Chapter 6- Cell-cell communication

1. Chapter 6- Cell-cell communication More definitions _________-interaction between two or more distinct cells or tissues _________- the cell of tissue producing the signal __________- the cell or tissue being induced _________ the ability to respond to a given inducer Fig. 6.2 Example- Pax6 is required for optic vesicle to respond to an inducer, but Pax6 is not the inducer Lacking nose and eyes Hence, Pax6 makes cells _____________ Pax6 null Wild-type

2. _____________ interaction-Tissue A requires tissue B to respond in a certain way (analogy to a book _______________) • Instructive vs permissive interactions ____________ interaction-Tissue A does not require tissue B to respond in a certain way, but only needs to be in a certain environment (analogy to a book ______________)

3. Epithelial-mesenchymal interactions Fig. 6.7 • __________- sheets of cells from any germ layer • ____________- unconnected cells (from mesoderm or neural crest) • All organs have both of these cell types Mesen- chyme Wing epithelium Wing _______ specificity Thigh Epithelial-mesenchymal interactions Foot ________ specificity Thus, mesenchyme __________ epithelium

4. Epithelial-mesenchymal interactions Newt with tadpole suckers Regional specificity Epithelial-mesenchymal interactions _______ specificity Frog gastrula Newt gastrula Thus, mesenchyme dictates _______ type, but epithelium dictates ____ of the organ Newt gastrula Frog gastrula Frog with newt “balancers”

5. Paracrine factors __________factors- diffusible molecules that can travel small distances to signal a neighboring cell __________signaling- cell-cell interactions by direct contact Example of _________ signaling • Fibroblast growth factor (FGF) – binds FGF receptors (FGFRs) • These are receptor tyrosine kinases FGF (________) FGFR (______) __________________ yields activation and subsequent phosphorylation of a second protein

6. Other well-known paracrine factors • _________ family- (3 in vertebrates)-create boundaries, induce development • _____ family- (15 in vertebrates)- limb polarity, muscle development • _________ superfamily (>30 members)- bone, kidney, neuronal , etc. differentiation

7. Cell surface receptor pathways BLA512 1/5/98 update Smad Smad Smad Smad STAT STAT STAT STAT Elk-1 NFkB CREB Gene Fos Fos jun jun AP1 AP1 P P Enzyme-linked Ion channel-linked No details shown G-protein-linked PDGF, EGF, IFa,b,and g, IL-2 ,IL-3, IL-4, IL-6 TNFa TGF-b PLCg Grb2 Jak G proteins TRADD Gs Golf FADD Gq Go GNRPs (e.g. Sos) TRAF Complexity! Ca++ from ER Ras IP3 PIP Adenylyl cyclase Cytoplasm Raf (a MAPKKK) PLCb DAG MEKK (a MAPKKK) NIK AMP cAMP MAPKK (e.g.MEK) PKC JNKK IKK PKA Caspases JNK IkB/NFkB IkB/NFkB MAP-kinase (ERK) Cell death Elk-1 jun JNK Gene Gene Nucleus Gene Gene

8. Signal transduction pathways Phosphorylation is key General pathway Fig. 6.14 A. Receptor tyrosine kinase (RTK) • ________ binds receptor • Receptor undergoes _______ • Receptor ____________ occurs • Receptor __________________ • Receptor binds _____________ • Adaptor protein binds ________ • G-protein recruits ____ • Raf phophorylates ____ • ____ phosphorylates ERK • ERK phosphoryates a • ____________________ • Transcription is ____________

9. B. TGF-b signalling- a simpler pathway • Ligand binds _______ • Two _______ receptors dimerize • _____________________occurs • Receptor phosphorylates ________ • SMADs ___________ • SMADs enter _________and bind ____ • Transcription is _____________ Fig. 6.20

10. C. JAK-STAT pathway- also a simpler pathway Fig. 6.21

11. D. Wnt signaling If mutate B-catenin, constitutive activation of myc gene– Tumor formation

12. Apoptosis Jacobson et al., Cell 88:347 (1997)

13. Fas-null mice Lymph nodes Spleen Too much and too little • Too much- ________________ disease • Alcohol-induced liver disease • Autoimmune disease • Primary biliarry cirrhosis • Wilson’s disease • Ischemia reperfusion injury • Virus hepatitis • Too little- ___________ • Splenomegaly • Lymphadenopathy • Cholangiocarcinoma • Hepatocellular Carcinoma

14. Apoptosis is required for normal development Elegans Mammals Fig. 6.28 Apaf-1 knock-out Wild-type Fig. 6.27

15. Cleavage of Death substrates • structural proteins (e.g. actin) • kinases (e.g. MEKK, PKC) • cell cycle proteins (pRb, PARP) • DNA repair enzymes • DNA nucleases • Anti-apoptotic proteins (Bcl-2) Receptor-mediated Apoptosis Caspases-3, -6 and -7 Protective

16. Monitoring Apoptosis by ______________ Hoechst

17. 0h 0h 3h 3h Monitoring Apoptosis by _______________ Fg-14 1000 bp ladder M38 100 bp ladder 7h 7h 5 kb 2 kb 1 kb 0.5kb

18. Monitoring Apoptosis by _________ 1000X M38 400X Fg14 1000X Hoechst TUNEL

19. An example of __________ signaling _________ signaling Cell 1 Delta (Ligand) Notch (receptor) Cell 2

20. Another example of ____________ signaling The ___________________ • The stuff between cells • Affects cell adhesion, _______________, epithelial sheet formation • Includes collagen, proteoglycans, fibronectin and laminin _______ are the _______________ for extracellular matrix molecules

21. Integrins interact with both extracellular and intracellular scaffolds Fibronectin Integrin Actin

22. Another form of communication- _______transmission of signals through __________________ Fig. 6.38 • Does not requires a __________, only regulation of small molecules through a port • Ports are composed of _______________

23. Signaling pathways exhibit ____________- • A major challenge in biology- How to get specificity from _______________pathways Example: Two pathways direct lymphocyte development Point of _________ Fig. 6.40

24. LPB LPS FasL TNF CD14 TNFR TLR-4 FasR FADD IL1R1 TRADD FADD Caspase 8 Caspase 8 TAK1 RIP TGFBR TRAF2 P38 MAPK MyD88 Sorb. P38 MAPK NIK TRAF6 IRAK TNF TPL-2 MEKK1, 2, 3 PKC EGFR JNKK IKK MEK Calyculin A, Okadaic Acid JNK PD098059 p65 ERK1,2 PDTC c-jun ALLN, HMA p105 Apoptosis IkB NF-kB Proteasome SN50 P105 phos, degraded NF-kB-responsive genes ROS A1, A20, ,Fas, FasL,TNF, Bcl2, TRAF1,2, c-IAP1,2 LPS-mediated apoptosis: Which pathway is defective?