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Imaging Target mRNA and siRNA‐Mediated Gene Silencing In Vivo with Ribozyme‐Based Reporters
Author(s) -
So MinKyung,
Gowrishankar Gayatri,
Hasegawa Sumitaka,
Chung JuneKey,
Rao Jianghong
Publication year - 2008
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.200800370
Subject(s) - ribozyme , gene silencing , rna splicing , biology , rna , microbiology and biotechnology , messenger rna , in vivo , reporter gene , gene expression , gene , genetics
Noninvasive imaging of specific mRNAs in living subjects promises numerous biological and medical applications. Common strategies use fluorescently or radioactively labelled antisense probes to detect target mRNAs through a hybridization mechanism, but have met with limited success in living animals. Here we present a novel molecular imaging approach based on the group I intron of Tetrahymena thermophila for imaging mRNA molecules in vivo. Engineered trans‐splicing ribozyme reporters contain three domains, each of which is designed for targeting, splicing, and reporting. They can transduce the target mRNA into a reporter mRNA, leading to the production of reporter enzymes that can be noninvasively imaged in vivo. We have demonstrated this ribozyme‐mediated RNA imaging method for imaging a mutant p53 mRNA both in single cells and noninvasively in living mice. After optimization, the ribozyme reporter increases contrast for the transiently expressed target by 180‐fold, and by ten‐fold for the stably expressed target. siRNA‐mediated specific gene silencing of p53 expression has been successfully imaged in real time in vivo. This new ribozyme‐based RNA reporter system should open up new avenues for in vivo RNA imaging and direct imaging of siRNA inhibition.