Direct evidence for rapid degradation of Bacillus thuringiensis toxin mRNA as a cause of poor expression in plants

It is well established that the expression of Bacillus thusingiensis (B.t.) toxin genes in higher plants is severely limited at the mRNA, level, but the cause remains controversial. Elucidating whether mRNA accumulation is limited transcriptionally or posttranscriptionally could contribute to effective gene design as well as provide insights about endogenous plant gene-expression mechanisms. To resolve this controversy, we compared the expression of an A/U-rich wild-type crylA(c) gene and a G/C-rich synthetic crylA(c) B.t.-toxin gene under the control of identical 5' and 3' flanking sequences. Transcriptional activities of the genes were equal as determined by nuclear run-on transcription assays. In contrast, mRNA half-life measurements demonstrated directly that the wildtype transcript was markedly less stable than that encoded by the synthetic gene. Sequences that limit mRNA accumulation were located at more than one site within the coding region, and some appeared to be recognized in Arabidopsis but not in tobacco (Nicotiana tabacum). These results support previous observations that some A/U-rich sequences can contribute to mRNA instability in plants. Our studies further indicate that some of these sequences may be differentially recognized in tobacco cells and Arabidopsis.