We previously cloned two distinct cDNA clones,
NGR1 and
NGR3, encoding S-like ribonucleases (RNases) induced by wounding and tobacco mosaic virus (TMV) infection, respectively, in
Nicotiana glutinosa leaves. To gain insight into the regulatory mechanism of the RNase genes, we analyzed nucleotide sequences of the genes
ngr1 (4.1 kbp) and
ngr3 (5.3 kbp), containing their structural genes as well as 5′-flanking regions. The
ngr1 gene is organized in three exons with two intervening introns, and
ngr3 has four exons interrupted by three introns. Primer extension analyses localized single transcription initiation sites at −32 and −99 upstream of the translation initiation codons ATG in the genes
ngr1 and
ngr3, respectively. The β-glucuronidase (GUS) reporter gene analysis with serial 5′-deletion mutants as well as a gel shift assay defined the wound-responsive region at residues −509 to −288 in gene
ngr1 and a TMV-responsive region at the residues −401 to −174 in
ngr3, respectively. Sequence search using PLACE and PlantCARE data bases showed that a wound-responsive element: the WUN-motif, occurs within the wound-responsive region in
ngr1, while
ngr3 contains several potential
cis-regulating elements, such as the elicitor responsiveness element: the W-box, a TMV responsive element: GT1, and the WUN-motif at positions between −401 and −174. These findings suggested that some of these
cis-elements may be involved in inducible expressions of
ngr1 and
ngr3. Furthermore, the gel shift assay suggested that the dissociation of protein factor(s) upon TMV-infection from the regulatory region may cause an inducible expression of
ngr3.
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