title: Reversible suppression of an essential gene in adult mice using transgenic RNA interference.
abstract: RNAi has revolutionized loss-of-function genetics by enabling sequence-specific suppression of virtually any gene. Furthermore, tetracycline response elements (TRE) can drive expression of short hairpin RNAs (shRNAs) for inducible and reversible target gene suppression. Here, we demonstrate the feasibility of transgenic inducible RNAi for suppression of essential genes. We set out to directly target cell proliferation by screening an RNAi library against DNA replication factors and identified multiple shRNAs against Replication Protein A, subunit 3 (RPA3). We generated transgenic mice with TRE-driven Rpa3 shRNAs whose expression enforced a reversible cell cycle arrest. In adult mice, the block in cell proliferation caused rapid atrophy of the intestinal epithelium which led to weight loss and lethality within 8-11 d of shRNA induction. Upon shRNA withdrawal, villus atrophy and weight loss were fully reversible. Thus, shRpa3 transgenic mice provide an interesting tool to study tissue maintenance and regeneration. Overall, we have established a robust system that serves the purpose of temperature-sensitive alleles in other model organisms, enabling inducible and reversible suppression of essential genes in a mammalian system.
authors.affiliation: Cold Spring Harbor Laboratory, Watson School of Biological Sciences, Cold Spring Harbor, NY 11724, USA.
authors.forename: Katherine
authors.initials: K
authors.lastname: McJunkin
authors.forename: Anthony
authors.initials: A
authors.lastname: Mazurek
authors.forename: Prem K
authors.initials: PK
authors.lastname: Premsrirut
authors.forename: Johannes
authors.initials: J
authors.lastname: Zuber
authors.forename: Lukas E
authors.initials: LE
authors.lastname: Dow
authors.forename: Janelle
authors.initials: J
authors.lastname: Simon
authors.forename: Bruce
authors.initials: B
authors.lastname: Stillman
authors.forename: Scott W
authors.initials: SW
authors.lastname: Lowe
mesh_terms: D000483:Alleles; D000818:Animals; D002453:Cell Cycle / Q000502:physiology*; D004261:DNA Replication / Q000502:physiology*; D005260:Female; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D034622:RNA Interference; D051716:Replication Protein A / Q000378:metabolism; D020218:Response Elements / Q000502:physiology*
chemical_list: D051716:Replication Protein A