Published in: Molecular and General Genetics - Abstract Volume 263 Issue 6 (2000) pp 995-1002:

"Both Sense and Antisense RNAs are Targets for the Sense Transgene-Induced Posttranscriptional Silencing Mechanism."

H. Van Houdt, M. Van Montagu, A. Depicker

Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie, Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium
E-mail: anna.depicker@rug.ac.be Tel.: +32-9-2645174; Fax: +32-9-2645349

Abstract:

Two stable transgenic tobacco lines were obtained as segregants from a primary transformant. Plants homozygous for a T-DNA inverted repeat locus (HOlo1) showed posttranscriptional gene silencing (PTGS) of the neomycin phosphotransferase II (nptII ) transgenes, whereas HOlo2 plants, homozygous for a single T-DNA insert, expressed the nptII genes normally. Transient expression of nptII genes newly introduced into leaves of both the HOlo2 and nptII-silenced HOlo1 plants was downregulated only in the silenced background. Different chimeric <beta>-glucuronidase (gus) genes with parts of the nptII transgene inserted in sense or antisense orientation into the 3'-untranslated region, which encoded transcripts that had homology or complementarity to nptII transcripts, showed reduced transient expression specifically in nptII-silenced tissue. Therefore, we conclude that RNAs of both polarities are targets for PTGS-induced RNA degradation, which supports the notion that double-stranded RNA acts as an inducing signal for silencing.

Acknowledgments:

The authors thank David Baulcombe and Jim English for performing the virus resistance assays, Isabelle Strobbe, Anni Jacobs, and Heidi Van Horebeek for technical assistance, Sylvie De Buck, Godelieve Gheysen, Frank Van Breusegem, Wout Boerjan, and Anton Gerats for critical reading of the manuscript and Martine De Cock for help preparing it. This work was supported by a grant from the European Union (BIOTECH BIO4-CT96-0253). The experiments were performed in compliance with the Belgian laws and regulations.