Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Tuesday Speaker Abstracts

Single mRNA Counting in Single Cell Reveals Function of RNA Stem Structure in

Coupling Dicing and Gene Silencing

Hye Ran Koh

1,2

, Amirhossein Ghanbariniaki

Sua Myong

1,2,3

Department of Physics, University of Illinois, Urbana, IL, USA,

Institute for Genomic Biology,

University of Illinois, Urbana, IL, USA,

Biophysics Department, Johns Hopkins University,

Baltimore, MD, USA,

Center for Physics of Living Cells, University of Illinois, Urbana, IL,

USA.

Micro (mi)RNAs possess secondary structures encompassing a loop and a stem with multiple

mismatches, but how they contribute to dicing and subsequent gene silencing efficiency remains

unclear. Using single molecule fluorescence in situ hybridization, we demonstrate that the

number of nuclear mRNA remains unchanged while cytoplasmic mRNA undergoes silencing.

Dicing rate and silencing efficiency both increase as a function of miRNA loop length in a

correlated manner. In contrast, mismatches in a stem drastically diminishes silencing efficiency

without reducing the dicing rate. We show that such decoupling effect is not due to the miRNA

uptake, cellular dicing or RISC loading. We postulate that the mismatches in a stem perturbs the

hand-over of cleaved miRNA from Dicer to Argonaute. Our result implies that the stem structure

in cellular miRNAs is intended for suboptimal silencing efficiency.