Gibberellin Biosynthetic Pathway inGibberella fujikuroi:Evidence for a Gene Cluster

Abstract

Differential screening of aGibberella fujikuroicDNA library was used to successfully clone and identify genes involved in the pathway of gibberellin biosynthesis. Several cDNA clones that hybridized preferentially to a cDNA probe prepared from mycelium induced for gibberellin production were isolated and characterized. The deduced amino acid sequences of two (identical) clones contained the conserved heme-binding motif of cytochrome P450 monooxygenases (FXXGXXXCXG). One of these cDNA fragments was used as a homologous probe for the screening of a genomic library. A hybridizing 6.7-kb genomicSalI fragment was cloned into pUC19. The sequencing of this clone revealed that a second cytochrome P450 monooxygenase gene was closely linked to the first one. Since at least four cytochrome P450 monooxygenase-catalyzed steps are involved in the synthesis of gibberellins, chromosome walking was performed to find a further gene of this family or other genes involved in gibberellin pathway. Next to the two P450 monooxygenase genes, a putative geranylgeranyl diphosphate synthase gene, the copalyl diphosphate synthase gene, which is the first specific gene of the gibberellin pathway, and a third P450 monooxygenase gene were identified. These results suggest that at least some of the genes involved in the biosynthesis of gibberellins are closely linked in a gene cluster inG. fujikuroi,as has been recently found for other “dispensable” pathways in fungi.