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13 Cards in this Set
- Front
- Back
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What cell components could provide protection and therefore act as biomarkers of robustness? |
mRNA (transcript itself), proteins (specific protein molecule), enzymes (activity) |
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How to assess what things are activated in mildly stressed cell? |
Check on TRANSCRIPTOME level (on microarray glass light) -> which mRNA in cells then (which genes are upregulated?) -> based on acid, heat, salt and H2O2 stress - isolate mRNA from unstressed and stressed cell -cDNA from both on microarray More green - higher cDNA from unstressed; more red - higher cDNA from stressed Yellow - comparable amount of cDNA from both |
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What 3 themes were found in gene expression in stressed cells? (Candidate biomarkers) |
1)region on stress regulator sigmaB 2)chaperones and proteases (Clp's; enzymes removing damaged cells) 3)scavengers of reactive oxygen species (catalases) |
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What is sigma B? |
Transcription factor which allows transcription of stress related genes It binds to core RNA polymerase which then binds to promoter of a gene specifically involved in stress response |
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What's the difference in survival during lethal heat between wild type and disavtivated sigma mutant? |
Both rapidly decreasing if exposed to lethal stress Exposed to mild heat treatment before lethal exposure - wild type survives better than mutant without sigB (wild type is much more robust than non adapted one and than the mutant) |
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What's the role of chaperones and proteases? |
Misfolded protein - chaperones refold it - active again Proteases cut misfolded proteins into pieces to get rid of them (degradation) |
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What's the role of ROS scavengers? |
Leakage of e- to oxyxen - superoxides formation (O2-) which are toxic Superoxide dismutase - superoxide to hydrogen peroxide which is then automatically transformed to hydroxyl radicals (•OH) which are even more toxic Catalases - H2O2 into H2O + O2 |
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How to know if something is a biomarker? |
If quantitative correlation between mildly stressed cells biomarkers and enhanced robustness |
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What happens if a cell is exposed to H2O2 in mild concentration? |
Most robust towards LETHAL concentration of H202 if exposed to mild H2O2 for 10-60 minutes Cross protection present towards heat and acid (also after 10 minutes) - better protection against heat than acid |
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How is it that H202 exposure resulted in protection against heat and acid? |
Measure radicals inside cells - the more fluorescence, the more radicals High heat - increase of signal - more radicals in the cells - adaptation leads to presence of superoxide dismutase and catalases which will get rid of them and hence cross protection Acid stress - exposure to acid will produce more radicals than in unstressed cells - also protection from oxidative stress treatment results |
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Could catalase activity work as a biomarker for mild stress induced robustness? |
-Mild oxidative stress treatment for 15-30 minutes best for catalase activity (brightest colour) - 1 log difference with unstressed cells
-Compare graphs of increased catalase activity and increased robustness towards stress like heat - similar trend with heat and H2O2 (good correlation-significant so catalase activity is a good biomarker) but not with acid
STRESS SPECIFIC - ok for heat but not ok for acid |
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How is quality of a biomarker strain specific? |
Bacillus group - same across species eg. Cereus and weihenstephanensis |
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Does mild stress adaptation enhance survival of probiotic bacteria? |
Yes - increased robustness towards heat and salt (osmotic stress) Means that it's not only for bacillus but also for other genera |
