Alex J. George Week 13

DNA Microarray Project

• After struggling for several minutes with GenMAPP and our data, we finally got our data uploaded to GenMAPP
• There were 70 errors while uploading
  • Here is our data saved from GenMAPP: Media:Leng Tai Yeast dataset.gex
• After uploading, we specified the Inreased and Decreased values for both Carbon and Nitrogen limiting cultures
  • Orange correlated to increase; Blue correlated to decreased
• From here, we used MAPPfinder to analyze our data
• The results of our data are found in the following .txt files:
  • Carbon-Limited Increased Expression- Media:Increasedcarbonlimited.txt
  • Carbon-Limited Decreased Expression- Media:Decreasedcarbonlimited.txt
  • Nitrogen-Limited Increased Expression- Media:Increasednitrogenlimited.txt
  • Nitrogen-Limited Decreased Expression- Media:Decreasednitrogenlimited.txt
• Here are the top 10 Gene Ontology terms for
  • Carbon-limited Decreased:

1. Mitochondrial electron transport chain
2. Energy Derivation by oxidation of organic compounds
3. DNA Transposition
4. Cellular Respiration
5. Aerobic Respiration
6. Amino acid permease activity
7. Carbohydrate metabolism
8. Cellular Carbohydrate metabolism
9. ATP synthesis coupled electron transport
10. Main Pathway of Carbohydrate Metabolism

• • Carbon-limited Increased:

1. Nucleolus
2. Cytoplasm organization and biogenesis
3. Ribosome biogenesis and assembly
4. Ribosome biogenesis
5. rRNA processing
6. rRNA Metabolism
7. Nuclear lumen
8. Membrane enclosed lumen
9. Organelle lumen
10. RNA Binding

• • Nitrogen-limited Decreased:

1. Disaccharide metabolism
2. Regulation of transcription from RNA polymerase to promoter
3. Phosphotransferase activity/ Alcohol group as acceptor
4. Trehalose metabolism
5. Kinase Activity
6. Transcription from RNA Polymerase to Promoter
7. Carbohydrate metabolism
8. Protein amino acid phosphorylation
9. Protein Kinase activity
10. Hydrolase activity/Hydrolyzing O-glycosyl compunds

• • Nitrogen-limited Increased:

1. Cytosolic Ribosome
2. Ribosome
3. Structural Constituent of Ribosome
4. Cytoplasm Organization and Biogenesis
5. Ribosome Biogenesis and assembly
6. Cytosolic Large Ribosomal Subunit
7. Ribonucleoprotein complex
8. Nucleolus
9. Ribosme Biogenesis
10. Large ribosomal subunit

• We searched for the gene "HXT2" and clicked on the "Cell Surface Receptor Linked Signal Transduction" to get the following map:

• We then clicked on the YCR072C gene to find its function which is a protein required for cell viability

Finding our data to work with

• We opened our data with excel in order to filter our results to find significant changes and GO terms
• The following is the criteria we used to cut the number of terms to about 20:
  • Permute p-value <0.05
  • Z score > 2
  • Number changed >3 and < 100
  • Percent Changed
    • Carbon-limited Increased: > 40%
    • Carbon-limited Decreased: > 60%
    • Nitrogen-Limited Increased: > 50%
    • Nitrogen-Limited Decreased: > 63%

Here are our 4 lists of about 20 GO terms each saved in Excel: Media:20 GO Terms for each culture.xls

Potential Explanations for Results:

• Nitrogen Limited Decreased Expression:

A large portion of the decreased processes in cold acclimatization of yeast involve sugar metabolism. A potential reason for the down-regulation of these molecules is freezing point depression. Having solutes in the cytoplasm lowers the freezing point of water and increases change of survival in cold temperatures. In addition, processes involving the movement of ions across the membrane are also down-regulated. The increase of ions in the cytoplasm would have the same effect. In addition, the biosynthesis of glycogen is down-regulated so that these molecules will not be used up to anabolize glycogen.

• Nitrogen Limited Increased Expression:

During acclimatization of yeast to cold temperatures, the organism needs energy molecules to carry out biological processes. Hence, the expression of GMP biosynthesis and GMP metabolism increase. Guanosine monophospate (GMP) is an ester of phosphoric acid containing a phosphate group, a pentose sugar and a guanine base. GMP plays an important role in nucleic acid synthesis for RNA transcription. In addition, both ribosomal subunits are expressed more indicating the necessity to translate proteins during acclimation to cold temperature. The increase in RNA methylation, RNA methyltransferase activity, tRNA methylation and tRNA methyltransferase activity, all indicate that new mRNA and tRNA molecules are being synthesized. Methylation is a postranscritional process that occurs soon after transcription, therefore an increase in these activities indicate that these molecules are being formed, most likely to be used for the production of proteins. IMP (Inosine monophosphate) dehydrogenase is also up-regulated because of its function relative to translation. IMPDH is an enzyme that catalyzes a key step in purine biosynthesis which is necessary for creating RNA during translation. Biotin is necessary for production of fatty acids and metabolism of fats and amino acids. In addition, biotin serves as a carrier of CO2 in the cell. Presumably, biotin would be useful in anaerobic, nitrogen-containing environments in both the breakdown of amino acids and the transport of CO2. Sulfur plays a wide range in cellular processes, therefore increased use of sulfur during cold acclimatization isn’t surprising. Suflur compounds are constituents of methionine (needed for transcription) and biotin.

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