Lecture 11

• Vesicular Trafficking: Lysosomes;

• endocytosis and exocytosis

pH is one of the mechanismsfor protecting contents in the cytosol

Vacuolar H+ ATPase:

LysosomesEndosomesSelected compartments ofGolgi ApparatusTransport vesiclesSecreted vesicles

All major classes of macromolecules are degradedin lysosomes

Lysosomal glycoproteinsare unusually highly glycosylated

40 hydrolytic enzymes

Lysosomes are highly heterogeneousShape and size

But all have acid hydrolases

The “stomach” of a cell

Stained for acid phosphatase:Phosphatase substrate

Huge vacuoles of plant cells

Related to lysosomes

Storage organelle

Degradative compartments

Turgor pressure

Homeostatic device

RubberOpiumGarlicProteinsPigmentsNoxious molecules

Three pathways to degradation in lysosomes

pH 6.0 pH 5.0

The structure of mannose 6-phosphate on a lysosomal enzyme

A unique marker for lysosomal proteins

The transport of newly synthesized lysosomal hydrolases to lysosomes

M6P receptor

pH6.5-6.7pH6.0

Recycling of M6P

Lysosomal storage diseases

Some lysosomes may undergo exocytosis

A signal patch in the hydrolase polypeptide chain provides the cue for M6P addition

Defects in the GlcNac phosphotransferase cause alysosomal storage disease in humans, inclusion-cellDisease (I-cell disease)

N-acetylglucosamine

Phagocytosis by a macrophate Phagocytosis by a neutrophil

Pinocytosis: cell drinking; Phagocytosis: cell eating

pseudopods

The formation of clathrin-coated vesiclesfrom the plasma membrane

Caveolae in the plasma membrane of fibroblast

Lipid rafts

caveolin

A low-density lipoprotein(LDL) particle

Receptor-mediated Endocytosis

Example: cholesterol uptake

Atherosclerotic plaques;When uptake is blocked

Atherosclerosis

3,000 Kd1500 cholesterolMolecules800 phospholipid500 unesterified cholesterol

500 Kd

Cholesterol estersAre hydrolyzed to Free choleterol in lysosomes

Normal and mutant LDL receptor

Coronary artery disease

Common endocytic signal: YXXbinding to adaptin)But LDL receptor:Asn-Pro-Val-Tyr

Possible fates for transmembrane receptor proteins

Different Rabsassociate withearly and lateendosomes

Recycling:LDL receptor

Receptor-mediated endocytosis of LDL

Sorting of transferrin (red)and opioid receptors (green)in the recycling endosomes

Transferrin receptor recycles withits ligand

DifferentRabs

Opioid receptors and EGF receptorsAre not recycled but degraded inLysosomes along with ligand: receptorDown-regulations

Some early endosomes Migrate slowly along MTToward the cell interiorAnd pinch off Vesicles to form MVBs

MVBs may fuse with a lateendosomal compartment orthey fuse with each otherto become late endosome

Lysosomes form when vesiclesfrom trans Golgi network fuse with late endosomes

Receptors and their ligandsare fully accessible to digestive enzymes in MVBs

Ubiquitin tagging facilitatethe uptake of receptors into endocytic vesicles andand sorting into theinternal membrane vesiclesof MVBs

Transcytosis: transferring macromolecules across ephithelial cell sheets

Remember transcelluartransport of glucose?

Early endosome torecycling endosome

Receptors havesorting signals

A newborn rat obtains antibodies from its mother’s milk

Exit of membrane proteins from the recycling endosomes can be regulated

Two early endosomal compartments and one common late endosomalcompartment in epethilial cells

Exocytosis: fusion of vesicleswith the plasma membrane

Two mechanisms of secretion

Three pathways of protein sorting in the trans Golgi network

Secretory proteins formselective aggregates

Secretory proteins becomehighly concentrated:aggregation and membraneretrieval

Proteins are often proteolytically processed during the formation of secretory vesicles

Size of the final products

Activity

In different cells

A prohormone

Regulated exocytosis can be a localized response of the plamsa membrane and its underlying cytoplasm

Mast cellsecreting histamineall over thecell surface whenon a solution ofsoluble stimulant

Localized exocytosis whenstimulant is presented from asolid bead

Two types of polarized cells

Polarized cells direct proteinsfrom the trans Golginetwork to the appropriatedomain of the plasma membrane

Two ways of sorting plasma membrane proteins

Live cellsGut epithelial cells

Lipid rafts in the trans Golgi networkGlycosphingolipids and cholesterol form rafts in the lipid bilayer

May mediate sorting of glycosphingolipids and GPI-anchored proteinsTo the apical plasma membrane

50 nm

The formation of synaptic vesicles

Summary

1. Lysosomes are where most of the intracellular degradationoccurs. Formation of lysosomes, sorting into lysosomes;

2. Phagocytosis, pinocytosis, receptor-mediated endocytosis,early endosomes, late endosomes, recycling endosomes;

3. Two types of exocytosis, sorting during exocytosis, synapticvesicles.