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Endocytosis in eukaryotic cell

Endocytosis in eukaryotic cell. Nutrient internalization : LDLR Signal transduction Composition of the plasma membrane Cell fate determination , tissue remodeling and motility LDLR and signaling. Pathways of receptor-mediated endocytosis. AP2. Rab5. PH6-6.5. Rab11. Rab7. PH4.5-5.5.

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Endocytosis in eukaryotic cell

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  1. Endocytosis in eukaryotic cell • Nutrient internalization : LDLR • Signal transduction • Composition of the plasma membrane • Cell fate determination , tissue remodeling and motility • LDLR and signaling

  2. Pathways of receptor-mediated endocytosis AP2 Rab5 PH6-6.5 Rab11 Rab7 PH4.5-5.5 Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  3. Endocytic transport itineraries of receptor tyrosine kinases TGF-a dissociate EGF bound Ligand free-recycle Ligand bound-recycle or to lysosome Sorting in MVBs Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  4. Dynamin-dependent endocytosis of RTKs With ubiquitination—lysosome (ErbB1) Without—recycling (ErbB2)

  5. Dynamin-dependent endocytosis of RTKs Shibire mutant

  6. Dynamin: A Molecular Motor with Pinchase ActionShibire mutant A highly conserved tripartite GTP- binding domain is located at the N terminus PHD: pleckstrin homology domain (a membrane-binding domain) CC: coiled-coil (may act as an effector domain to activate the dynamin GTPase activity) PRD: a proline-rich domain (interact with numerous other proteins)

  7. Endocytic transport itineraries of G-protein-coupled receptors b-adrenergic receptor D2 dopamine receptor: dynamin dependent and independent Different postendocytic transport Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  8. Desensitization and resensitization of G-protein-coupled receptors during endocytosis Endocytosis = Rapid resensitization b-arrestin Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  9. Termination of receptor-tyrosine-kinase signaling in multivesicular bodies Cytoplasmic domain Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  10. MVBs

  11. Endocytic protein complexes modified by signaling receptors • Clathrin • Dynamin tyrosine phosphorylation and ubiquitylation • Eps15 • e.g. Egfr --Y phosphorylated recruits Cbl • then Egfr is ubiquitylated • d. Ral • e Rab5 dominant negative inhibit endocytosis of Egfr and insulin receptor Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  12. Ubiquitination : Targeting of proteinsfor degradation by the proteasome

  13. The ubiquitylation pathway Nat. Rev. Mole. Cell Biol. 4, 497, 2003

  14. Protein ubiquitylation Mono-ubiquitylation Multi-ubiquitylation poly-ubiquitylation Nat. Rev. Cell Biol. 2, 195, 2001

  15. RTK journey to the lysosome Oncogene, 23, 2057, 2004

  16. Ubiquitin and ubiquitin receptor in endosome fusion Vps: vacuolar protein sorting Vps9=GEF Activates Vps21 to regulate Rab5 (Rabex-5) Ubiquitin interacts with Vps9 to release inhibition E3 ligase Nat. Rev. Mole. Cell Biol. 4, 497, 2003

  17. Protein complexes modified by signaling receptors in endosomes Rab 5 promote membrane fusion and endocytosis Egfr—Ras—RIN—activated Rab5 --Vps34—PI(3)P– membrane composition Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  18. Hypothetical model of signaling complexes in endosomes Signaling protein found in endosomal compartment Different from plasma membrane? Small GTPases translocate to endosome with activated Egfr Nat. Rev. Mole. Cell Biol. 3, 600, 2002

  19. Multiple pathways of endocytosis Cell, 115, 513, 2003

  20. Endocytosis of membrane receptor Eps15 + clathrin Eps15 + ubiqutin Prolonged endosomal signaling UIM –ubiquitin-interacting motif Epsin Epsin15 Epsin15R PNAS, 102, 2679, 2005

  21. Endocytosis and cell fate determination Notch pathway Wingless pathway

  22. Notch signaling pathway nervous system: selection of a single neuroblast (lateral inhibition) Notch –transmembrane protein DSL family—Delta, Serrate, Lag-2 Kuzbanian—cleave Notch ECD Presenilin—cleave Notch ICD

  23. Cleavage of Notch Kuzbanian—cleave Notch ECD Presenilin—cleave Notch ICD

  24. Dissociated NotchECD colocalizes with Delta in control retinas discs. shi Ts—320C wt red green green red Shi=dynamin surface vesicles vesicles Development, 127, 1373, 2000

  25. The endocytosis of Notch ligands is required for notch activation Cell 124,897

  26. Ligand-engaged and unliganded Notch Cell 124,897,2006

  27. Creation of asymmetry in pIIa and pIIb cells 1. Notch endocytosis (Numb and a-adaptin) 2. Sanpodo (required for Notch signaling) endocytosis 3. Delta internalization to Rab11 (+) recycling endosome and recycle to plasma membrane Rab11 based mechanism is suppressed Cell 124,897,2006

  28. The Endocytic Pathway as the Switch-offfor Receptor Signaling Regulated Endocytic Routing Modulates Wingless Signaling in Drosophila Embryos Cell, 105, 613-624, 2001 Laurence Dubois, Magalie Lecourtois, Cyrille Alexandre, Elisabeth Hirst, and Jean-Paul Vincent

  29. Intercellular signaling set up Segment boundary Segments Wingless (Armadillo and engrailed) Hedgehog Developmental Biology by Scott F. Gilbert Sinauer Associates, publishers Six edition, 2000

  30. D-frizzled 2-receptor Dishevelled-cytosolic phosphoprotein Zw3/GSK3-Ser/Thr kinase Arm + Pan/lef1— transcription Arm/b-catenin interacts with Cadherin Colon cancer-APC Oncogenic-c-myc Developmental Biology by Scott F. Gilbert Sinauer Associates, publishers Six edition, 2000

  31. A Transition in the Distribution ofWingless Red—Engrailed , Green--Wingless

  32. A Transition in the Distribution of Wingless Until early stage 10, Wingless spreads into the engrailed domain and then recedes sometime during stage 10. At stage 12, Wingless-containing vesicles are infrequently detected in engrailed-expressing cells. At this stage, rhomboid begins to be expressed at the posterior of each engrailed stripe, leading to the activation of EGFR signaling in and around its domain of expression. The zone of Rhomboid influence corresponds roughly to where denticles form at the end of embryogenesis, while Wingless signaling is associated with the absence of denticles (bald cuticle). PS, parasegment boundary; SB, segment boundary.

  33. An Active HRP-Wingless Fusion Protein Sp:signal peptide L: linker • HRP-horseradish peroxidase • Reacts with DAB— • electron dense deposit • Unaffected by fixation • Stability—late endosome • and lysosome Arm-Gal4—naked D. Wg Gal4 rescues wg mutant E.Secreted –acts at a distance

  34. GAL4/UAS system

  35. HRP-Wingless Is Preferentially Targeted to Lysosomes at the Posterior In all panels, HRP-Wingless is expressed under the control of the engrailed-gal4 driver

  36. EGFR Signaling Modulates the Rate of Wingless Degradation Red-engrailed Green-Wingless Hg prevent of Wg spread enGAL4 UAS-nucLacZ; UAS-Wingless Pointed too

  37. Regulated Endocytic Routing ModulatesWingless Signaling in Drosophila Embryos

  38. Dpp forms a DV gradient in the Drosophila embryo Gene & Development, 16, 1314, 2002

  39. Models of morphogen gradient formation transcytosis

  40. Distinct endocytic pathways regulate TGF-b receptor signaling and turnover Smad7-smurf ubiquitylated Receptor and smad 1 /2 Nat. Cell Biol.5,410-421

  41. Endocytosis in tissue remodeling and cell migration

  42. Endocytosis of E-cadherin regulate epithelial polarity and tissue remodeling Epithelial-mesenchymal transition HGF activate Src phosphorylates E-cadherin E3 ligase binding ubiquitination (lysosomal degradation) Curr. Opin. Cell Biol. 18,407-415, 2006

  43. Specific cell boundaries are established : protocadherin recruits exocyst component Sec5 Curr. Opin. Cell Biol. 18,407-415, 2006

  44. Endocytosis regulates cell motility and invasiveness • Rac • Rho at raft controlled by integrin signal • MT1-MMP • Src phosphorylates endophilin A2, prevent endocytosis of MT1-MMP Cell, 124, 897-900, 2006

  45. LDLR and Wnt signaling Coaxing the LDL Receptor Family into the Fold Cell, Vol. 112, 289–292, February 7, 2003, Joachim Herz* and Peter Marschang* LRP6 holds the key to the entry of anthrax toxin Cell, 124, 1119, 2006

  46. Structural organization of LDL receptor family members VLDLR and ApoER2 Bind to Reelin Adaptor Dab1 recruits src and PI3 kinase to cause tau Phosphorylation Alzheimer’s disease LDL receptor-related protein LRP6-Wnt3 Arrow-Wg Cell, 112, 289, 2003

  47. D-frizzled 2-receptor Arm/b-catenin interacts with Cadherin Colon cancer-APC Oncogenic-c-myc Developmental Biology by Scott F. Gilbert Sinauer Associates, publishers Six edition, 2000

  48. The LDL Receptor family biosynthesis and secretion Coreceptor Fly: Arrow-Wg (Frizzled) Mouse: LRP6-Wnt3 Mesd: mesoderm development Wnt3 deficient mutant In ER, a molecular chaperone – Protein Folding In Mesd mutant-LRP fail to reach the cell surface –insoluble, aggregate in the ER Cell, 112, 289, 2003

  49. LRP6 holds the key to the entry of Anthrax Toxin 炭疽桿菌 Toxin: LF-lethal factor:metalloproteinase (cleaves MAKK) EF-edema factor:adenylate cyclase PA-protective antigen (import) Prepore to pore (low pH) furin Cell, 124, 1119-1121, 2006 Antisense screen for LRP6

  50. Endocytosis and signaling transduction 1. When cell biology meets development: Endocytic regulation of signaling pathway Genes & Development 16-1314-1336, 2002 2. Signal transduction and endocytosis: close encounters of many kinds Nat. Rev. Mol. Cell Biol. 3, 600-614, 2002 3. Endosome dynamics during development Curr. Opin. Cell Biol. 18, 407-415, 2006 4. Endocytosis conducts the cell signaling orchestra Cell 124,897-900, 2006

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