Polyadenylation (poly(A)) of eukaryotic mRNA is a critical step for gene 
expression. In plants, poly(A) signals leading to the formation of polyadenosine 
tails after mRNAs include the far upstream elements, the AAUAAA-like signals, 
and the mRNA cleavage sites for poly(A). Multiple AAUAAA signals leading to 
alternative polyadenosine formation have been found in many genes, but the 
effects of each AAUAAA signal on gene expression remain to be uncovered. A DNA 
fragment, whose transcript contains two canonical AAUAAA signals from the 
3'-untranslation region of endochitinase gene of tobacco (Nicotiana tabacum L. 
cv. W38), was mutated and constructed into the downstream of beta-glucuronidase 
(GUS) coding region. Transient expression of GUS gene from these constructs 
indicated that the distal AAUAAA signal from the stop codon was more important 
than the proximal one in stimulating gene expression. Also, the sequence rather 
than the distance between the stop codon and the AAUAAA signal region was 
critical for gene expression. Transgenic tobaccos with these constructs were 
also generated, and the position of the polyadenosine tail formation in this 
region was mapped. Results revealed that both AAUAAA signals were functional, 
and that polyadenosine tails of most transcripts were directed by the distal 
AAUAAA signal. Finally, the RNA stabilities of these variants in transgenic 
plants were measured. RNAs from the variants with the functional distal AAUAAA 
signal were more stable than those with the functional proximal one only. The 
possible secondary structure in this poly(A) signal region was predicted and 
discussed.