The major gibberellin (GA) controlling stem elongation in pea (Pisum sativum L.) is GA 1 , which is formed from GA 20 by 3␤-hydroxylation. This step, which limits GA 1 biosynthesis in pea, is controlled by the Le locus, one of the original Mendelian loci. Mutations in this locus result in dwarfism. We have isolated cDNAs encoding a GA 3␤-hydroxylase from lines of pea carrying the Le, le, le-3, and le d alleles. The cDNA sequences from le and le-3 each contain a base substitution resulting in single amino acid changes relative to the sequence from Le. The cDNA sequence from le d , a mutant derived from an le line, contains both the le ''mutation'' and a single-base deletion, which causes a shift in reading frame and presumably a null mutation. cDNAs from each line were expressed in Escherichia coli. The expression product for the clone from Le converted GA 9 to GA 4 , and GA 20 to GA 1 , with K m values of 1.5 M and 13 M, respectively. The amino acid substitution in the clone from le increased K m for GA 9 100-fold and reduced conversion of GA 20 to almost nil. Expression products from le and le-3 possessed similar levels of 3␤-hydroxylase activity, and the expression product from le d was inactive. Our results suggest that the 3␤-hydroxylase cDNA is encoded by Le. Le transcript is expressed in roots, shoots, and cotyledons of germinating pea seedlings, in internodes and leaves of established seedlings, and in developing seeds.

The amount of active gibberellin (GA) in plant tissues is determined in part by its rate of catabolism through oxidation at C-2. In pea (Pisum sativum L.) seeds, GA 2-oxidation is controlled by the SLN (SLENDER) gene, a mutation of which produces seedlings characterized by a slender or hyper-elongated phenotype. We cloned a GA 2-oxidase cDNA from immature pea seeds by screening an expression library for enzyme activity. The clone contained a full-length open reading frame encoding a protein of 327 amino acids. Lysate of bacterial cultures expressing the protein converted the C 19 -GAs, GA 1 , GA 4 , GA 9 , and GA 20 to the corresponding 2␤-hydroxy products. GA 9 and GA 20 were also converted to GA 51 and GA 29 catabolites, respectively. The gene appeared to be one member of a small family of GA 2-oxidases in pea. Transcript was found predominantly in roots, flowers, young fruits, and testae of seeds. The corresponding transcript from sln pea contained a point mutation and did not produce active enzyme when expressed heterologously. RFLP analysis of a seedling population segregating for SLN and sln alleles showed the homozygous mutant allele co-segregating with the characteristic slender phenotype. We conclude that SLN encodes GA 2-oxidase.

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