Week 5 + 6 Intracellular Compartments and Endomembrane System Flashcards
What is purpose of ER?
Synthesize most lipids, proteins (for distribution to many organelles and to plasma membrane)
What is the purpose of the Golgi apparatus?
Modify, sort and package proteins and lipids for secretion or organelle delivery
What is purpose of lysosomes?
Intracellular degredation
What is purpose of endosomes?
Sorting of endocytosed materials
What is purpose of peroxisomes?
Oxidative breakdown of toxic molecules
What is protein sorting?
How proteins reach target destinations
What are the 3 ways proteins are transported into organelles?
- Nuclear pores
- Transport across membranes
- Transport by vesicles.
What are signal peptides/patches?
Proteins are targeted to specific organelles via amino acid sequences that act like locator. Distinct signal sequences for each organelle
How are signal sequences studied?
via engineering proteins w/attached sequence and identifying where they locate.
Describe the structure of nuclear pore.
Complex of about 30 proteins - act like gate - arranged to allow small molecules through but selectively control large ones - nuclear basket extends into nucleus and cytosolic fibrils extend out into cytosol.
How do proteins move through nuclear pores?
Proteins are directed to nucleus by nuclear localization signal. Nuclear import receptors bind the signal sequence and guide protein through pore.
Describe process of energy for movement through the pores.
The energy required is derive from GTP hydrolysis, aided by GTPase called Ran.
Nuclear import receptor binds Ran-GTP
When GTP is hydrolyzed, it falls off and a protein with nuclear localization signal can bind.
This complex can then move through nuclear pore
Inside nucleus, Ran-GTP binds to receptor, and nuclear protein is delivered to nucleus.
Ran cycles between GDP and GTP with aid of Ran-GAP and Ran-GEF.
Where do most mitochondrial proteins come from?
Most mitochondrial proteins are made in cytosol from nuclear genes and translocated to mitochondrial matrix. Mitochondrial DNA only encodes for some proteins.
Describe process of getting proteins into mitochondria from cytoplasm
- Mitochondrial proteins made in cytosol
- Protein with signal sequence binds to import receptor protein in outer membrane
- Protein moves into intermembrane space with aid of translocator in outer membrane
- Complex diffuses through membrane and meets translocator in inner membrane
- Protein moves into mitochondrial matrix and signal peptide is cleaved
- Also requires chaperone proteins and ATP
Where do ribosomes come from?
Common pool of ribosomes used to synthesize all proteins. mRNA translates on free ribosomes. Ribosomes translating proteins with ER signal sequence are directed to it, otherwise remain in cytosol.
Describe how proteins enter the ER
An SRP binds to signal sequence and ribosome, pausing translation and then interacts with SRP receptor in ER membrane. The SRP is released and ribosome passes from SRP receptor to protein translocator in ER membrane. Protein synth. resumes and translocator starts to transfer growing polypeptide across lipid bilayer.
Describe how proteins enter fully enter the ER lumen
Proteins enter ER while being synthesized by ribosome. Soluble proteins cross ER membrane and released into lumen. Signal pep is cleaved by signal peptidase.
Describe how mature single-pass transmembrane ER proteins are made
Stop transfer sequences can stop the translocation of proteins, resulting in transmembrane protein in lipid bilayer. After the stop transfer sequence enters, the rest of protein is translated in cytosol.
Describe how double-pass transmembrane proteins in ER are made.
A double-pass transmembrane protein will have internal ER signal sequence that acts as a start-transfer sequence. The start and stop sequences don’t get cleaved and remain in part of the protein.
What is the endomembrane system?
A collection of organelles connected by vesicle movement.
what is the major secretory pathway?
ER > Golgi > plasma membrane - some vesicles move from Golgi to endosomes to lysosomes
What is major endocytic pathway?
Plasma membrane > lysosomes
How are vesicles created?
Vesicles are pinched off an existing membrane with help of proteins that form a coat around it. Proteins that need to be moved have sequence that allows them to bind to receptors in membrane and are carried along as vesicle pinches off.
Describe steps of budding with clathrin proteins
Cargo receptors and cargo are bound by adaptin proteins. Clathrins bind to adaptins and shape budding vesicle. Dynamin and other proteins wrap around neck of bud
- Hydrolysis of GTP bound to dynamin helps pinch off vesicle
- The vesicle is then uncoated and can go on to fuse elsewhere
Describe vesicle fusing
- A vesicle will eventually reach its target membrane
- Tethering proteins on target organelle capture the vesicle by interaction with vesicle-specific Rab proteins
- Vesicle docking to target membrane depends on binding of v-SNAREs and t-SNAREs
- Vesicle and organelle membranes fuse.
What is the purpose of complementary SNAREs?
Complementary SNARES on the vesicle and target membranes bring the structure very close together, squeezing out all water so the phospholipids can interact and fusion can occur with aid of additional proteins. SNAREs can then by recycled.
Describe the secretory path in the ER.
Proteins carried through this path are synthesized co-translationally in ER - many covalently modified in ER after synth.
Short branched oligosaccharides are added to Asp residues in process of glycosylation. Further modification in ER then Golgi. Proteins in ER also form disulphide bonds which aids in their proper folding and stability.
What happens during secretory path in ER if proteins are left behind?
Some proteins are retained in ER (via ER retention sequence). Proteins that don’t fold properly don’t exit ER but are retained with aid of chaperones - may be exported to cytoplasm to be degraded.
If too many misfolded proteins - the unfolded protein response is generated - more chaperones, ER and associated proteins are produced. If ER is overwhelmed by amount - may trigger apoptosis.
Describe secretory path in the Golgi
Golgi’s have 3-20 cisternae. Proteins move from one to next. Cisternae may also mature and move towards trans face.
Vesicles that have exited from ER move to cis face of Golgi and fuse. Are sequentially processed with addition of more complex oligosaccharides, moving through stack and exit on trans face to head to destinations.
Describe secretion phase (and its two paths) of secretory path
Constitutive exocytosis pathway - provides steady stream of proteins and lipids to plasma membrane and cell exterior - no signal sequence needed
Regulated exocytosis pathway - operates in secretory cells - large amounts of required substance is stored in vesicles and released in response to extracellular signal.
How is amount of phospholipid bilayer retained?
Amount of phospholipid bilayer leaving cell via exocytosis is balanced by that entering the cell through endocytosis
What is pinocytosis vs. phagocytosis
Fluids/small molecules = pinocytosis. Large particles = phagocytosis
Describe pinocytosis
vesicles are pinched off with aid of coat proteins ie. clathrin. Material in fluid enters cell too. Vesicle fuses with endosomes and lysosomes and material inside is degraded. Specific material can be taken up by cells through receptor-mediated endocytosis - ie. LDL
Describe LDL cycle.
LDL binds to LDL receptors and is internalized in clathrin-coated vesicles. The vesicles lose their coat and then fuse with endosomes - where the LDL dissociates from receptors because of acidity. The LDL ends up in lysosomes where is degraded to release free cholesterol. LDL receptors are returned to plasma membrane via transport vesicles to be recycled.
How do some viruses enter cells?
Viral entry into cells by receptor-mediated endocytosis. Clathrins.
What happens to vesicles once in cell? What are the three main things that happen?
Vesicles fuse with early endosomes which become late endosomes. The interior is acidic, causing receptors to release cargo. The receptors can be recycled to degraded in lysosome.
1. Recycling 2. degradation 3. transcytosis
Where does most of the endocytosed material end up?
Much of the material endocytosed ends up in lysosomes where it is digested and degraded. Lysosomes have large variety of hydrolytic enzymes that break down macromolecules and function best at low pH. Digested material is transported out of lysosome into cytoplasm for reuse
What are the different paths for materials destined for degredation in lysosomes?
Materials destined for degradation in lysosomes follow diff paths to get to lysosomes. Cells can digest old or worn out organelles or groups of proteins in process of autophagy.
