United States Department of Energy Joint Genome Institute, Walnut Creek, United States
Morgann C Reilly
Joint BioEnergy Institute, Emeryville, United States; Chemical and Biological Processes Development Group, Pacific Northwest National Laboratory, Richland, United States
Ya-Fang Cheng
Energy Biosciences Institute, Berkeley, United States
Stefan Bauer
Energy Biosciences Institute, Berkeley, United States
Igor V Grigoriev
United States Department of Energy Joint Genome Institute, Walnut Creek, United States; Department of Plant and Microbial Biology, University of California, Berkeley, Berkeley, United States; Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, United States
Joint BioEnergy Institute, Emeryville, United States
Blake A Simmons
Joint BioEnergy Institute, Emeryville, United States; Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, United States
Rachel B Brem
The Buck Institute for Research on Aging, Novato, United States; Department of Plant and Microbial Biology, University of California, Berkeley, Berkeley, United States
Energy Biosciences Institute, Berkeley, United States; Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, United States; Department of Bioengineering, University of California, Berkeley, Berkeley, United States
Energy Biosciences Institute, Berkeley, United States; Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, United States; Department of Bioengineering, University of California, Berkeley, Berkeley, United States
The basidiomycete yeast Rhodosporidium toruloides (also known as Rhodotorula toruloides) accumulates high concentrations of lipids and carotenoids from diverse carbon sources. It has great potential as a model for the cellular biology of lipid droplets and for sustainable chemical production. We developed a method for high-throughput genetics (RB-TDNAseq), using sequence-barcoded Agrobacterium tumefaciens T-DNA insertions. We identified 1,337 putative essential genes with low T-DNA insertion rates. We functionally profiled genes required for fatty acid catabolism and lipid accumulation, validating results with 35 targeted deletion strains. We identified a high-confidence set of 150 genes affecting lipid accumulation, including genes with predicted function in signaling cascades, gene expression, protein modification and vesicular trafficking, autophagy, amino acid synthesis and tRNA modification, and genes of unknown function. These results greatly advance our understanding of lipid metabolism in this oleaginous species and demonstrate a general approach for barcoded mutagenesis that should enable functional genomics in diverse fungi.
