Rapid production of novel pre‐microRNA agent hsa‐mir‐27b in Escherichia coli using recombinant RNA technology for functional studies in mammalian cells

Abstract

MicroRNAs (miRNAs or miRs) are a large family of noncoding RNAs that have been identified as critical epigenetic factors in the regulation of various cellular processes including drug metabolism and disposition. However, the study on miRNA functions is limited to the use of synthetic RNA and recombinant DNA agents. Herein, we show that novel pre‐miRNA‐27b (mir‐27b) agents could be biosynthesized in Escherichia coli using tRNA‐based recombinant RNA technology, and recombinant tRNA/mir‐27b chimera was readily purified to a high degree of homogeneity (> 95%) using anion‐exchange fast protein liquid chromatography. The tRNA‐fusion mir‐27b was revealed to be processed to mature miRNA miR‐27b in human carcinoma LS‐180 cells in a dose and time dependent manner. Moreover, recombinant tRNA/mir‐27b agents were biologically active in reducing the mRNA and protein expression levels of cytochrome P450 3A4 (CYP3A4) which consequently led to a lower midazolam 1'‐hydroxylase activity. These findings demonstrate that human pre‐miRNAs may be produced in Escherichia coli by recombinant RNA technology as novel ncRNA agents for functional studies in drug metabolism.