Article

Beyond Secondary Structure: Primary-Sequence Determinants License Pri-miRNA Hairpins for Processing.

Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
Cell (Impact Factor: 33.12). 02/2013; 152(4):844-58. DOI: 10.1016/j.cell.2013.01.031
Source: PubMed

ABSTRACT To use microRNAs to downregulate mRNA targets, cells must first process these ∼22 nt RNAs from primary transcripts (pri-miRNAs). These transcripts form RNA hairpins important for processing, but additional determinants must distinguish pri-miRNAs from the many other hairpin-containing transcripts expressed in each cell. Illustrating the complexity of this recognition, we show that most Caenorhabditis elegans pri-miRNAs lack determinants required for processing in human cells. To find these determinants, we generated many variants of four human pri-miRNAs, sequenced millions that retained function, and compared them with the starting variants. Our results confirmed the importance of pairing in the stem and revealed three primary-sequence determinants, including an SRp20-binding motif (CNNC) found downstream of most pri-miRNA hairpins in bilaterian animals, but not in nematodes. Adding this and other determinants to C. elegans pri-miRNAs imparted efficient processing in human cells, thereby confirming the importance of primary-sequence determinants for distinguishing pri-miRNAs from other hairpin-containing transcripts.

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