Cholera, Cystic Fibrosis and Hangovers G proteins and second messengers

1. Cholera, Cystic Fibrosis and HangoversG proteins and second messengers

3. Cholera • Cholera is a severe diarrheal disease caused by the bacterium Vibrio cholerae. • The toxin released by the bacteria causes increased secretion of water in the intestine, which can produce massive diarrhea. • Various countries have weaponized cholera toxin for ChemBio warfare

4. The 1991 South American Epidemic • Incidence: 1 in 100,000 worldwide • Infection via contaminated food and water • 1991 Peru Epidemic • Over 1 million cases • Nearly 10,000 fatalities

5. Cholera in the news in the past few hours! Cholera outbreak infects over 100 students in northern NigeriaAFP - 39 minutes agoKANO, Nigeria (AFP) — At least 116 female students in northern Nigeria have been hospitalized with cholera after consuming contaminated beans, ... Cholera kills two in TanzaniaThe Times, South Africa - 5 hours agoDAR ES SALAAM - Two people have died from an outbreak of cholera in southern Tanzania, a local health official said. They were among five people admitted to ... Angola: No Cholera in 48 HoursAllAfrica.com, Washington - 8 hours agoAngola's central Huambo province capital city has not recorded any new case of cholera in the last 48 hours, apart from the 134 people affected since the ... Two die of cholera in MurshidabadTimes of India, India - 20 hours agoBEHRAMPORE: Two people have died of cholera at Beldanga over the last five days. Another 60-odd people are suffering from the disease. ... Cholera claims a woman's life in Kliptown, SowetoAnarkismo.net - 13 hours agoOne of two confirmed cases of cholera in Kliptown, Kelebogile Malefane's death has seen the Gauteng Department of Health spring into an awareness campaign ... …

6. Progression • Cholera is one of the most rapidly fatal illnesses known. A person may become hypotensive within an hour of the onset of symptoms and may die within 2-3 hours if no treatment is provided. • More commonly, the disease progresses from the first liquid stool to shock in 4-12 hours, with death following in 18 hours to several days. • Severe dehydration can cause death. • Given adequate fluids, most people will make a full recovery. • Caused by cholera toxin, a Gprotein inhibitor.

7. Signaling molecules operate over various distances Receptor proteins exhibit ligand-binding and effector specificity

8. Ligand solubility determines receptor location • Steroid (cytosolic) receptors • Cell surface receptors

9. Receptor equations Dissociation Constant Fractional occupancy 1 Occupancy [H] KD

10. Conserved signaling molecules • GTPase switch proteins (G proteins) • Protein kinases • Second messengers

11. GTPase switch proteins (a.k.a. G proteins) • Two roles: • Molecular switches • Molecular timers “Ras” is just the proper name of a particular G protein

12. Protein kinases

13. Second messenger functions • Second messengers are small molecules that convey the message from the receptor to the cell interior. • They provide for: • Amplification - many second messengers are generated per signalling event • Diffusion - most are small and diffusible, so they can go where the signaling molecule cannot.

14. The most common second messengers • Ca2+ calcium is the most common! • IP3 inositol triphosphate • DAG diacylglycerol • NO· nitric oxide • cAMP cyclic AMP

15. Adenylyl cyclase makes cyclic AMP (cAMP) cAMP is broken down by phosphodiesterase Second messengers may be generated by enzymes

16. G protein-coupled receptors • All G protein-coupled receptors (GPCRs) contain 7 membrane-spanning regions • GPCRs are involved in a range of signaling pathways: • light detection, • odorant detection • detection of many hormones • Detection of many neurotransmitters

17. Specificity in GPCRs • The extracellular domain determines ligand specificity • The cytoplasmic domain determines G protein specificity • Together, these two domains link a particular hormone to a particular signaling pathway

18. G protein-coupled receptors and their effectors • Many different cell-surface receptors are coupled to trimeric G proteins • So called because they actually consist of three subunits. • α, β, and γ • We’ll talk about “monomeric” G proteins later. • Ligand binding activates the receptor, …which activates the G protein, …which activates an effector enzyme…to generate an intracellular second messenger • Note: G proteins can either stimulate (Gs) or inhibit (Gi) effector enzymes

19. G proteins and cAMP

20. Inositol triphosphate (IP3) • Another second messenger • Generated by the effector enzyme phospholipase C • PLC cleaves phosphatidyl inositol bis-phosphate (PIP2) in the membrane into IP3 and diacyl glycerol (DAG)

21. IP3 causes calcium release from the ER

22. GPCR NE a a ER/SR PLC PLC IP3 IP2

23. So what has all this to do with cholera?

24. P CFTR Cl- Cl- Basolateral Na+ Na+ Na+ Na+ K+ K+ Apical H2O H2O

25. P CFTR cAMP Basolateral GPCR VIP Cl- Cl- a a Na+ PKA cAMP Na+ PKA AC AC Apical ATP H2O H2O

26. Cholera toxin • Cholera toxin ADP-ribosylates the Gα subunit of the G protein that activates adenylyl cyclase • Gα can no longer hydrolyze GTP

27. R P In Cholera… CFTR cAMP Basolateral GPCR Cl- Cl- ADP-ribosylation PKA cAMP PKA cAMP Na+ Na+ a PKA AC AC Apical ATP H2O H2O

28. P Quick fix for cholera and hangovers • A simple treatment for cholera, or ANY dehydration, is to mix half a teaspoon of table salt and 8 teaspoons of sugar in 1 quart of water. • = lots of water + small amounts of salt (NaCl) + glucose (sucrose = glucose-fructose disaccharide). • Sodium drives the transport of glucose into the epithelial cell • Glucose is osmotically active, and trapped in the body, thus helping retain water. • Gatoraid works too! Basolateral Cl- CFTR Na+ Na+ K+ glucose H2O H2O