Intracellular Compartments and Protein Sorting

1. Intracellular Compartments and Protein Sorting Compartmentalization of Cells Pages 695-712

2. Proteins Characterize Organelles -They catalyze the reactions that occur in each organelle -They selectively transport small molecules in and out of its interior -They serve as organelle-specific surface markers that direct new deliveries of proteins and lipids to the appropriate organelle

3. Major Intracellular Compartments

4. Organelle Function Nucleus – Contains the genome and is the site for DNA and RNA synthesis Endoplasmic Reticulum – Produces most of the lipid for the rest of the cell -Functions in transport of proteins to the Golgi -Functions as a store for calcium Golgi Apparatus – Receives proteins and lipids from the ER and dispatches them to several destinations Mitochondria – Generates most of the ATP used by cells Lysosomes – Degrades intracellular organelles and macromolecules taken in from outside the cell Endosomes – Contain material taken in from outside the cell Peroxisomes – Contain enzymes involved in oxidative reactions

5. TABLE 12–1 Relative Volumes Occupied by the Major Intracellular Compartments in a Liver Cell (Hepatocyte) INTRACELLULAR COMPARTMENT PERCENTAGE OF TOTAL CELL VOLUME Cytosol 54 Mitochondria 22 Rough ER cisternae 9 Smooth ER cisternae plus Golgi cisternae 6 Nucleus 6 Peroxisomes 1 Lysosomes 1 Endosomes 1

7. EM of a Liver Cell -Organelles often have characteristic positions in the cytosol depending on interactions with the cytoskeleton. -The ER and Golgi depend on the microtubule array. -Eucaryotic cells are 10-20 times larger linearly, but 1,000-10,000 times greater in volume.

8. Specialization of Membrane Function Development of Plastids

9. Evolution of Cell Nucleus and ER

10. Evolution of Mitochondria

11. Relationships between Compartments

12. Protein Traffic Map

13. Types of Protein Transport 1. Gated transport – Protein traffic between the nucleus and cytosol occurs between topographically equivalent spaces, which are connected through the nuclear pore complexes 2. Transmembrane transport – Membrane-bound protein translocators directly transport specific proteins across a membrane from the cytosol into a space that is topologically distinct 3. Vesicular transport – Membrane-enclosed transport intermediates ferry proteins from one compartment to another

14. Vesicle Transport

15. Types of Sorting Signals Each type of protein transfer is usually guided by sorting signals in the transported protein that are recognized by receptors. Most receptors recognize classes of proteins rather than just one protein. 15-60 AA

16. Signal Sequences

17. Organelles Cannot be Constructed from Scratch During division, cells must duplicate their organelles They do it by enlarging existing organelles by incorporating new molecules into them and then dividing Each daughter cell inherits their organelles from their mother

19. Nuclear Envelope Defines the nuclear compartment Inner membrane contains specific proteins that interact with chromatin and the nuclear lamina Outer membrane is continuous with the membrane on the ER

20. Nuclear Pore Complexes -Composed of over 30 different proteins called nucleoporins -The more active the nucleus is in transcription the more complexes the envelope contains, typically there’s 3000-4000 Nuclear side

21. Nuclear Pore Complex 500 macromolecules per second

22. Free Diffusion through Nuclear Pores 9 nm diameter limit for free diffusion, but up to 39 nm can be brought through by transport receptors

23. Function of a NLS NLS is rich in positively charged amino acids, lysine and arginine Nuclear proteins can be transported through a pore complex while they are in a fully folded conformation

24. Visualizing Active Transport

25. Nuclear Import Receptors The import receptors are soluble cytosolic proteins that bind both the NLS on the protein to be transported and to nucleoporins. Many of the nucleoporins contain phenylalanine-glycine (FG)-repeats that serve as binding sites for the import receptors

26. Nuclear Export Relies on nuclear export signals on proteins that are bound by nuclear export receptors Both types of receptors belong to the family of nuclear transport receptors In yeast there are 14 genes in this family, many more in humans A single pore complex conducts traffic in both directions Import into nucleus -Pro-Pro-Lys-Lys-Lys-Arg-Lys-Val- Export from nucleus -Leu-Ala-Leu-Lys-Leu-Ala-Gly-Leu-Asp-Ile-

27. Compartmentalization of Ran-GDP and Ran-GTP GAP – GTPase-activating protein GEF – Guanine exchange factor Ran is a GTPase

28. Directionality of Nuclear Transport Ran Binding Protein

29. Model for Cargo Release by Ran-GTP

30. Nuclear Transport in Drosophila -Gene regulatory protein called dorsal stained brown -Expressed in the ventral nuclei

31. Control of Nuclear Import during T-cell Activation NF-AT – Nuclear factor of activated T cells Regulation of nuclear localization is done by phosphorylation

32. The Nuclear Lamina Nuclear lamins - are Intermediate filaments -Gives shape and stability to the nuclear envelope -Interacts with chromatin

33. Assemble/Disassembly of the Nuclear Lamina -Lamin Phosphorylation -NPCs disassemble, disperse, bind nuclear import receptors -Motor proteins are involved with disassembly -Nuclear envelope reassembles around chromosomes