title: Mechanism of imidazolium ionic liquids toxicity in Saccharomyces cerevisiae and rational engineering of a tolerant, xylose-fermenting strain.
abstract: BACKGROUND
  Imidazolium ionic liquids (IILs) underpin promising technologies that generate fermentable sugars from lignocellulose for future biorefineries. However, residual IILs are toxic to fermentative microbes such as Saccharomyces cerevisiae, making IIL-tolerance a key property for strain engineering. To enable rational engineering, we used chemical genomic profiling to understand the effects of IILs on S. cerevisiae.
  
  
  RESULTS
  We found that IILs likely target mitochondria as their chemical genomic profiles closely resembled that of the mitochondrial membrane disrupting agent valinomycin. Further, several deletions of genes encoding mitochondrial proteins exhibited increased sensitivity to IIL. High-throughput chemical proteomics confirmed effects of IILs on mitochondrial protein levels. IILs induced abnormal mitochondrial morphology, as well as altered polarization of mitochondrial membrane potential similar to valinomycin. Deletion of the putative serine/threonine kinase PTK2 thought to activate the plasma-membrane proton efflux pump Pma1p conferred a significant IIL-fitness advantage. Conversely, overexpression of PMA1 conferred sensitivity to IILs, suggesting that hydrogen ion efflux may be coupled to influx of the toxic imidazolium cation. PTK2 deletion conferred resistance to multiple IILs, including [EMIM]Cl, [BMIM]Cl, and [EMIM]Ac. An engineered, xylose-converting ptk2∆ S. cerevisiae (Y133-IIL) strain consumed glucose and xylose faster and produced more ethanol in the presence of 1 % [BMIM]Cl than the wild-type PTK2 strain. We propose a model of IIL toxicity and resistance.
  
  
  CONCLUSIONS
  This work demonstrates the utility of chemical genomics-guided biodesign for development of superior microbial biocatalysts for the ever-changing landscape of fermentation inhibitors.
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. quinnd2222@gmail.com.
authors.forename: Quinn
authors.initials: Q
authors.lastname: Dickinson
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. scott.bottoms.1@gmail.com.
authors.forename: Scott
authors.initials: S
authors.lastname: Bottoms
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. lhinchman@glbrc.wisc.edu.
authors.forename: Li
authors.initials: L
authors.lastname: Hinchman
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. sean.mcilwain@wisc.edu.
authors.forename: Sean
authors.initials: S
authors.lastname: McIlwain
authors.affiliation: RIKEN Center for Sustainable Resource Science, Wako, Saitama, Japan. sheenaclaire.li@riken.jp.
authors.forename: Sheena
authors.initials: S
authors.lastname: Li
authors.affiliation: Department of Computer Science and Engineering, University of Minnesota-Twin Cities, Minneapolis, MN, USA. cmyers@cs.umn.edu.
authors.forename: Chad L
authors.initials: CL
authors.lastname: Myers
authors.affiliation: Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, Canada. charlie.boone@utoronto.ca.
authors.forename: Charles
authors.initials: C
authors.lastname: Boone
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. jcoon@chem.wisc.edu.|Biomolecular Chemistry, University of Wisconsin, Madison, WI, USA. jcoon@chem.wisc.edu.
authors.forename: Joshua J
authors.initials: JJ
authors.lastname: Coon
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. ahebert@wisc.edu.|Biomolecular Chemistry, University of Wisconsin, Madison, WI, USA. ahebert@wisc.edu.
authors.forename: Alexander
authors.initials: A
authors.lastname: Hebert
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. tksato@glbrc.wisc.edu.
authors.forename: Trey K
authors.initials: TK
authors.lastname: Sato
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. landick@biochem.wisc.edu.|Departments of Biochemistry and Bacteriology, University of Wisconsin, Madison, WI, USA. landick@biochem.wisc.edu.
authors.forename: Robert
authors.initials: R
authors.lastname: Landick
authors.affiliation: DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, Madison, WI, 53726, USA. jpiotrowski@wisc.edu.
authors.forename: Jeff S
authors.initials: JS
authors.lastname: Piotrowski
mesh_terms: D005285:Fermentation / Q000502:physiology; D052578:Ionic Liquids / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D014994:Xylose / Q000378:metabolism*
chemical_list: D052578:Ionic Liquids; D014994:Xylose