MIC 329 The Gram-positive World

MIC 329 The Gram-positive World

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MIC 329 The Gram-positive World

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1. MIC 329 The Gram-positive World* *(Well, really a talk about my favorite Gram (+) organism, Bacillus subtilis)

6. The ongoing schism of the genus Bacillus Geobacillus Thermobacillus Virgibacillus Salibacillus Paenibacillus Gracibacillus

7. Why study Bacillus subtilis? Best-characterized Gram-positive bacterium Biochem., metabolism well-studied Good genetic system (transformation, transduction) Advanced molecular biology techniques Entire genome sequenced / annotated Easy to grow, manipulate in culture Widely used in industry, agriculture ?Simple? model for cellular differentiation

9. B. subtilis spore anatomy

10. Endospores are resistant to: Heat (both wet and dry) Ultraviolet (UV) radiation Extreme desiccation (including vacuum) Lysozyme Chemicals (organic solvents, oxidizing agents, etc.)

12. Abundance of spores in extreme locales

13. Solar UV: ~10 J /m2 sec UV-B (noon) ~25 J/m2 sec UV-A (noon) Temperature extremes: Avg. -7 to +46 ?C (air) ~70-80 ?C (surfaces) Desiccation: Avg. 13%-30% RH Avg. 28 cm rainfall / year

14. Spores are 1-2 orders of magnitude more UV resistant than vegetative cells

16. DNA Protective Factors in Spores Spore coat proteins Spore pigment in coat Dipicolinic acid in core SASP in core

21. Spore photochemistry is due to SASP-DNA interaction SASP are Small, Acid-soluble Spore Proteins SASP are synthesized at Stage III of sporulation SASP bind to DNA and shift its conformation from B to A UV irradiation of SASP-DNA complexes results in formation of SP and not T<>T

23. The UV photochemistry of DNA in vegetative cells and spores is different

26. SP is repaired in germinating spores by SP lyase and NER

27. Spores of B. subtilis DNA repair mutants respond differently to lab UV and Solar UV

28. Do spores exposed to solar UV accumulate different types of DNA damage(s)?

29. Probing DNA damages with EndoV and alkali

32. SP is repaired in germinating spores by SP lyase and NER

33. SP lyase-mediated DNA repair in B. subtilis Encoded by splB gene. Synthesized at Stage III of sporulation, packaged in the dormant spore. Active during spore germination. Direct reversal of SP to thymines in situ. ?Dark repair? process.

34. Organization and expression of the splAB operon in B. subtilis

35. ?Radical SAM? Model for SP repair

36. CONCLUSIONS In the laboratory: Spores are highly UV resistant. SP is the major DNA damage. CPD, ss, ds breaks negligible at biol. relevant UV doses. SP lyase > NER during germination.

37. Survival and persistence of bacterial endospores in extreme environments Patricia Fajardo Mario Pedraza Lilian Chooback Roberto Rebeil Heather Glanzberg Paul Riesenman Jocelyn Law Tony Slieman Rachel Mastrapa Yubo Sun Heather Maughan Yaming Xue

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