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After stationary phase (3), two additional phases reported PowerPoint Presentation
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After stationary phase (3), two additional phases reported

After stationary phase (3), two additional phases reported

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After stationary phase (3), two additional phases reported

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  1. After stationary phase (3), two additional phases reported 4. Death phase (day 3-4 for E. coli lab culture) 5. Long term stationary phase (5-7 days) Death and Survival Long term stationary phase Lloven 2010 FEMS Micro. Rev. 34:476 Fig 1

  2. A. Stationary Phase Response A variety of protective phenotypic changes Increase expression of 50 genes Mediated by RpoS (σS or σ38) – affects 10% of genome Fig 13.5 Microbe

  3. 2. Regulation of RpoS Regulation of RpoS levels and activity Lloven2010 FEMS Micro. Rev. 34:476

  4. 2. Regulation of RpoS A. Transciptional Regulation of RpoS levels • Several factors act in positive and negative ways to regulate transcription of rpoS • Crp-camp-catabolite repression Lloven2010 FEMS Micro. Rev. 34:476

  5. 2. Regulation of RpoS B. Translational Control A couple of small RNAs are induced in stationary phase and by other stresses DsrA, RprA and OxyS DsrA and RprA act to stimulate RpoS translation by binding to mRNA and removing inhibitory secondary structure Most turn down expression(translation of genes)

  6. A. Stationary Phase Response B. Translational Control sRNAs that affect translation of rpoS mRNA and/or other regulators Fig 13.6 Microbe

  7. 2. Regulation of RpoS B. Translational Control DsrAsRNA binds to rpoS mRNA and changes mRNA configuration so that mRNA is translated Fig 13.7 Microbe Lloven2010 FEMS Micro. Rev. 34:476

  8. 2. Regulation of RpoS C. Post-Translational Control – Protein Stability No stress – RssB-P associates with RpoS and RpoS is rapidly degraded by ClpXP Stress – RssB is not phosphoylated, RpoS is not degraded Lloven2010 FEMS Micro. Rev. 34:476

  9. 90-99% of cells die Why do the cells die? B. Death Phase Long term stationary phase Lloven 2010 FEMS Micro. Rev. 34:476 Fig 1

  10. B. Death Phase Possible Causes of Cell Death 1. Stochastic cell death – some cells die due to progressive accumulation of damaged molecules in starved cells, oxidative damage 2. Programmed cell death – Altruistic death response of many cells in population mediated by toxin-antitoxin modules? Toxin – RNase, cleaves mRNA and inhibits protein synthesis Antitoxin – sRNA or labile inhibitory protein

  11. Toxin – Antitoxin module MazEF Toxin – antitoxin system • Stress – mazEF transcription reduced • Antitoxin MazE is unstable • MazFcleaves mRNA which inhibits most protein synthesis • A few small proteins are still made • Most cells do PCD • Some may enter stasis Lloven 2010 FEMS Micro. Rev. 34:476 Fig 4

  12. B. Death and Survival If you had a million cells in the population and 99% of them died, how many cells would be die and how many would be left? 10,000

  13. B. Death and Survival What would be the advantage to the population as a whole for cells to commit suicide (undergo programmed cell death) after being in stationary phase for a while? fewer cells competing for the limited resources Dead cells would lyse and release their cell components which would be nutrients to remaining cells Release dna which could be taken in by transformation and generate more genetic diversity in survivors

  14. C. How do cells survive survive long term? Death and Survival Long term stationary phase Lloven 2010 FEMS Micro. Rev. 34:476 Fig 1

  15. C. Survival Responses Phenomena observed Make reserve material while in log phase GASP (growth advantage in stationary phase) phenotype Mutator phenotype Viable but nonculturable (VBNC) state Persisters – resistant to antibiotics SCDI – stationary phase contact-dependent inhibition Sporulation

  16. 1. Reserve materials Most bacteria accumulate one or more reserve materials during log phase Use during starvation (endogenous metabolism) See Section 13.1 Types Carbohydrates Lipids Polypeptides Polyphosphates

  17. 1. Reserve materials Carbohydrates – Glycogen – chain of glucose (1,4 and 1,6) Trehalose – disaccharide, two glucoes with 1,1 linkage

  18. 1. Reserve Materials b. Lipids Triacylglycerides – fats or oils, in actinomycetes Polyhydroxyalkanoates (PHA) such as poly-B-hydroxybutyrate made from acetyl-CoA Length – 4 to 18 units Fig 13.3 Bact Met & Phy

  19. 1. Reserve Materials c. Polypeptides Phycocyanin – asparate and Arg Cyanophycin - pigment d. Polyphosphates 4- 100s of phosphates linked together

  20. C. Survival Responses 2. GASP phenotype Stable mutations that confer a greater fitness or advantage in stationary phase such as a mutation in rpoS or lrp and transposon insertion that increases expression of ABC transporter for aspartate and glutamate Have observed different subpopulations growing in long term stationary phase Long term stationary phase

  21. Survival Responses 3. Mutator phenotype in stressed cells Induction of RpoS(σS) also causes repression of methyl-mismatch repair and induction of error-prone DNA pol IV Is this Lamarckism? Why is this beneficial? Get more mutations in dna Increases genetic diversity of cells May make a mutation that allows cells to grow

  22. Survival Responses 4. Viable but nonculturable (VBNC) state On prolonged starvation, some cells appear to enter a state of low metabolic activity (viable) and non-growing (dormant) Observed now in several species Mechanism unknown Some have been resuscitated by placing in favorable growth conditions

  23. Survival Responses 5. Persisters Observed in antibiotic treated individuals A few cells survive antibiotic treatment, later grow again in persistent or chronic infection Persistersthought to be dormantsubpopulation of cells same as VBNC? Enriched in biofilms

  24. Survival Responses 6. SCDI – stationary phase contact-dependent inhibition Mutant subpopulation that kill or inhibit growth of the parental strain One example – had mutant that overproduced glycogen may be another example of GASP