Endocytosis Flashcards
(46 cards)
1
Q
What is Clathrin-Mediated Endocytosis?
A
- receptor-mediated endocytosis
- how cells bring in specific molecules (nutrients or receptors) from the cell surface by forming vesicles.
2
Q
What are the key players in CME?
A
- Clathrin Triskelion
-made of 3 heavy chains and 3 light chains
- forms a lattice coat on the membrane
- helps curve the membrane to form a pit - Adaptor Proteins (AP-2)
- helps clathrin bind to the membrane
- works on plasma membrane
- recruits cargo receptors - cargo receptors
- concentrate the molecules (cargo) the cell wants to take in - Dynamin
- GTPase that cuts off the vesicle from the membrane
- forms a ring around the neck of the vesicle
- Uses GTP hydrolysis to pinch and release the vesicle
3
Q
What are the 4 stages of CME?
A
- Initiation
- Clathrin and AP-2 gather at the membrane
- Cargo receptors are recruited - Propagation
- The clathrin-coated pit begins to invaginate (fold inward) - Budding
- Pit keeps folding inward
- Dynamin cuts the vesicle from the membrane - Uncoating
- Clathrin coat is removed
- Vesicle is free to fuse with early endosomes.
4
Q
What is Caveolin-Mediated Endocytosis?
A
- way for cells to bring in molecules using small, flask-shaped pits in the membrane called caveolae (means “little caves”).
- based on lipid rafts and the protein caveolin.
5
Q
What are the key features of Caveolae?
A
- small invaginations in the plasma membrane
- coated with caveolin-1 (integral membrane protein) -> binds to cholesterol, helps form and shape caveolae.
- rich in cholesterol and sphingolipids
- formed in the golgi and the moved to cell surface.
- lipid rafts:
- specialized membrane areas rich in cholesterol and sphingolipids.
- favor the localization of specific proteins, including GPI-anchored proteins, signaling molecules, and transmembrane proteins
6
Q
What is the function of caveolae?
A
- Endocytosis (Caveolar-dependent endocytosis)
- Caveolae can pinch off with help from dynamin
- They form caveosomes (neutral pH compartments, different from endosomes)
- Used for transcytosis, especially in endothelial cells (e.g., transporting albumin across capillaries) - Transcytosis of Albumin
- Albumin binds to gp60 receptor
- Triggers endocytosis and movement across the endothelial cell
- Involves dynamin - Signalling Platforms
- In many cells, caveolae do not endocytose, they stay on the surface
- Act as hubs for signaling because many signaling proteins prefer raft-like regions
- Some signaling proteins bind to a scaffolding domain on caveolin-1 to concentrate locally
- Examples of molecules enriched here: - Lipids: Sphingomyelin, ceramide, cholesterol
- Receptors: EGF receptor, PDGF receptor
- Transducers: PI3K, adenylyl cyclase
7
Q
What is Macropinocytosis?
A
- used to internalize fluid, particles, and membrane — no need for receptors
- nonspecific
- happens in all cells
8
Q
How does macropinocytosis work?
A
- membrane forms projections:
- Lamellipodia (sheet-like structures)
- Circular ruffles
- Blebs (bulging part of the membrane)
- These projections fold back and trap fluid in large vesicles called macropinosomes.
- Macropinosomes are large vesicles, over 500 nm in diameter.
- After internalization, the cargo is usually sent to lysosomes for breakdown.
9
Q
What is CLIC-GEEC?
A
- special type of endocytosis that does not use clathrin or dynamin.
- involves tubular structures, not coated pits of vesicles.
- mainly happen at leading edge of migrating cells.
- Takes in fluid, membrane, and GPI-anchored proteins
- Also internalizes parts of the lipid raft membrane
10
Q
How does CLIC-GEEC work?
A
- CLICs (Clathrin-Independent Carriers):
- These are tubular, uncoated vesicles that first carry the material into the cell
- They are small and mobile - GEECs (GPI-anchored protein Enriched Early Endosomal Compartments):
- Several CLICs fuse together to form a larger vesicle called a GEEC.
- GEECs contain GPI-anchored proteins and continue moving through the cell - After a few minutes, GEECs fuse with early endosomes (part of the regular endocytic pathway).
- GEECs are more acidic than regular early endosomes.
11
Q
What is Phagocytosis?
A
- Phagocytosis is a specialized form of endocytosis where certain immune cells engulf and digest large particles (usually >500 nm), like bacteria or dead cells.
12
Q
Which cells perform phagocytosis?
A
- only specialized cells:
1. macrophages
2. neutrophils
3. dendritic cells
all part of innate immune system
13
Q
how does phagocytosis work?
A
- Receptor-dependent recognition:
- The cargo (e.g., a bacterium) is often tagged with antibodies (Ab).
- These antibodies bind to Fc receptors on the phagocyte surface. - Actin rearrangement:
- Binding of the Fc receptor triggers Rho GTPases to reorganize the actin cytoskeleton.
- This causes the cell to form pseudopods (arm-like extensions). - Engulfment:
- The pseudopods wrap around the cargo in a zipper-like fashion.
- The cell then engulfs the cargo, forming a vesicle called a phagosome.
14
Q
Explain Phagosome Maturation
A
- once the cargo is inside:
- The phagosome fuses first with early endosomes, then with late endosomes.
- It gradually acquires endosomal proteins, especially LAMPs (lysosome-associated membrane proteins).
- When enough LAMPs are present, the phagosome fuses with lysosomes, forming a phagolysosome.
- Inside the phagolysosome, the cargo is broken down and degraded.
15
Q
How can pathogens escape destruction?
A
- Convert the phagosome into:
- An autophagosome or an ER-like vesicle
- A hybrid autophagosome–RAB7-positive phagosome
- A rough ER-like compartment - Inhibit phagosome maturation:
- Block the transition at the RAB5-positive stage (early endosomal stage)
- Interfere with RAB7 function, preventing late-stage maturation.
16
Q
What are the major endocytic compartments?
A
- Endocytic vesicles
- Early endosomes
- Recycling endosomes
- Later endosomes/multivesicular bodies
- Lysosomes
17
Q
Describe the pathway of endocytosis of LDL (Low-density lipoprotein)?
A
- The LDL particle, which carries cholesterol, binds to its specific LDL receptor on the plasma membrane of the cell.
- The LDL-receptor complex is then internalized into the cell through clathrin-mediated endocytosis. (clathrin helps form a clathrin-coated vesicle that brings the LDL particle into the cell)
- Once inside cell, clathrin coat removed, the vesicles becomes an early endosome. Environment inside is mildly acidic, LDL particle dissociates from its receptor.
- LDL receptor is recycled, returned to the plasma membrane, used again to bind additional LDL particles.
- LDL particle (without its receptor) is transported to a lysosome, where it is degraded. The breakdown products of LDL include cholesterol, amino acids, and fatty acids, which are used by the cell for various functions, like membrane synthesis and energy production.
18
Q
What is the function of ARH protein?
A
- specifically involved in LDL receptor-mediated endocytosis in certain cell types, particularly in liver cells (hepatocytes), where LDL uptake is crucial for regulating cholesterol levels.
- It interacts with the LDL receptor and helps to mediate the internalization of the receptor-LDL complex in clathrin-mediated endocytosis.
19
Q
what is EGF and EGFR?
A
EGF: Epidermal Growth Factor (signalling molecule that binds to EGFR)
EGFR: receptor tyrosine kinase that activates cell signaling pathways involved in growth, differentiation, and survival when EGF binds.
20
Q
What is the pathway of endocytosis of EGF and EGFR?
A
- EGF binds to EGFR at the plasma membrane.
- This causes EGFR to dimerize and become activated (autophosphorylation of tyrosine residues on its intracellular tail).
- The binding of EGF to EGFR triggers endocytosis of the EGF–EGFR complex.
- this internalization typically occurs via clathrin-mediated endocytosis, but non-clathrin pathways can also be used depending on conditions (e.g., high EGF concentrations).
- Once inside the cell, the EGF–EGFR complex is delivered to early endosomes.
- EGF remains bound to EGFR in the acidic environment of early endosomes—the complex is stable here.
- EGFR is inactivated not by dissociation, but by being sorted into intraluminal vesicles (ILVs) inside the early endosome.
- These ILVs form part of multivesicular bodies (MVBs).
- This sequestration removes EGFR from the cytoplasm, preventing further signaling even though the receptor is still present in the cell.
- After being packaged into ILVs, the MVB fuses with lysosomes, and the EGF–EGFR complex is degraded.
- This ensures that EGFR signaling is terminated, and receptor levels are downregulated.
21
Q
Why is Iron transport Important?
A
- essential for many cellular processing, especially in cells making lots of hemoglobin (like reticulocytes, the precursors to red blood cells).
- Iron in blood travels as Fe(III) (ferric iron), bound to a glycoprotein called transferrin.
22
Q
What are the two key forms of transferrin?
A
- Holo-transferrin: Transferrin bound to iron (Fe(III)).
- Apo-transferrin: Transferrin with no bound iron.
23
Q
Describe the pathway of the endocytosis of Transferrin
A
- Iron Transport in Blood:
- Fe(III) binds to transferrin, forming holo-transferrin.
- This is the main form of iron that circulates in the blood. - Two holo-transferrins bind to transferrin receptors on the surface of reticulocytes (immature red blood cells).
- The holo-transferrin–receptor complex is internalized by clathrin-dependent endocytosis.
- In the acidic environment of the early endosome:
- Fe(III) is released from transferrin.
- STEAP3, an enzyme in the endosome, reduces Fe(III) to Fe(II) (ferrous iron).
- Fe(II) is then transported into the cytoplasm via DMT1 (Divalent Metal Transporter 1). - The apo-transferrin–receptor complex (now iron-free) is recycled back to the plasma membrane.
- At neutral pH (outside the cell), apo-transferrin dissociates from the receptor, making it available to bind iron again.
24
Q
Describe the pathway of transcytosis of immunoglobulins (IgA) (across epithelial cells)
A
- occurs mainly in mucosal tissue
- plasma cells secrete dimeric polymeric IgA (plgA) at the basolateral sice.
- pIgA binds to IgA receptors on epithelial cells
- The complex is clathrin-dependent endocytosed
- Vesicles move the complex across the cell to the apical side
- At the apical membrane, pIgA is cleaved:
1. Transmembrane part stays in the cell
2. Extracellular part + IgA = secretory IgA (sIgA) - sIgA is released into the lumen
- Transcytosis links endocytosis + exocytosis, allowing movement across epithelial barriers
25
What is Transcytosis?
Process where molecules are:
1. Endocytosed (taken in) on one side of a polarized cell (e.g., epithelial cell),
2. Transported across the cell inside vesicles, and
3. Exocytosed (released) on the opposite side.
commonly used in epithelial and endothelial cells to move cargo from the basolateral side to the apical side or vice versa.
26
Why is Transcytosis of IgA Important?
- IgA (immunoglobulin A) is a key antibody found in mucosal tissues (e.g., gut, lungs, reproductive tract).
- It provides immune protection at mucosal surfaces by neutralizing pathogens in secretions like saliva, tears, and mucus.
27
Describe the process of Transcytosis of IgA
1.
- Plasma cells in mucosal tissue secrete dimeric IgA.
- This polymeric IgA (pIgA) binds to the polymeric Ig receptor (pIgR) located on the basolateral surface of epithelial cells.
2.
- pIgA–pIgR complex is internalized via clathrin-dependent endocytosis.
3. The complex is transported in vesicles across the epithelial cell toward the apical surface (facing the lumen).
4.
- at the apical membrane, plgR is cleaved.
- extracellular portion of the receptor remains attached to the IgA.
- This cleaved complex is now called secretory IgA (sIgA).
5.
- slgA is released into the lumen of the mucosal surface, where it can neutralize pathogens.
28
Explain transport of
albumin by caveolin
- Albumin is transported across endothelial cells by caveolae (small plasma membrane invaginations).
- gp60 receptor on the luminal side binds albumin.
- Binding activates Src kinases, triggering caveolin-1–dependent vesicle formation.
- Albumin is internalized via caveolae (clathrin-independent, dynamin-dependent).
- Vesicles move across the cell to the basolateral side.
- Exocytosis releases albumin into the tissue.
29
Explain folate uptake via the GEEC pathway.
- folate binds to the folate receptor, which is a GPI-anchored protein.
- folate receptor is located in lipid rafts.
- this receptor-folate complex is internalized through the CLIC-GEEC pathway.
- The CLIC-GEEC pathway forms tubular, non-coated vesicles.
- Internalized vesicles fuse to form GEECs, which later merge with early endosomes.
- Folate is delivered into the cell for metabolic use.
30
What is the zipper model of phagocytosis?
- describes how a phagocyte engulfs a particle step by step in a controlled, receptor-dependent manner.
1. Ligands (antibodies) on the surface of a particle bind to receptors (Fc receptors) on the phagocyte's membrane.
2. this binding triggers localized actin polymerization, which extends the membrane around the particle.
3. As more receptors engage sequentially around the particle, the membrane "zippers" up around it—like closing a zipper.
4. The pseudopods meet and fuse at the top, fully enclosing the particle into a phagosome.
31
What is autophagy?
- Cargo acquired from within cell
- Catabolic process for reusing components rather than de novo
32
What are the purposes of autophagy?
- Obtain nutrients under starvation conditions
- Turnover of defective organelles
- Removal of protein aggregates from cytoplasm
- Removal of bacteria/infectious agents
- Specialized purposes (e.g. liberation of cholesterol from lipid droplets)
33
What are the three kinds of autophagy?
1. Chaperone-mediated autophagy
2. Microautophagy
3. Macroautophagy
34
what is chaperon-mediated autophagy?
- target: Single, specific proteins with a KFERQ-like motif.
process:
- KFERQ-tagged proteins bind to Hsc-70 (a chaperone).
- The complex is delivered to the lysosome.
- Protein is translocated across the lysosomal membrane via LAMP-2A.
selective, not bulk degradation.
35
What is microautophagy?
- target: small volumes of cytoplasm or surface proteins.
process:
- cytosolic proteins and/or Hsc-70 bind to phosphatidylserine on the surface of late endosomes.
- the membrane invaginates and buds inward, capturing material into internal vesicles.
- non-selective or semi-selective, direct lysosomal engulfment.
36
how does macroautophagy 'autophagy' work?
- target: large structures like protein aggregates, damaged organelles (e.g., mitochondria -> mitophagy)
37
what is the process of macroautophagy?
1.
- formation of phagophore (isolation membrane)
- may originate from ER, mitochondria, or other organelles.
2. LC3 processing:
- LC3-I: Soluble form, generated by cleavage of pro-LC3.
- LC3-II: LC3-I is conjugated to PE (phosphatidylethanolamine) by Atg12-Atg5-Atg16L complex → becomes membrane-bound LC3-II.
- LC3-II integrates into the phagophore membrane and helps with membrane expansion and closure.
3. Cargo selection:
- Damaged material often ubiquitinated.
- Autophagy receptor p62:
binds to ubiquitin on cargo
binds to LC3 on the phagophore membrane
helps link cargo to autophagosome
4.
- Phagophore expands, sealing around the cargo to form a double-membrane autophagosome.
5.
- Autophagosome fuses with endosomes, becoming more endosome-like (early → late).
- Finally fuses with a lysosome, forming an autolysosome.
- Contents are degraded, and LC3-II on the outer membrane is recycled.
38
What are Rab Proteins?
- small GTPases, part of the Ras superfamily.
- regulate vesicle trafficking (e.g., targeting, tethering, fusion)
- they are molecular switches
- active (on) when bound to GTP
- inactive (OFF) when bound to GDP
- have fatty acid modifications at their C-terminus to anchor into membranes (only in GTP-bound state)
39
When Rab is in a GTP-bound state, what effector proteins does it bind?
Tethering factors
SNARE proteins
Motor proteins (e.g., kinesin, dynein)
Lipid-modifying enzymes
40
How does the Rab GTPase Cycle work?
1. Activation
GEF exchanges GDP for GTP.
Rab becomes active and associated with membranes.
2.
Active Rab-GTP binds to specific effector proteins for vesicle transport steps.
3. Inactivation.
- GAP stimulates GTP hydrolysis -> Rab becomes inactive (Rab-GTP).
- Rab-GDP detaches from effectors.
4. Recycling.
- GDP extracts Rab-GDP from the membrane and holds it in the cytosol.
- Rab is then recycled for another round.
41
What are effectors for Rab1?
- p115 (vesicle tethering)
- GM130 (cis-golgi matrix protein)
- TRAPP complex (GEF that also acts as a tether)
- COPII and COPI vesicle regulators
42
What are effectors for Rab5?
- EEA1 (Early Endosome Antigen 1): filamentous tether that aids in early endosome fusion. Binds Rab5 and P13P.
- Rabenosyn-5: involved in endosomal trafficking and recycling.
- PI 3-kinase: produces PI3P, which is required for recruitment of EEA1/Rabenosyn-5.
- SNARE proteins: syntaxin 6 and syntaxin 13 via EEA1 and Rabenosyn-5
43
What are effectors for Rab7?
- RILP: connects late endosomes to dynein-dynactin for minus-end microtubule transport
- ORP1L: cholesterol sensor
- βIII spectrin: anchors endosome to the cytoskeleton
- FYCO1: links vesicles to kinesin for plus-end movement
- HOPS complex: tethers vesicles for fusion between late endosomes and lysosomes.
44
Where is Rab5 localized?
- early endosomes and phagosomes.
it is a key regulator of early endocytic trafficking, determines endosome identity, and orchestrates endosome fusion and sorting.
45
Where is Rab7 localized?
- late endosomes, lysosomes, autophagosomes.
- drives maturation of endosomes.
- controls cargo delivery to lysosomes.
- regulates positioning and fusion of late endocytic compartments.
46
How does a Rab5-positive early endosome convert into a Rab7-positive late endosome?
1. Rab5 activation on early endosome recruits a Rab7 GEF to the membrane.
2. Rab7 GEF catalyzes exchange of GDP for GTP on Rab7, activating it.
3. Activated Rab7 then recruits TBC2, a Rab5 GAP, which inactivates Rab5 by promoting GTP hydrolysis.
4. Inactive Rab5-GDP dissociates from the membrane.
5. Simultaneously, the complex that Rab5 recruited displaces Rabex5 (Rab5’s own GEF), accelerating Rab5 removal.
6. The endosome now contains Rab7, and not Rab5—defining it as a late endosome.
