Alternative splicing of prosystemin pre-mRNA produces two isoforms that are active as signals in the wound response pathway

Systemin and its precursor protein, prosystemin, play an essential role in the systemic wound response pathway of tomato plants. We report here the isolation from tomato of a novel prosystemin cDNA (prosysB) that differs from the reported cDNA sequence (prosysA) by the addition of a CAG trinucleotide. Inspection of the prosystemin genomic sequence, which was mapped to the central region of chromosome 5, indicated that prosysA and prosysB transcripts are generated by an alternative splicing event that utilizes different 3' splice sites within intron 3. Quantitative RT-PCR analysis showed that prosysB transcripts accumulated to approximately twice the level of prosysA in all tissues that express the prosystemin gene. The relative abundance of the two mRNAs was unaffected by wounding or methyl jasmonate treatment, conditions that increase the level of total prosys mRNA. These findings indicate that alternative splicing of prosys pre-mRNA is a constitutive process. The amino acid sequence of prosysB is predicted to differ from that of prosysA by replacement of Arg-57 with Thr-Gly in the non-systemin portion of the protein. Over-expression of the prosysB cDNA in transgenic tomato plants conferred constitutive expression of defense genes that are regulated by wounding and systemin. We conclude that prosysB is the major prosystemin-encoding transcript in tomato, and that this isoform is active as a signal in the wound response pathway.