchiya1, Miki Nakajima, Shingo Takagi
1Drug Metabolism and Toxicology, Division of Pharmaceutical Sciences, Graduate School of Medical Science, Kanazawa University, Kanazawa, Japan.
Abstract:
MicroRNAs (miRNA) are small noncoding RNAs that regulate gene expression through translational repression or mRNA cleavage. Here, we found that cytochrome P450 (CYP), a superfamily of drug-metabolizing enzymes, is a target of miRNA. Human CYP1B1, which is highly expressed in estrogen target tissues, catalyzes the metabolic activation of various procarcinogens and the 4-hydroxylation of 17beta-estradiol. CYP1B1 protein is abundant in cancerous tissues. We identified a near-perfect matching sequence with miR-27b in the 3'-untranslated region of human CYP1B1. Luciferase assays revealed that the reporter activity of the plasmid containing the miR-27b recognition element was decreased in MCF-7 cells (miR-27 positive) but not in Jurkat cells (miR-27b negative). Exogenously expressed miR-27b could decrease the luciferase activity in Jurkat cells. In MCF-7 cells, the antisense oligoribonucleotide for miR-27b restored the luciferase activity and increased the protein level and enzymatic activity of endogenous CYP1B1. These results suggested that human CYP1B1 is post-transcriptionally regulated by miR-27b. The expression levels of miR-27b and CYP1B1 protein in breast cancerous and adjacent noncancerous tissues from 24 patients were evaluated. In most patients, the expression level of miR-27b was decreased in cancerous tissues, accompanied by a high level of CYP1B1 protein. A significant inverse association was observed between the expression levels of miR-27b and CYP1B1 protein. Thus, the decreased expression of miR-27b would be one of causes of the high expression of CYP1B1 protein in cancerous tissues. This is the first study to show that miRNAs regulate not only essential genes for physiologic events but also drug-metabolizing enzymes.
Insights
MicroRNAs (miRNAs) regulate gene expression. This study shows that miR-27b targets and downregulates cytochrome P450 1B1 (CYP1B1) protein, an enzyme often elevated in breast cancer tissues.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- MicroRNAs (miRNAs) are key regulators of gene expression, impacting cellular processes through translational repression or mRNA cleavage.
- Cytochrome P450 (CYP) enzymes, particularly CYP1B1, are involved in metabolizing drugs and hormones, and their dysregulation is linked to cancer.
- CYP1B1 is highly expressed in estrogen-target tissues and cancerous tissues, playing a role in activating procarcinogens and metabolizing 17beta-estradiol.
Purpose of the Study:
- To investigate the regulatory relationship between microRNAs and the drug-metabolizing enzyme Cytochrome P450 1B1 (CYP1B1).
- To determine if miR-27b post-transcriptionally regulates human CYP1B1 expression.
- To explore the potential role of miR-27b and CYP1B1 in breast cancer pathogenesis.
Main Methods:
- Bioinformatic analysis to identify potential miRNA binding sites on CYP1B1 mRNA.
- Luciferase reporter assays in cell lines (MCF-7 and Jurkat) to validate miR-27b binding and regulatory effects on CYP1B1.
- In vitro experiments using antisense oligoribonucleotides to modulate miR-27b levels and assess impacts on CYP1B1 protein and activity.
- Quantitative analysis of miR-27b and CYP1B1 protein expression in patient-derived breast cancerous and adjacent noncancerous tissues.
Main Results:
- A conserved binding site for miR-27b was identified in the 3'-untranslated region of human CYP1B1 mRNA.
- Luciferase assays confirmed that miR-27b directly targets and represses CYP1B1 expression in a sequence-dependent manner.
- Inhibition of miR-27b led to increased CYP1B1 protein levels and enzymatic activity in breast cancer cells.
- Patient tissue analysis revealed significantly lower miR-27b levels and higher CYP1B1 protein levels in cancerous tissues compared to adjacent noncancerous tissues.
- A significant inverse correlation was observed between miR-27b expression and CYP1B1 protein levels in breast cancer patients.
Conclusions:
- Human CYP1B1 is a direct post-transcriptional target of miR-27b.
- Decreased expression of miR-27b contributes to the overexpression of CYP1B1 protein in breast cancer tissues.
- This study establishes a novel miRNA-mediated regulatory pathway for drug-metabolizing enzymes, with implications for cancer biology and therapy.
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