The effect of sequences with high AU content on mRNA stability in tobacco.

Abstract

Little is known about the mechanisms that target transcripts for rapid degradation in plants. In mammalian cells, sequences with a high AU content and multiple AUUUA motifs have been shown to cause mRNA instability when present in the 3' untranslated regions of several transcripts. This precedent, coupled with the poor accumulation of AU-rich foreign transcripts in plants (e.g., BT-toxin mRNAs), prompted us to test whether AU sequences could destabilize transcripts in tobacco. To address this question, we made a set of constructs containing sequences with high AU content inserted into the 3' untranslated regions of reporter genes. The stability of the corresponding transcripts was then assayed in stably transformed cell lines of tobacco. These experiments showed that a 60-base sequence containing 11 copies of the AUUUA motif (AUUUA repeat) markedly destabilized a beta-glucuronidase reporter transcript compared to a no-insert control or a 60-base spacer sequence (GC control). Another sequence with an identical A+U content had little effect. The same results were obtained when each sequence was assayed within the 3' untranslated region of a beta-globin reporter transcript. In regenerated transgenic plants, the AUUUA repeat decreased the accumulation of the beta-globin transcript by approximately 14-fold, compared to the GC control. Taken together, our results indicate that the AUUUA repeat is recognized as an instability determinant in plant cells and that the effect is due to the sequence of the element, not simply to the high AU content.