Lkelly9 Week 10

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Definitions

1. In vivo: (Of a biological process) made to occur within the living organism
   • http://www.biology-online.org/dictionary/In_vivo
2. Diurnal: A biological rhythm that primarily express a periodicity during daylight hours
   • http://www.biology-online.org/dictionary/Diurnal_rhythm
3. Mannoproteins: yeast cell wall components that are proteins with large numbers of mannose groups attached; highly antigenic.
   • http://www.biology-online.org/dictionary/Mannoproteins
4. Transcriptome: all the RNAs present in a cell type
   • http://www.biology-online.org/articles/embryonic_stem_cells_prospects/expression_profiling_embryonic_stem.html
5. Motif: The smallest group of atoms in a polymer that, when under the influence of a rotation-translation operator, will assemble the rest of the atoms in the chain.
   • http://www.biology-online.org/dictionary/Motif
6. Biogenesis:(1) The process in which life forms arise from similar life forms., (2) It asserts that living things can only be produced by another living thing, and not by a non-living thing.
   • http://www.biology-online.org/dictionary/Biogenesis
7. Desaturase: Any of several enzymes that put double bonds into the hydrocarbon areas of fatty acids.
   • http://www.biology-online.org/dictionary/Desaturase
8. Prototrophic Strains: Strain's that have the same nutritional requirements as the wild-type strain.
   • http://www.biology-online.org/dictionary/Prototrophic_strains
9. Orthologues: Any gene that can be found in two or more different species that can be traced back to the same common ancestor.
   • http://www.biology-online.org/dictionary/Orthologous_Gene
10. Exogenous: Developed or originating outside the organism, as exogenous disease.
    • http://www.biology-online.org/dictionary/Exogenous

Outline

• What is the main result presented in this paper?
  • DNA microarray analysis showed that 494 genes in the glucose-limited cultures and 806 genes in the nitrogen-limited cultures had significantly different transcript levels at the two temperatures.
  • Other environmental parameters have a significant effect on the transcriptional response to a stimulus.
    • Cannot change one parameter without any impact on others.
  • The use of both glucose- and ammonium-limited cultures enabled the identification of core sets of genes that have a context-independent response to single environmental stimuli.
  • Growing evidence that the specific growth rate affects genome-wide transcription
  • Acclimatized growth at low temperatures does not involve a Msn2/Msn4-complex regulatory system. It is based on something else that has not been identified.
  • Identified a set of 235 genes that showed a consistent transcriptional response to low temperature, regardless of the limiting nutrient.
  • A group of genes involved in lipid metabolism was the only group that was clearly regulated in both low-temperature chemostats and batch cultures.
  • HSP26 and HSP42 were down regulated at low temperature in the chemostat culture and up regulated at low temperature in the batch cultures.
  • Study overall demonstrates that responses to low temperatures and low specific growth rate can be separated by using chemostat cultures.
• What is the importance or significance of this work?
  • This work pointed out important differences between batch cultures and chemostat cultures, such as the importance of discriminating different phases in the adaptation to environmental change. The chemostat cultures are able to control more variables.
  • It is also emphasized that low-temperature acclimation of yeast involves more than transcriptional reprogramming.
    • Intracellular metabolite levels are also adjusted.
• Create a flow chart to describe their methods.
  • 4 Different Growth Conditions
    • 12°C, glucose-limited
    • 12°C, ammonium-limited
    • 30°C, glucose-limited
    • 30°C, ammonium-limited
  • Utilized various analytical methods and microarray analysis and compared their data to other S. cerevisiae low-temperature transcriptome datasets.
• How did they treat the cells (what experiment were they doing?)
• What strain(s) of yeast did they use? Was the strain haploid or diploid?
  • Strain: prototrophic, haploid reference S. cerevisiae strain CEN.PK113-7D (MATa)
• What media did they grow them in? Under what conditions and temperatures?
  • 2.0 L chemostats, with a working volume of 1.0 L
    • dilution rate (D) = 0.03 h-1
    • pH kept at 5.0
    • Stirrer speed = 600 rpm
  • Grown at both 12°C and 30°C.
  • Defined synthetic medium limited by carbon or by nitrogen. All other growth requirements were in excess.
  • Anaerobic conditions
  • Before sampling, biomass dry weight, metabolites, dissolved oxygen, and gas profiles were constant for at least 3 volume changes.
• What controls did they use?
  • A combination of both glucose- and ammonium-limited cultures reduced the impact of secondary effects.
• How many replicates did they perform per condition?
  • In the microarray analysis, the results for each growth condition were derived from three independently cultured replicates.
  • It is unclear how many chemostats with each set of conditions were used.
• What mathematical/statistical method did they use to analyze the data?
  • Cells sampled from chemostats, preparation of probes, and hybridization to Affymetrix Genechip microarrays. (Precise methods described in Piper et al. (2002))
  • Agilent 2100 Bioanalyzer: determined RNA quality
  • Microsoft Excel running the significance analysis of microarrays add-in was used for pair-wise comparisons.
  • Expressionist Analyst version 3.2: Generated venn diagrams and heat-map visualizations of transcript data
  • Regulatory Sequence Analysis (RSA) Tools: performed promoter analysis
  • Database for Annotation, Visualization and Integrated Discovery (DAVID) 2006: Statistical assessment of over-representation of GO biological processes categories among sets of significantly changed transcripts
  • Fisher's test: overrepresentation of transcription-factor binding sites as defined by chromatin immunoprecipitation (ChIP)-on-chip analysis
    • Probability was calculated using the equation on the right.
• What transcription factors did they talk about?
• Briefly state the result shown in each of the figures and tables.

Acknowledgments

• Except for what is noted above, this individual journal entry was completed by me and not copied from another source.

Lauren M. Kelly 12:33, 26 March 2017 (EDT)