1. Day 38 Announcements Friday, April 13: Junctions, the nucleus: pp. 496- 501, 541-550. –Homework problems 15-3, 17-9 due Monday, April 16: the cell cycle, pp. 586-598. Wednesday, April 18: protein synthesis, pp. 684- 695. Friday, April 20: protein targeting, pp. 695-711. –Homework problems 19-10, 14 due.

2. Day 38 Outline/Objectives A.Cellular junctions 1.Adhesive 2.Tight 3.Gap B.The nucleus and nuclear transport C.The cell cycle: mitotic spindle movements After reading the text, attending lecture, and reviewing lecture notes, you should be able to: Describe the different types of cellular junctions: including adhesive, tight and gap junctions. Solve problems involving the ECM and cell junctions. Discuss how entry and exit from the nucleus is regulated. Discuss the forces and movements during metaphase and anaphase, including types of spindle microtubules and motor proteins.

3. Types of Cellular Junctions (Animal Cells)

4. Adhesive Junctions AdhesionAdhesive JunctionCytoskeletal linkage cell-substratefocal contactsmicrofilaments cell-substratehemidesmosomesintermediate filaments cell-celladhesion beltsmicrofilaments cell-celldesmosomesintermediate filaments cell-substrate adhesion involves integrins cell-cell adhesion involves cadherins

5. Focal Adhesions Cell-substrate adhesion In migrating cells Integrins connected to actin microfilaments via linker proteins. Cells migrate using integrin “feet” on ECM “road.”

6. Hemisdesmosomes Cell-substrate adhesion Bind epithelial cells to the basal lamina. Integrins connected to intermediate filaments via hemidesmosome plaques. Blisters form when skin cell layers detach from ECM.

7. Adhesion Belts and Desmosomes Bind Epithelial Cells, Distribute Forces Cell-cell adhesion Cadherins connected to intermediate filaments via desmosome plaque. Cell-cell adhesion A continuous, contractile zone Cadherin receptors connect to actin microfilaments through linker proteins.

8. Tight Junctions Functions: Seals epithelial cell layer; blocks diffusion of molecules between cells. Blocks lateral diffusion of membrane proteins, maintaining apical- basolateral polarity.

9. Gap Junctions Allow ions, small molecules < 1200 MW through. 6 connexin proteins form connexon pore structure. Functions: –electrical, chemical linkage of cytoplasm in heart, CNS, embryonic cells. –For rapid or synchronous functions, e.g. coordinated heart beat, coordinated tissue behavior during embryogenesis Regulation: –Extracellular signals, e.g. dopamine –Decrease in cytosolic pH closes (?) –Increase in cytosolic Ca2+ closes (seals off damage)

10. The Nucleus Nuclear envelope –Inner, outer MBs –Continuous with ER Nuclear lamins –Intermediate filaments –Structural support Nucleolus –Site of rRNA synthesis, assembly Nuclear pores –Regulate import/export of macromolecules

11. Nuclear Pores About 3000 per nucleus Nuclear pore complex regulates passage of RNAs, proteins. Composed of about 100 polypeptides Nuclear localization signals (8-30 amino acids) in proteins target entry.

12. Evidence for nuclear localization signals 8-30 amino acids at beginning of nuclear proteins. If remove NLS from nuclear protein, it no longer enters nucleus. If attach NLS to cytoplasmic protein, it goes into nucleus. Gold beads > 9 nm in diameter cannot pass through nuclear pores. If attach NLS to gold beads > 9 nm and < 27 nm, then they can pass through nuclear pores.

13. Transport Through Nuclear Pores

14. Active transport through nuclear pores requires importin, exportin, Ran, GTP

15. The Cell Cycle Phases: MMitosis G1Gap 1 SDNA Synthesis G2Gap 2 G0Resting, differentiation G0

16. Mitosis

17. Three Kinds of Spindle Microtubules

18. Kinetochore Microtubules Minus ends of MTs attach to centrosome, plus ends to kinetochore, which attaches to centromere region of DNA. Balance of pulling forces to the poles at metaphase

19. Anaphase Movements Anaphase A: –Signal at M/A transition splits chromosomes, which move to poles. –Kinetochore MTs shorten Anaphase B: –Polar MTs lengthen, push apart –Astral MTs shorten, pull

20. Mitotic Motors Movements due to MT motors at: –Kinetochore (Minus end directed) –Overlap of Polar MTs (Plus end directed) –Motor in cell cortex, connecting to Astral MTs (Minus end directed) MT assembly, disassembly also involved.