Matthew K. Oki Individual Journal 15

Week 15 Individual Journal

Outline

Intro

Organisms undergo natural temperature cycles throughout a day and night cycle It is uncertain if organisms are able to adapt and acclimate within a 24 hour cycle, or if they deal with “continuous temperature shock” Saccharomyces cerevisiae used Optimal temperature 33 C Often used in sub-optimal temperatures in industrial applications Temperature ranges vary by strain “The aim of this study is to investigate the impact of diurnal temperature cycles (DTC) on S. cerevisiae and to assess the extent to which these responses can be predicted from steady-state analyses.”

“This system was recently used to specifically investigate the impact of temperature dynamics on yeast glycolysis, based on integrated modeling and experimental analysis of the in vivo kinetics of glycolytic enzymes”

Methods and Results

prototrophic haploid yeast strain Saccharomyces cerevisiae CEN.PK113-7D (MATa) (33, 34) was used in this study

Kept in an anaerobic controlled environment

Used sequential batch reactors instead of single batches

After three volume cycles, (not sure what this means) with no significant changes in biomass, a cycle began with formula T (°C) = 21  9 sin{[t (h)  6] /12}

“Samples were taken during the fifth and/or sixth temperature cycle. To minimize disturbance, sampling volumes did not exceed 5% of the reactor volume during a single temperature cycle, and intervals of at least 3 h were maintained between sampling points”

Spelled out analytical methods

Measured glucose consumption during DTC Determined the percentage of cells carrying a bud Cell cycle phase distribution analyzed “Samples for microarray analysis were taken during the fifth and sixth temperature cycles from two independent duplicate cultures” “Sample points from the fifth and sixth temperature cycles were combined”

“All genes with a P value of 0.002 were considered to be significantly changed (1,102 genes)”

“Steady-state chemostat cultures at constant temperatures of 12°C and 30°C (independent duplicate cultures at both temperatures) were sampled for microarray analysis”

“For each sampling point during DTC, an average expression profile was calculated based on the two biologically independent arrays.”

“The complete data set (17 arrays) was deposited at the Gene Expression Omnibus database (http://www .ncbi.nlm.nih.gov/geo) under accession number GSE55372.”

Results

CO2 production revealed a clear cyclic variation in fermentative activity CO2 decreased production as temperatures dropped “For both CO2 and residual glucose, the amplitude of this fluctuation decreased as yeast cells acclimated to the DTC and became steady and reproducible after three cycles” “residual glucose concentration was inversely correlated with temperature” in 5th and 6th cycle

“glucose concentrations decreased much faster when the temperature increased after passing the temperature minimum than they increased as the temperature minimum was approached. This asymmetry was also visible in the off-gas CO2 profile”
“concentrations of extracellular metabolites (ethanol, glycerol, lactate, succinate, and pyruvate) (Fig. 3D to G) were largely unaffected by the cyclic temperature variation, with the notable exception of acetate, the concentration of which rhythmically varied by circa 70%”
“transcriptome analysis was performed, covering six time points during the 24-h temperature cycle”
“microarray analysis revealed major reprogramming of gene expression”

1102 genes showed transcription changes

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Acknowledgements

• I would like to thank my partners, Mia Huddleston, Matthew R Allegretti, and Colin Wikholm, for the assistance on this weeks project both in the understanding of our paper in class and completion of the powerpoint outside of class
• I would also like to thank Kam D. Dahlquist, Ph.D. for providing the instructions and information for this assignment both in class and on this document: BIOL368/F16:Week 8.
• Even though I worked with the people noted above, this individual journal entry was completed by me and not copied from another source.
• Matthew K. Oki 00:25, 5 December 2016 (EST):

References

1. BIOL368/F16:Week 14
2. Hebly, M., de Ridder, D., de Hulster, E. A. F., de la Torre Cortes, P., Pronk, J. T., & Daran-Lapujade, P. (2014). Physiological and transcriptional responses of anaerobic chemostat cultures of Saccharomyces cerevisiae subjected to diurnal temperature cycles. Applied and Environmental Microbiology, 80(14), 4433-4449. doi: 10.1128/AEM.00785-14

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