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Temporal and fluoride control of secondary metabolism regulates cellular organofluorine biosynthesis.

ABSTRACT: Elucidating mechanisms of natural organofluorine biosynthesis is essential for a basic understanding of fluorine biochemistry in living systems as well as for expanding biological methods for fluorine incorporation into small molecules of interest. To meet this goal we have combined massively parallel sequencing technologies, genetic knockout, and in vitro biochemical approaches to investigate the fluoride response of the only known genetic host of an organofluorine-producing pathway, Streptomyces cattleya. Interestingly, we have discovered that the major mode of S. cattleya's resistance to the fluorinated toxin it produces, fluoroacetate, may be due to temporal control of production rather than the ability of the host's metabolic machinery to discriminate between fluorinated and non-fluorinated molecules. Indeed, neither the acetate kinase/phosphotransacetylase acetate assimilation pathway nor the TCA cycle enzymes (citrate synthase and aconitase) exclude fluorinated substrates based on in vitro biochemical characterization. Furthermore, disruption of the fluoroacetate resistance gene encoding a fluoroacetyl-CoA thioesterase (FlK) does not appear to lead to an observable growth defect related to organofluorine production. By showing that a switch in central metabolism can mediate and control molecular fluorine incorporation, our findings reveal a new potential strategy toward diversifying simple fluorinated building blocks into more complex products.

SUBMITTER: Walker MC

PROVIDER: S-EPMC5818262 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Temporal and fluoride control of secondary metabolism regulates cellular organofluorine biosynthesis.

Walker Mark C MC Wen Miao M Weeks Amy M AM Chang Michelle C Y MC

ACS chemical biology 20120706 9

Elucidating mechanisms of natural organofluorine biosynthesis is essential for a basic understanding of fluorine biochemistry in living systems as well as for expanding biological methods for fluorine incorporation into small molecules of interest. To meet this goal we have combined massively parallel sequencing technologies, genetic knockout, and in vitro biochemical approaches to investigate the fluoride response of the only known genetic host of an organofluorine-producing pathway, Streptomyc ...[more]

PMID: 22769062

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Project description:Fungi are rich sources of secondary metabolites of pharmaceutical importance, such as antibiotics, antitumor agents, and immunosuppressants, as well as of harmful toxins. Secondary metabolites play important roles in the development and pathogenesis of fungi. LaeA is a global regulator of secondary metabolism and was originally reported in Aspergillus nidulans; however, its role in secondary metabolism in Magnaporthe oryzae has not yet been reported. Here, we investigated the role of a gene homologous to LAEA (loss of AflR expression) of Aspergillus spp. in Magnaporthe oryzae, named M. oryzae LAEA (MoLAEA). Studies on MoLAEA overexpression and knockdown strains have suggested that this gene acts as a negative regulator of sporulation and melanin synthesis. However, it is not involved in the growth and pathogenesis of M. oryzae Transcriptomic data indicated that MoLAEA regulated genes involved in secondary metabolism. Interestingly, we observed (for the first time, to our knowledge) that this gene is involved in benzylpenicillin (penicillin G) synthesis in M. oryzae Overexpression of MoLAEA increased penicillin G production, whereas the silenced strain showed a complete absence of penicillin G compared to its presence in the wild type. We also observed that MoLaeA interacted with MoVeA, a velvet family protein involved in fungal development and secondary metabolism, in the nucleus. This study showed that though MoLAEA may not make any contribution in rice blast fungal pathogenesis, it regulates secondary metabolism in M. oryzae and thus can be further studied for identifying other new uncharacterized metabolites in this fungus.IMPORTANCE M. oryzae causes blast disease, the most serious disease of cultivated rice affecting global rice production. The genome of M. oryzae has been shown to have a number of genes involved in secondary metabolism, but most of them are uncharacterized. In fact, compared to studies of other filamentous fungi, hardly any work has been done on secondary metabolism in M. oryzae It is shown here (for the first time, to our knowledge) that penicillin G is being synthesized in M. oryzae and that MoLAEA is involved in this process. This is the first step in understanding the penicillin G biosynthesis pathway in M. oryzae This study also unraveled the details of how MoLaeA works by forming a nuclear complex with MoVeA in M. oryzae, thus indicating functional conservation of such a gene across filamentous fungi. All these findings open up avenues for more relevant investigations on the genetic regulation of secondary metabolism in M. oryzae.

Dataset Information

Temporal and fluoride control of secondary metabolism regulates cellular organofluorine biosynthesis.

Publications

Temporal and fluoride control of secondary metabolism regulates cellular organofluorine biosynthesis.

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