Expression and sequence requirements for nitrite reductase co-suppression

We have previously reported that the introduction of a full-length tobacco nitrite reductase Nii1 cDNA under the control of the 35S promoter triggers co-suppression of endogenous Nii genes in 25% of tobacco transformants. Her e we show that introduction of chimeric Nii1-uidA, uidA-Nii1 and Nii1-uidA- Nii1 transgenes carrying 186 bp of the 5' end and/or 241 bp of the 3' end o f the Nii1 cDNA do not trigger co-suppression of endogenous Nii genes. In a ddition, we show that when introduced by crossing or transformation into co -suppressed transgenic tobacco lines carrying full-length Nii1 transgenes, these chimeric transgenes are not silenced. These results therefore suggest that the 5' and 3' ends of the Nii1 cDNA are not sufficient to trigger co- suppression and are not targets for homology-dependent RNA degradation. Sur prisingly, co-suppression was released in a double transformant obtained by introduction of one of these constructs into the co-suppressed transgenic tobacco line 461-2.1 homozygous for a full-length Nii1 transgene, and in on e plant regenerated from untransformed leaf discs (plant 461-2.1*). The rea ppearance of co-suppression at very low frequency (less than 10(-3)) in the F-2 progeny of plant 461-2.1* and the apparent absence of structural modif ication of the transgene locus suggest a metastable epigenetic modification . The steady-state level of Nii mRNAs in the plant 461-2-.1* was higher tha n in wild-type plants but lower than in hemizygous plants 461-2.1 which nev er trigger silencing. These results therefore confirm that transcription of the transgene above a particular threshold is required to trigger co-suppr ession.