Sequence of a Nicotiana plumbaginifolia beta(1,3)-glucanase gene encoding a vacuolar isoform.

/3(l,3)-glucanases are a group of hormonally and developmentaUy regulated hydrolytic plant enzymes, which are also induced upon pathogen infection. A cDNA and genomic clone of the Nicotiana plumbaginifolia /3(l,3)-glucanase gnl gene have been analyzed previously (1, 2). Here, we report the nucleotide and deduced amino acid sequences of a different N. plumbaginifolia 0(1,3)glucanase gene designated gn2. The coding region of gn2 is contained within two exons and yields a precursor protein of 365 amino acids. In the absence of the corresponding cDNA, the length of the single intron present in gn2 was determined by primer extension and sequencing of the transcribed mRNA by the dideoxy procedure (3). A synthetic oligonucleotide complementary to nucleotides 16 to 56 downstream from the 5' end of the second exon was utilized for this experiment. Similarly, primer extension analysis using the same oligonucleotide defined the cap site (arrow in the figure) of the transcript derived from gnl at 14 base pairs (bp) upstream from the first ATG of the coding region. The sequences TATAAAT and CCAAT (underlined in the figure) likely corresponding to the gnl TATA and CAAT boxes, reside at positions 2 9 to -35 and 7 8 to -82 from the cap site, respectively. The N-terminal amino acid sequence of the GN2 protein has previously been determined (4) and the start of the mature GN2 was identified (boxed in figure). The nucleotide sequence data revealed that gnl encodes a precursor protein containing a 32-amino acid N-terminal signal peptide for translocation to the endoplasmic reticulum.