Gibberellin 2-oxidation and the SLN gene of Pisum sativum

Two cDNAs encoding gibberellin 2-oxidases were isolated from maturing pea seeds. The first, PsGA2ox1, was isolated by activity screening of a Lambda-ZAP cDNA library excised into phagemid form and expressed in Escherichia coli. The second, PsGA2ox2, was obtained initially as a PCR product using degenerate primers designed according to conserved regions of plant 2-oxoglutarate-dependent dioxygenases. E. coli heterologous expression products of PsGA2ox1 and PsGA2ox2 converted GA 1 to GA 8, as shown by HPLC-radiocounting, and gas chromatography-MS. PsGA2ox1 converted GA 20 to GA 29, but GA 20 was a poor substrate for the PsGA2ox2 expression product. Furthermore, PsGA2ox1 converted GA 29 to GA 29-catabolite at a low level of efficiency while PsGA2ox2 did not catalyse this step. A cDNA of PsGA2ox1 isolated from plants of genotype sin contained a single base deletion which was predicted to produce a truncated protein and gibberellin 2-oxidase activity could not be demonstrated from this cDNA. A 10 bp size difference between the introns of the SLN and sin PsGA2ox1 genes was used to show co-segregation between the SLN and sin phenotypes and the size of the PCR products. PsGA2ox1 transcripts were more abundant in cotyledons than in shoots, while the reverse was the case for PsGA2ox2. The expression patterns of the genes, together with the effects of the sin mutation, indicate that PsGA2ox1 plays a major role in GA 20 deactivation in both shoots and maturing seeds, while the PsGA2ox2 gene might be important for GA 1 deactivation in the shoot.