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 GA1 to GA8, as shown by HPLC-radiocounting, and gas chromatography-MS. PsGA2ox1 converted GA20 to GA29, but GA20 was a poor substrate for the PsGA2ox2 expression product. Furthermore, PsGA2ox1 converted GA29 to GA29-catabolite at a low level of efficiency while PsGA2ox2 did not catalyse this step. A cDNA of PsGA2ox1 isolated from plants of genotype sln 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 sln PsGA2ox1 genes was used to show co-segregation between the SLN and sln 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 sln mutation, indicate that PsGA2ox1 plays a major role in GA20 deactivation in both shoots and maturing seeds, while the PsGA2ox2 gene might be important for GA1 deactivation in the shoot.
Lester
D R
DR
School of Plant Science, University of Tasmania, Hobart, Australia.
Ross
J J
JJ
Smith
J J
JJ
Elliott
R C
RC
Reid
J B
JB
D000595:Amino Acid Sequence; D001483:Base Sequence; D001667:Binding, Competitive; D015152:Blotting, Northern; D017931:DNA Primers; D018076:DNA, Complementary; D017343:Genes, Plant; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D018532:Pisum sativum / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D018076:DNA, Complementary; D006899:Mixed Function Oxygenases; C121463:gibberellin 2-dioxygenase