Cell Communication

1. Cell Communication Single cell Multicellular organism

2. Budding yeast cells responding to mating factor One haploid cell Another haploid cell Sexual mating (requires many downstream signal pathways to be activated)

3. GENERAL PRINCIPLES OF CELL COMMUNICATION Extracellular signal molecules bind to specific receptors

4. Extracellular signal molecules can act over either short or long distance

6. Autocrine signaling can coordinate decision by groups of identical cells “Community effect” in early development In tumor biology---cancer cells stimulate their own proliferation

7. Gap junctions allow signaling information to be shared by neighboring cells Ca2+, cAMP etc. but not for proteins or nucleic acids Intracellular electrodes, small water-soluble dyes Connexin 43 deficiency --- abnormal heart development

8. Each cell is programmed to respond to specific combinations of extracellular signal molecules

9. Different cells can respond differently to the same extracellular signal molecules

10. The concentration of a molecule can be adjusted quickly only if the lifetime of the molecule is short

11. Nitric oxide gas signals by binding directly to an enzyme inside the target cell Nitroglycerine --- angina Viagra --- PDE inhibitor CO

12. Nuclear receptors are ligand-activated gene regulatory proteins

13. Ligand-binding domain

15. The three largest classes of cell-surface receptor proteins are ion-channel-linked, G-proteins-linked, and enzyme-linked receptors

16. Most activated cell-surface receptors relay signals via small molecules and a network of intracellular signaling proteins

17. Some intracellular signaling proteins act as molecular switches Monomeric GTPase Trimeric GTPase 2% of human genes

18. Signal integration by protein phosphorylation

19. Intracellular signaling complexes enhance the speed, efficiency, and specificity of the response

20. Complex forms transiently

21. Interactions between intracellular signaling proteins are mediated by modular binding domains

22. www.cellsignal.com PDZ Domain Domain binding and function:PDZ domains bind to the C-terminal 4–5 residues of their target proteins, frequently transmembrane receptors or ion channels. These interactions can be of high affinity (nM Kd). The consensus binding sequence contains a hydrophobic residue, commonly Val or Ile, at the very C-terminus. Residues at the –2 and –3 positions are important in determining specificity. PDZ domains can also heterodimerize with PDZ domains of different proteins, potentially regulating intracellular signaling. In addition to engaging in protein-protein interactions, several PDZ domains including those of syntenin, CASK, Tiam1 and FAP are capable of binding to the phosphoinositide PIP2. PIP2-PDZ domain binding is thought to control the association of PDZ domain-containing proteins with the plasma membrane. Structure Reference: Doyle, D.A. et al. (1996) Cell 85(7), 1067–1076. The third PDZ domain from PSD-95.

23. Binding Examples: PDZ domain proteins Binding partners domain binding sites Post-synaptic Density Protein 95 (PSD-95) NMDA receptor B via PDZ1 and PDZ2 of PSD-95 – IESDV-COOH Post-synaptic Density Protein 95 (PSD-95) Kvl1.4 Shaker-type K+ channel via PDZ1 and PDZ2 of PSD-95 – VETDV-COOH Post-synaptic Density Protein 95 (PSD-95) Neural Nitric Oxide Synthase (nNOS) via PDZ2 PDZ/PDZ interaction

24. Lipid raft Enriched in cholesterol and glycolipids c-Src tyrosine kinase

25. Cells can respond abruptly to a gradually increasing concentration of an extracellular signal effector/target : 1~16 Chicken oviduct cells Stimulated by estradiol maximal activation

26. One type of signaling mechanism expected to show a steep thresholdlike response

27. A cell can remember the effect of some signals Signals trigger muscle cell determination Autophosphorylation of Ca2+/CaM-kinase II

28. Cells can adjust their sensitivity to a signal

29. SIGNALING THROUGH G-PROTEIN-LINKED CELL-SURFACE RECEPTORS 1. The largest family of cell-surface receptors 2. 5% of the C. elegans genes 3. Signal molecules: hormones, neurotransmitters and local medicators 4. Rhodopsin-light receptor 5. Genome sequencing --- vast numbers of new family members 6. Major targets for drug discovery

30. Trimeric G proteins disassemble to relay signals from G-protein-linked receptors Transducin-G protein in visual transduction

31. The disassembly of a activated G-protein into two signaling components

32. The switching off of the G-protein a subunit by the hydrolysis of its bound GTP RGS proteins --- regulators of G protein signaling, act as a subunit-specific GTPase activating proteins (GAPs) ~25 RGS proteins in the human genome

33. Some G-proteins signal by regulating the production of cyclic AMP ~5 X 10-8 M >10-6 M Nerve cell culture, preloaded with a fluorescent protein that changes its fluorescence when it binds to cAMP. (Science 260:222-226, 1993)

34. cAMP-dependent protein kinase (PKA) mediate most of the effects of cyclic AMP Role of cAMP, PKA in glycogen metabolism

35. How gene transcription is activated by a rise in cAMP concentration (CRE, cAMP response element) Role of protein phosphatases?

36. Some G-proteins activate the inositol phospholipid signaling pathway by activating phospholipase C-b (<1% of total phospholipids)

38. The two branches of the inositol phospholipid pathway

39. Ca2+ functions as a ubiquitous intracellular messenger Ca2+ signaling in fertilization of starfish, detected by Ca2+-sensitive fluorescence dye

40. The main ways eucaryotic cells maintain a very low concentration of free Ca2+ in their cytosol

41. The frequency of Ca2+ oscillations influences a cell’s response In a liver cell

42. Ca2+/calmodulin-dependent protein kinases (CaM-kinases) mediate many of the actions of Ca2+ in animal cells A peptide derived from CaM-Kinase II The structure of Ca2+/calmodulin

43. The activation of CaM-kinases II ~2% of total mass in some brain regions, especially in synapses • It can function as a molecular memory device --- • Learning defect (where things are in space) in mutant mice that • lack the brain-specific subunit of CaM-kinase II • (2) Same defect also observed in mutant mice that • have their CaM-kinase II mutated at the autophosphorylation site

44. CaM-kinase II is immobilized on a solid surface +a brain protein phosphatase +repetitive pulse of Ca2+/calmodulin at different frequency Kinase activity assay CaM-kinases II as a frequency decoder of Ca2+ oscillations What a nice experiment it is!

45. Smell and vision depend on G-protein-linked receptors that regulate cyclic-nucleotide-gated ion channels

46. Cyclic GMP A rod photoreceptor cell

47. The response of a rod photoreceptor cell to light

48. Extracellular signals are greatly amplified by the use of small intracellular mediators and enzymatic cascades Amplification in the light-induced catalytic cascade in vertebrate rods

49. G-protein-linked receptors desensitization depends on receptor phosphorylation

50. SIGNALING THROUGH ENZYME-LINKED CELL-SURFACE RECEPTORS Six classes: 1. Receptor tyrosine kinases 2. Tyrosine kinase-associated receptors 3. Receptorlike tyrosine phosphatases 4. Receptor serine/threonine kinases 5. Receptor guanylyl cyclases 6. Histidine-kinase-associated receptors