Lecture 19- Vesicular Transport of Secretory Proteins (cont.) & Receptor-Mediated Endocytosis

Anterograde transport vesicles

COP II

Retrograde transport vesicles

COP I

formed from large systolic complexes called coatomers, composed of 7 polypeptide subunits

COP I coat

ARF GTPase

COP I

Which vesicles retrieve v-SNARE proteins and components of the ER membrane back to provide the necessary material for additional rounds of vesicle budding from the ER

COP I

Which transport also retrieves missorted ER-resident proteins from the cis-Golgi

COPI-mediated retrograde

Which mutants cannot recycle key membrane proteins back to the rough ER

COP I

Sorting signal for targeting a protein to the ER lumen at C terminal

Lys-Asp-Glu-Leu (or KDEL)

Proteins with “KDEL” signal bind to “KDEL” receptors on

ER and cis-Golgi membrane

KDEL receptor binds more tightly to its ligand at low or high pH

low

Receptor binds to KDEL peptides in the cis-Golgi and release them in ER because the pH of the Golgi is slightly lower or higher than ER

lower

Lys-Lys-X-X at the very end of C- terminus binds to

COP I subunits

Gologi complex is organized into 3 compartments (stacked set of flattened sacs)

cisternae

Each cisternae has different set of enzymes that modify the proteins in different ways

glycosidases and glycosyltransferases

Retrograde transport of COPI vesicles results in progressive maturation of the cisternae from 

cis to medial to trans

Transport of proteins from trans-Golgi to late endosomes and lysosomes is achieved through vesicles coated

Clathrin and Adapter proteins

Purified clathrin molecules have 3-limbed shaped

triskelions

Each triskelions contains

1 clathrin heavy chain and 1 clathrin light chain

What polymerizes to form a polygonal lattice with an intrinsic curvature

Triskelions

When clathrin polymerizes on a donor membrane, it associates with what, which fill the space between the clathrin lattice and the membrane

Adapter Protein complexes

4 different adaptor protein complex

AP1

AP2

AP3

GGA

monomeric adapter complex

GGA

What determine the cargo that gets included in the vesicle by interacting with systolic face of membrane proteins

Adapter proteins

binds to membrane proteins with “Tyr-X-X-𝜙” 𝑠𝑒𝑞𝑢𝑒𝑛𝑐𝑒 (𝜙 − ℎ𝑦𝑑𝑟𝑜𝑝ℎ𝑜𝑏𝑖𝑐 𝑎𝑚𝑖𝑛𝑜𝑎𝑐𝑖𝑑

AP1

binds to membrane proteins with “Asp-X-Leu-Leu” or “Asp-Phe-Gly-X-𝜙” 𝑠𝑒𝑞𝑢𝑒𝑛𝑐𝑒

GGA

What complex vesicles take cargo from trans-Golgi to lysosome (bypass late endosome)

clathrin-AP3

What complex vesicles are involved in endocytosis

clathrin-AP2

required for pinching off clathrin-coated vesicles

cytosolic protein

Dyamin

At the early or later stages of bud formation, dynamin polymerizes around the neck portion of the bud then hydrolyzes GTP

later

Membrane bound vesicles found exclusively in animal cells

Lysosomes

Lysosomes have enzymes called

acid hydrolases

Degrades polymers into their monomeric subunits

Lysosomes

The lumen of lysosomes have an acidic pH of

5

Acidic pH also helps to do what, making them accessible to the action of the lysosomal hydrolases

denature proteins

The lysosomal hydrolases work most efficiently at what pH

acidic

Sorting signal that directs soluble lysosomal enzymes from the trans-Golgi network to the late endosomes

mannose-6-phosphate(M6P)

At where does, M6P receptors binds very tightly to M6P residues on lysosome-destined proteins

trans-Golgi

Clarithrin/AP1 coated vesicles containing which receptor bound to lysosomal enzymes bud from the trans-Golgu network which lose their coats then fuse with a late endosome

M6P

Acidic pH (of what) in endosomes causes M6P to dissociate from its receptor

-a phosphate removes phosphate from M6P (prevents rebinding with its receptor)

5-5.5

M6P-receptor is recycled back to Golgi in vesicles coated with a protein complex called

retromer

Late endosomes with lysosomal enzymes fuse with

lysosome

Group of disease caused by the absence of one or more lysosomal enzymes

-Undigested glycolipids and extracellular components accumulate in lysosomes as large inclusions

Lysosomal Storage Diseases

Patients exhibit developmental, neurological and physiological abnormalities

Example: I-Cell Disease

Lysosomal Storage Diseases

Severe lysosomal storage disease

Multiple lysosomal enzymes are missing

Cells from individuals lack N-acetylglucosamine phosphotransferase in cis-Golgi that is required to add Mannose-6-Phosphate on lysosomal targeted proteins

Lysosomal enzymes are secreted instead of being sorted and delivered to lysosomes

I-Cell Disease

A specific receptor on the cell surface binds tightly to an extracellular macromolecular ligand that it recognizes

Receptor-Mediated Endocytosis

ligand complex is recruited to a plasma membrane region that buds inward to form a coated transport vesicle

Receptor

Clathrin/AP2 coated pits

About ?% of surface of some cells (hepatocytes, fibroblast)

2%

Many internalized ligands are observed in

clathrin coated pits

Functions as intermediates in the endocytosis of most ligands bound to cell surface receptors

clathrin coated pits

Some receptors are clustered in clathrin/AP2 coated pits even in the absence of

ligand

Cells take up lipids from the blood in the form of

large lipoprotein complexes

what particles made of proteins and lipids

Carries cholesterol through blood to cells

Lipoproteins

All classes of lipoproteins that the same or different general structure

-Shell composed of apolipoprotein and a phospholipid monolayer containing cholesterol

-Hydrophobic core composed of cholesteryl esters/triglycerides

same

Contains only a single molecule of one type of apolipoprotein (ApoB-100) wrapped around the outside of the particle

20-25 nm diameter

LDL particle

Asn-Pro-X-Tyr Signal on LDL receptor binds to what complex and incorporates the receptor in clathrin/AP2 pits

AP2

Mutations in LDL receptor gene causes

Familial hypercholesterolemia

LDL in the blood is increased about 2 folds

Heterozygotes

LDL in the blood is increased about 4 to 6 folds

Homozygotes

Lead to excessive deposition of LDL forming atherosclerotic plaque in coronary arteries

Homozygotes

Without medical intervention which group usually die of heart attacks before reaching their late 20’s

Homozygotes

Mature endosomes containing numerous vesicles in their interior

multivesicular endosomes

Transport vesicles bud “where” -into the cytosol

outward

Multivesicular endosomes bud “where” – away from the cytosol

inward

Sorting of proteins in what determines which proteins will remain on the lysosomal surface and which ones will be incorporated into internal vesicles

endosomal membrane

Has a mono-ubiquitin tag-sorting signal

Cargo proteins

a peripheral membrane protein

Hrs protein

Ubiquitin tagged Hrs protein (a peripheral membrane protein) recruits protein complex to where

membrane

recruited protein complexes

ESCRT proteins (Endosomal Sorting Complexes Required for Transport)

example of ESCRT proteins (Endosomal Sorting Complexes Required for Transport)

ubiquitin binding protein Tsg101

ESCRT protein complex drives where vesicle budding and loading of cargo proteins in the vesicle

inward

Vps4 ATPase drives the what of ESCRT proteins for next round

disassembly

When cells are placed under stressful conditions, they have the capacity to recycle macromolecules for use as nutrients in a process that involves lysosomal degradation (“eating oneself”)

Autophagy

Delivers cytosolic proteins and organelles to lysosomes for degradation

Autophagic pathway