title: An RNAi-based approach to down-regulate a gene family in vivo.
abstract: Genetic redundancy poses a major problem to the analysis of gene function. RNA interference allows the down-regulation of several genes simultaneously, offering the possibility to overcome genetic redundancy, something not easily achieved with traditional genetic approaches. Previously we have used a polycistronic miR155-based framework to knockdown expression of three genes of the early B cell factor family in cultured cells. Here we develop the system further by generating transgenic mice expressing the RNAi construct in vivo in an inducible manner. Expression of the transgene from the strong CAG promoter is compatible with a normal function of the basal miRNA/RNAi machinery, and the miR155 framework readily allows inducible expression from the Rosa26 locus as shown by Gfp. However, expression of the transgene in hematopoietic cells does not lead to changes in B cell development and neuronal expression does not affect cerebellar architecture as predicted from genetic deletion studies. Protein as well as mRNA levels generated from Ebf genes in hetero- and homozygous animals are comparable to wild-type levels. A likely explanation for the discrepancy in the effectiveness of the RNAi construct between cultured cells and transgenic animals lies in the efficiency of the sequences used, possibly together with the complexity of the transgene. Since new approaches allow to overcome efficiency problems of RNAi sequences, the data lay the foundation for future work on the simultaneous knockdown of several genes in vivo.
authors.affiliation: Institute of Clinical Molecular Biology and Tumor Genetics, Helmholtz Zentrum München, German Research Center for Environmental Health, Munich, Germany.
authors.forename: Jeehee
authors.initials: J
authors.lastname: Kim
authors.forename: Aurora
authors.initials: A
authors.lastname: Badaloni
authors.forename: Torsten
authors.initials: T
authors.lastname: Willert
authors.forename: Ursula
authors.initials: U
authors.lastname: Zimber-Strobl
authors.forename: Ralf
authors.initials: R
authors.lastname: Kühn
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Wurst
authors.forename: Matthias
authors.initials: M
authors.lastname: Kieslinger
mesh_terms: D000818:Animals; D002460:Cell Line; D002531:Cerebellum / Q000378:metabolism; D015536:Down-Regulation; D053595:Embryonic Stem Cells / Q000378:metabolism; D015870:Gene Expression; D005786:Gene Expression Regulation; D023061:Gene Order; D020868:Gene Silencing; D018390:Gene Targeting; D051379:Mice; D008822:Mice, Transgenic; D005810:Multigene Family; D054448:Precursor Cells, B-Lymphoid / Q000378:metabolism; D034622:RNA Interference; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D019076:Transgenes
chemical_list: C489921:Ebf1 protein, mouse; D015534:Trans-Activators