R. Rodríguez-Ortiz; C. Michielse; M. Rep; M.C. Limón; J. Avalos
Universiteit van Amsterdam
The phytopathogenic fungus Fusarium oxysporum is a model organism in the study of plant-fungus interactions. As other Fusarium species, illuminated cultures of F. oxysporum exhibit an orange pigmentation because of the synthesis of carotenoids, and its genome contains orthologous light-regulated car genes for this biosynthetic pathway. By chemical mutagenesis, we obtained carotenoid overproducing mutants of F. oxysporum, called carS, with upregulated mRNA levels of the car genes. To identify the regulatory gene responsible for this phenotype, a collection of T-DNA insertional mutants obtained by Agrobacterium mediated transformation was screened for carotenoid overproduction. Three candidate transformants exhibited a carS-like phenotype, and two of them contained T-DNA insertions in the same genomic region. The insertions did not affect the integrity of any annotated ORFs, but were linked to a gene coding for a putative RING-finger (RF) protein. Based on its similarity to the RF protein CrgA from the zygomycete Mucor circinelloides, whose mutation results in a similar carotenoid deregulation, this gene (FOXG_09307) was investigated in detail. Its expression was not affected in the transformants, but mutant alleles were found in several carS mutants. A strain carrying a partial FOXG_09307 deletion, fortuitously generated in a targeted transformation experiment, exhibited the carS phenotype. This mutant and a T-DNA insertional mutant holding a 5-bp insertion in FOXG_09307 were complemented with the wild type FOXG_09307 allele. We conclude that this gene is carS, encoding a RF protein involved in down-regulation of F. oxysporum carotenogenesis.