Lecture 12: Vesicular Trafficking I

These flashcards cover the key principles of vesicular transport, including GTPase cycles, the specifics of COPII and COPI systems, adaptor proteins, and sorting signals based on the BIS225 Lecture 11 notes.

How are vesicles targeted to their destination while preventing leakage of soluble cargo?

Vesicles are targeted with high fidelity to their destination as an asymmetric lipid bilayer to ensure cargo is delivered without leakage.

What is the purpose of maintaining molecule topology during vesicular transport?

It ensures that membrane-bound cargo molecules maintain their specific orientation within the membrane during the transport process.

What are the roles of v-SNARES and t-SNAREs in vesicular transport?

v-SNARES (on the vesicle) and t-SNAREs (on the target compartment) are involved in the docking and fusion of the vesicle with the target membrane.

Why is transport for large macromolecules mediated by vesicles and tubules rather than channels?

Large macromolecules like proteins are too bulky and have a variety of charges that prevent them from passing through channels.

What must happen to a transport vesicle's coat before fusion can occur?

The coat needs to come off for fusion to occur.

How do vesicles move through the secretory pathway to reach the Golgi?

Vesicles hook onto microtubules and move via motor proteins.

What are the three essential components required for the formation of all transport vesicles?

A GTPase, adaptor proteins, and a coat.

According to the molecular switch model, what are the two states of small GTPases?

They switch between an inactive GDP-bound form and an active GTP-bound form.

What is the function of Guanine nucleotide exchange factors (GEFs)?

GEFs facilitate the exchange of GDP for GTP to activate the GTPase.

What is the function of GTPase Activating Proteins (GAPs)?

GAPs stimulate GTP hydrolysis to convert the GTPase back to its inactive GDP-bound form.

Where are the GDP and GTP forms of a GTPase typically located?

The GDP form is usually found in the cytosol, while the GTP form is membrane-associated.

Which protein is considered the founding member of the small GTPase family?

Ras.

Which specific GTPase is used for the formation of COPII coated vesicles?

Sar1, which is a member of the Arf family.

What are the adaptor and coat proteins required for COPII vesicle formation?

The adaptor is Sec23/24 and the coat is Sec13/31.

What is the role of chaperones in the Endoplasmic Reticulum (ER)?

Chaperones ensure the proper folding of newly synthesized proteins, including soluble and membrane proteins.

What structural change occurs in Sar1 when it is activated from Sar1-GDP to Sar1-GTP?

A hidden amphiphilic helix flips out, allowing Sar1 to anchor to the donor membrane (ER).

Which protein acts as the GEF for Sar1?

Sec12.

What is the specific role of the Sec23/24 adaptor complex in COPII formation?

Sec23/24 recognizes signals in cargo receptors and its bowtie-shaped crystal structure allows for membrane curvature.

How is the disassembly of the COPII coat promoted?

Sec23 acts as a GAP for Sar1, and this activity is enhanced following the recruitment of the Sec13/31 coat, promoting disassembly.

In reconstitution experiments, what are the minimal components required to form a COPII vesicle from ER membranes?

Cytosol, ATP, and GTP.

What is the difference between Ribophorin and p58 in COPII experimental assays?

Ribophorin is a resident ER protein that should not be in the vesicles, while p58 is a COPII cargo protein that should be incorporated.

What is the effect of expressing the Sar1GDP mutant?

It inhibits COPII formation by sequestering GEFs, resulting in a dominant negative effect.

What is the primary role of the COPI coat in the secretory pathway?

COPI is involved in retrieval transport from the vesicular tubular cluster or Golgi back to the ER.

Which GTPase and coat are used for the transport of lysosomal proteins from the TGN?

The GTPase is Arf1 and the coat is Clathrin.

What are the two primary functions of adaptor proteins?

They recognize and select cargo to ensure specificity and link the coat to the membrane.

What are the subcellular localizations for the adaptor complexes AP1 and AP2?

AP1 is localized to the TGN, and AP2 is localized to the Plasma Membrane (PM).

What is an example of a Tyrosine-based sorting signal and its associated receptor?

The signal is YxxPhi, found in the Transferrin Receptor (TfR).

What is the sorting signal for the LDL receptor?

FXNPXY.

Which sorting signal is recognized for the internalisation of the epidermal growth factor receptor (EGFR)?

Ubiquitin.

Which subunits of the AP2 adaptor complex are responsible for recognizing sorting signals?

$\mu$ and $\sigma$ subunits.