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Organelles promoting high level terpenoid biosynthesis in filamentous fungi.


EMSL Project ID
48096

Abstract

Terpenoid compounds are a promising source of renewable biofuels that have many favorable characteristics such as high energy content and compatibility with existing energy-use technologies. While most attention has been paid to terpenoids produced by plants, filamentous fungi also can naturally synthesize high levels of these compounds. However, there is a lack of knowledge of the cellular location of terpenoid biosynthetic pathways, the channeling of precursor molecules and intermediates, and the coordination of primary metabolism and terpenoid biosynthetic pathways that result in maximum terpenoid synthesis. Recently we have described terpenoid assembly organelles (TAO) in the fungus Fusarium graminearum where terpenoid biosynthetic enzymes are localized upon induction. HMG CoA reductase, catalyzing synthesis of the terpenoid precursor mevalonate, normally localizes to the ER, but under terpenoid induction also localizes to the TAO. This proposal seeks to further characterize the structure and content of these cellular terpenoid factories with the ultimate goal of using that information for improved terpenoid engineering and output. Specifically we propose to isolate fluorescently tagged TAO by flow cytometry and determine their protein and terpenoid content utilizing analytical resources at EMSL. The transcriptome of the fungus under terpenoid inducing and non-inducing conditions will be determined by RNAseq performed at JGI. The TAO proteome data will be used to determine whether all known F. graminearum terpenoid biosynthetic enzymes are located there and to detect the presence of other unknown proteins that may be important for terpenoid synthesis or TAO biogenesis. Proteome data also will be used to determine whether TAO-associated proteins and TAO themselves may be conserved among members of the fungal kingdom and to predict signals and pathways targeting proteins to the organelle. Metabolomic data of TAO would be used to confirm the organelle as the site of terpenoid biosynthesis and to detect potential terpenoid pathway intermediates. Comparison of the transcriptome of the fungus under terpenoid inducing and non-inducing conditions will be used to predict whether gene products essential for terpenoid synthesis are restricted to the TAO and whether transcript modification may account for dual cellular targeting of enzymes such as HMG CoA reductase. Knowledge gained by this research will be used to guide functional studies aimed at expression of heterologous terpenoids, maximizing terpenoid expression, and engineering terpenoid pathways in filamentous fungi.

Project Details

Project type
FICUS Research
Start Date
2013-10-01
End Date
2015-12-31
Status
Closed

Team

Principal Investigator

Harold Kistler
Institution
United States Department of Agriculture - Agricultural Research Service Cereal Disease Laboratory

Co-Investigator(s)

Claudia Schmidt-Dannert
Institution
University of Minnesota

Team Members

Scott Baker
Institution
Environmental Molecular Sciences Laboratory

Related Publications

Boenisch MJ, KL Broz, SO Purvine, WB Chrisler, CD Nicora, L Connolly, M Freitag, SE Baker, and HC Kistler. 2017. "Structural reorganization of the fungal endoplasmic reticulum upon induction of mycotoxin biosynthesis." Scientific Reports 7:Article No. 44296. doi:10.1038/srep44296