MICRB 265 General Microbiology

1. MICRB 265 General Microbiology Topic 5D – Signal Transduction

2. Last Topic • Intercellular signaling – quorum sensing • Case study: bioluminescence by Vibrio fischeri

3. This Topic • Signal Transduction and two-component regulatory systems • 12: 9.5; 11E: 8.12 • Case study: OmpC/OmpF osmolarity response in E. coli • Know this one well! • Case study: regulation of chemotaxis • Not covered in detail

4. Signal Transduction • Bacteria respond to a wide variety of environmental signals • Changes in pH, temp., salinity, available nutrients, etc. • Recall from negative and positive control: environmental signal could be an effector (e.g. inducer – allolactose; AHL-quorum sensing) • Small molecule in environment serves as a signal (single component-DNA binding protein)

5. Two-Component Systems • External signal sensed by a sensor protein (first component) – then signal transmitted to regulatory protein (second component) • Signal transduction - process of signal moving from environment to affect gene expression

6. Two-Component Systems • Sensor kinase protein • Kinases – phosphorylate other molecules • Uses ATP • Sensor kinases autophosphorylate  then phosphate transferred to next component • Response regulator protein • Accepts phosphate from #1  changes shape • DNA-binding protein

7. 2-Component Figure

8. Case Study: OmpF/OmpC • E. coli has > 50 2-component systems • Many examples in eukaryotes • OmpF/OmpC – outer membrane porin proteins • Allow diffusion of small hydrophilic molecules across OM

9. Case Study: OmpF/OmpC • OmpF is wider in diameter than OmpC • Allows in more nutrients (and other things) • OmpC protects against bile salts • OmpF used when osmotic pressure is low; OmpC when pressure high

10. OmpC/OmpF Gene Regulation • 2-component system: • Sensor kinase: EnvZ (senses osmotic pressure; high osmolarity  autophosphorylate) • Response regulator protein: OmpR • If OmpR phosphorylated  activates Ts of ompC, represses Ts of ompF • Serves as an activator and repressor

11. Another Example • Chemotaxis in E. coli • Not a gene expression system Sensor kinase Response regulator

12. Sample Question In a conventional two-component regulatory system in Gram negative bacteria, • the sensor kinase is located in the outer membrane (LPS layer) to sense the external environment. • the response regulator is always an activator protein. • the sensor kinase forms a complex with two other proteins to span the cell membrane and outer membrane. • the response regulator is a soluble (cytoplasmic) protein. • the response regulator protein may also have phosphatase activity.

13. Sample Question If you mutated E. coli so that the response regulator OmpR could not be dephosphorylated, then put the mutant E. coli into the human gut, what might happen? • It might die because it would be inhibited by bile salts in the gut. • It should survive well because it would have OmpF porins with large pores to take up all the nutrients. • It should survive because it would produce OmpC porins with small pores. • It might die because it produced OmpF porins with large pores. • It might survive but produce both kinds of porins (OmpC and OmpF).

14. Sample Question Instead, if you mutated the envZ gene in E. coli so that EnvZ could not autophosphorylate, and inoculated it into bile esculin medium, what might happen? • The E. coli would grow well, as if it was the wildtype (i.e., no effect on growth). • EnvZ would not be able to phosphorylate OmpC, so the cells would be inhibited by the bile salts. • The E. coli might produce only OmpC porins with small pores. • micF RNA would bind to the OmpC mRNA to prevent production of OmpC porins. • None of the above.