The terminal loop region controls microRNA processing by Drosha and Dicer

microRNAs are widely expressed, ∼22-nt-long regulatory RNAs. They are first transcribed as much longer primary transcripts, which then undergo a series of processing steps to yield the single-stranded, mature microRNAs, although the mechanisms are incompletely understood. Here, we show that the terminal loop region of human primary microRNA transcripts is an important determinant of microRNA biogenesis. Mutations that restrain the terminal loop region inhibit Drosha processing of primary microRNA transcripts as well as Dicer processing of precursor microRNA transcripts in vitro. The inhibition may result from lower enzyme turnover on the mutant transcripts. Consequently, the mutations reduce miRNA maturation in transfected human cells. We conclude that a flexible terminal loop region is critical for microRNA processing.