(466b) High-Pass Filtering and Noise Suppression in Intragenic miRNA-Mediated Host Regulation

Authors: 
Kang, T., The University of Texas at Dallas
Bleris, L., The University of Texas at Dallas
Li, Y., The University of Texas at Dallas
Ehrhardt, K., The University of Texas at Dallas
Singh, A., University of Delaware
Quarton, T., The University of Texas at Dallas
Nowak, C. M., The University of Texas at Dallas
MicroRNAs (miRNAs) are short non-coding RNA molecules that regulate gene expression post-transcriptionally by binding to target messenger RNAs (mRNAs). The majority of human miRNAs are intragenic, located within exons or introns of protein-coding sequence (host). Intriguingly, a percentage of intragenic miRNAs negatively regulate the host transcript forming an incoherent feedforward motif topology. Here, we study intragenic miRNA-mediated host gene regulation using a synthetic gene circuit stably integrated in a human kidneys within a genomic cell safe harbor locus. We engineered an artificial intron by separating the coding sequence of a reporter gene using splice sites and incorporated a synthetic miRNA within an artificial intron. When this intragenic miRNA is directed to inhibit the host gene transcript via engineered targets in its 3’UTR we observe a reduction in fluorescent reporter expression accompanied by output filtering and noise reduction. Specifically, the system operates as a high-pass filter with respect to promoter strength, allowing output above and attenuating below certain promoter strength, with the threshold being robust to promoter strength and measurement time. Additionally, we show that the intragenic miRNA host regulation yields reduced noise compared to a splicing-alone architecture (i.e. without 3’UTR targets). Our results provide a new insight to intragenic miRNA mediated host gene expression, with direct implications to gene therapy and synthetic biology applications.