Quantitative design of microRNA-responsive mRNA from expression data

Limiting expression to target cell types is a longstanding goal in gene therapy, which could be met by sensing endogenous microRNA. However, an unclear association between microRNA expression and activity currently hampers such an approach. Here, we probe this relationship by measuring the stability of synthetic microRNA-responsive 3'UTRs across 10 cell lines in a library format. By systematically addressing biases in microRNA expression data and confounding factors such as microRNA crosstalk, we demonstrate that a straightforward model can quantitatively predict reporter stability purely from expression data. We use this model to design constructs with previously unattainable response patterns across our cell lines. The rules we derive for microRNA expression data selection and processing should apply to microRNA-responsive devices for any environment with available expression data. Plasmid libraries containing reporters with microRNA target sequences in their 3'UTR were transiently transfected into either 4 (library 1) or 10 (library 2) cell lines for n=2 replicates. After two days, poly(A) RNA was extracted, reverse transcribed, and sequenced. The original plasmid libraries were also sequenced.