1. Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4.1.2-08/1/A-2009-0011

2. SECOND MESSENGERS Tímea Berki and Ferenc Boldizsár Signal transduction Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4.1.2-08/1/A-2009-0011

3. TÁMOP-4.1.2-08/1/A-2009-0011 Second messengers Hydrophylic molecules: cAMP, cGMP, IP 3, Ca 2+ Hydrophobic molecules (lipids): diacylglycerol (DAG), phosphatidylinositols Gases: NO,CO, (H 2 S)

4. TÁMOP-4.1.2-08/1/A-2009-0011 General scheme of intracellular signaling Second messengers activates intercellular process G-protein is activated and produces effector Effector stimulates second messenger synthesis Agonist activates membrane bound receptor

5. TÁMOP-4.1.2-08/1/A-2009-0011 Discovery of second messengers - cAMP E. W. Sutherland Adrenaline effect of the liver is mediated through cyclic-AMP 1971. Nobel Prize in Physiology and Medicine

6. TÁMOP-4.1.2-08/1/A-2009-0011 Synthesis of cAMP O OH ATP cAMP AMP H H H CH 2 NH 2 N N N N O O O O P O O O P O O O P Adenylyl cyclase PP i O OH H H H CH 2 NH 2 N N N N O O O P H H cAMP phosphodiesterase H2OH2O H O OH H H H CH 2 NH 2 N N N N O H O O P O

7. TÁMOP-4.1.2-08/1/A-2009-0011 cAMP activates Protein kinase A Inactive PKA Activated PKA R R C C C C cAMP cAMP cAMP cAMP R R C C C C C C C C R R cAMP cAMP cAMP cAMP Regulatory subunit Catalytic subunit

8. TÁMOP-4.1.2-08/1/A-2009-0011 PKA targets Enzymes Stucture proteins Transcription factors: CREB CREB: dimeric form binds to cAMP-responsive elements (CRE) CREB CRE

9. TÁMOP-4.1.2-08/1/A-2009-0011 Phospholipase C Hormone Receptor Plasma membrane Cytoplasm IP3 opens Ca 2+ channel Lumen of smooth endoplasmatic reticulum IP 3 R DAG PKC Ca 2+ IP 3 GTP    G protein PIP 2 GTP PLC 

10. TÁMOP-4.1.2-08/1/A-2009-0011 IP3 receptor pathway Hormone Receptor Plasma membrane Cytoplasm IP 3 R DAG GTP  PLC PIP 2 IP 3 Ca 2+ Lumen of smooth endoplasmatic reticulum Ca 2+ Pump Ca 2+ channel + - GTP    G protein Ca 2+

11. TÁMOP-4.1.2-08/1/A-2009-0011 Gases as second messengers: NO way! “Endothelium-derived relaxation factor” (EDRF): mediator(s) produced by endothelial cells causing vasodilatation 1980s: NO!! [Nature 1983 Nov 10- 16;306(5939):174-6.] 1992: NO:“molecule of the year” (Science); “Nitric Oxide Society” established 1998: Nobel Prize in Physiology or Medicine – F. Murad, R.F. Furchgott, L. Ignarro, S. Moncada ~3000 papers/year published about NO!

12. TÁMOP-4.1.2-08/1/A-2009-0011 NO synthesis NO synthase (NOS): 1 1eNOS: endothelial 2 2iNOS: inducible (eg. macrophages) 3 3nNOS: neuronal 2 major domains: 1 1N-term. Oxygenase (Heme-thiolate proteins) 2 2C-term. Reductase (~NADPH-cytochromeP450 reductase) 3 3Linker: calmodulin-binding sequence L-Arg H2NH2N + NH 2 NH NH 3 + H O O – NADPH + O 2 NADP + + H 2 O RNOHLA 1/2 NADPH + O 2 1/2 NADP + + H 2 O N NH 2 NH NH 3 + H O O – HO L-Citrulline H2NH2N + NO NH NH 3 + H O O – O

13. TÁMOP-4.1.2-08/1/A-2009-0011 NO-cGMP pathway Vascular lumen Endothelial cell Smooth muscle cell Acetylcholine Shear stress Bradykinin L-arginine L-citrulline NO GTP cGMP GC eNos Ca 2+ ↑ Vasodilatation Platelet aggregation ↓ Smooth muscle growth ↓ Neutrophil adhesion ↓ NO inactivation Oxidative stress ONOO -.OH M B2

14. TÁMOP-4.1.2-08/1/A-2009-0011 cGMP

15. TÁMOP-4.1.2-08/1/A-2009-0011 Protein kinase G Ser/Thr protein kinase activated by cGMP Expressed by vascular smooth muscle cells, platelets, endothel, heart muscle, fibroblasts, renal cells, leukocytes, nervous system Regulates smooth muscle relaxation, platelet function, sperm metabolism, cell division and nucleic acid synthesis

16. TÁMOP-4.1.2-08/1/A-2009-0011 Functions of NO Vascular effect: vasodilatation –e.g. Nitroglycerin: treatment of coronary disease (Angina pectoris) –e.g.Viagra: erection Heart: contractility and heart rate ↓ Immune system: macrophages produce NO to kill bacteria BUT in severe systemic infection (Sepsis) this leads to generalised vasodilatation and SHOCK (Septic shock)