Crabtree-positive yeasts
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Evolutionary Biology The Crabtree effect and its influence on fitness of yeast populations from natural isolates Presented by Emelyne Cunnington Co-supervised by Thomas Pfeiffer and Austen Ganley ATP ~ An energetic compound Energy metabolism The Crabtree effect Research questions Methodology Pilot study Conclusion ATP ~ As a component of yeast fitness Energy metabolism The Crabtree effect Glucose Research questions Glycolysis ATP Methodology Pilot study Conclusion RESPIRATION Pdh TCA Cycle OXPHOS CO2 Acetyl-CoA ATP Pyruvate FERMENTATION Pdc Acetaldehyde Adh Ethanol Crabtree effect Energy metabolism The Crabtree effect Research questions Methodology Pilot study ~Aerobic alcoholic fermentation~ • Certain yeasts produce ethanol in aerobic condition • Crabtree-positive yeasts • Ferment sugar in addition to respiration • In presence of oxygen • Above a critical [sugar] • By opposition: Crabtree-negative yeasts Conclusion • Use solely the respiration pathway Crabtree, Biochem. J. (1929) Crabtree effect Energy metabolism The Crabtree effect Research questions Methodology Pilot study ~Aerobic alcoholic fermentation~ • Crabtree-positive yeasts • Typical growth • On batch culture • Diauxic shift • Two time growth Conclusion Monje-Casas, Biochem, J. (2004) Crabtree effect Energy metabolism The Crabtree effect Research questions Methodology Pilot study ~Aerobic alcoholic fermentation~ • Crabtree-positive yeasts • Typical growth • On chemostat • Dilution rate • Drop in growth yield Conclusion Dijken, Antonie Van Leeuwenhoek (1993) Costs and Benefits Energy metabolism The Crabtree effect Research questions • Why do yeasts use the ‘wasteful’ fermentation pathway in presence of oxygen? • Rate of ATP production Methodology Pilot study • Yield in ATP production Conclusion Neijsel, Mol. Microbiol. (1994) Costs and Benefits Energy metabolism The Crabtree effect Research questions Methodology • Why do yeasts use the ‘wasteful’ fermentation pathway in presence of oxygen? • Aerobic fermentation = higher rate in ATP production • Competitive advantage • Come at the cost of lower yield of ATP production Pilot study ο Trade-off between yield and rate Conclusion Pfeiffer, Nature (2001) Costs and Benefits Energy metabolism The Crabtree effect • Yield/Rate Trade-off theory: • Growth rate at the cost of growth yield Research questions Selection for growth yield Coupled with decrease in growth rate Methodology Pilot study Conclusion Jasmin, Proc. R. Soc. B (2012) Investigation of the Crabtree effect Energy metabolism • Selection for growth yield The Crabtree effect Research questions Methodology Pilot study Conclusion Such as Environment dependant • Unstructured environment ο Competitive trait Crabtree-positive: advantage of higher rate • Structured environment ο Co-operative trait Crabtree-negative: advantage of higher yield MacLean, Nature (2006) Energy metabolism Experimental evolution ~Selection for growth yield~ The Crabtree effect Research questions Methodology Pilot study Conclusion Bachemann, PNAS (2013) Energy metabolism Distinction between natural isolates without engineered populations The Crabtree effect Research questions Methodology Pilot study Conclusion 72h growth Acknowledgement • • • • Thomas Pfeiffer Austen Ganley Rainey’s group Bas Teusink Thank you for your attention Energy metabolism The Crabtree effect Research questions PERSPECTIVES On the investigation of the Crabtree effect • Relevance of the Yield/Rate Trade-off theory • ‘Typical’ yeast metabolic behavior in nature Methodology • Influence of the Crabtree effect on population fitness Pilot study Conclusion • Influence of the population structures & interactions Yield in ATP production Respiration Fermentation Glycolysis Pyruvate oxidation (x2) Glycolysis Ethanol production ATP 2 -- 2 -- NADH 6 12 -- -- FADH2 -- 2 -- -- GTP -- 1 -- -- Total 8 15 (x2) 2 0 Pathway NADH = 3 ATP 38 - 2 FADH2 = 2ATP - GTP = ATP Aerobic Chemostat Culture Controlled culture in Bioreactor F R verduyn, Appl. Microbiol. Biotechnol. (1984) Ethanol production as a function of sugar concentration Costs and Benefits Energy metabolism The Crabtree effect • Why do yeasts use the ‘wasteful’ fermentation pathway in presence of oxygen? M.A.C Theory Glucose Research questions Glycolysis ATP RESPIRATION Methodology Pilot study Conclusion Pyruvate FERMENTATION Pdh Pdc TCA Cycle OXPHOS Adh Ald, Acs Acetaldehyde CO2 Acetyl-CoA ATP ATP Ethanol Yield/Rate trade-off in ATP production Pfeiffer, Trend in Biochemical Sciences (2005) Fitness as a function of sugar concentration Population frequency: ππ‘ = π0 × π ππ Population growth rate: ππππ₯ = Monod’s equation: π= log(ππ‘ −π0 ) π π ππππ₯ × πΎπ +π ππ = πΎπ π while ππΉ = πΎπ πΉ + πΎπ π Monod, Herman et Cie (1946) Population frequency to Population fitness Population frequency: ππ‘ = π0 × π ππ Population growth rate: ππ = In competition: log(ππ‘ −π0 ) π‘ ππ = 1 − ππ and ππ = 1 − ππ πππππ = ππ − ππ πππππ = log ππ‘ −π0 π‘ πππππ = log − log ππ‘ π0 × π0 ππ‘ ππ‘ −π0 π‘ Experimental design ~ Competition Day 1 Strain A Strain B Overnight culture Day 3 1ml 1ml 0h 0h 4h 4h 8h 8h 12h 12h Camera setting AxB T12 Strain A AxB T8 AxB T0 AxB T4 Strain B Energy metabolism Distinction between natural isolates without engineered populations The Crabtree effect Research questions Methodology Pilot study Conclusion 0h growth 72h growth • Using time lapse photography to record colony growth on agar plates ο Collection of picture • Subtraction of light background ο Isolation of pixel’s colonies Camera setting Camera setting
