Crabtree-positive yeasts

Report
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
 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  Competitive trait
Crabtree-positive: advantage of higher rate
• Structured environment  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:
 = 0 ×  
Population growth rate:
 =
Monod’s equation:
=
log( −0 )


 ×
 +
 =   while  =   +  
Monod, Herman et Cie (1946)
Population frequency to Population fitness
Population frequency:
 = 0 ×  
Population growth rate:
 =
In competition:
log( −0 )

 = 1 −  and
 = 1 − 
 =  − 
 = log
 −0

 = log
− log

0
×
0

 −0

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  Collection of picture
• Subtraction of light background  Isolation of
pixel’s colonies
Camera setting
Camera setting

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