3/2 Lecture

Vesicle Pinching Off: The process of generating a vesicle from a membrane involves a critical final step where the vesicle must be pinched off from the original membrane. The protein responsible for this process is called Dynamin.
- Function of Dynamin: It helps squeeze and extract the vesicle from the membrane, effectively ejecting it into the cytosol after preparation.

Vesicle Formation Overview: Discussed the role of specific phospholipids (PIPs) in determining which vesicles emerge from certain membranes.
Clathrin-coated Vesicles: An earlier type of vesicle discussed, focusing on its formation.

Definition and Function: COP II coated vesicles are essential for transporting materials from the rough endoplasmic reticulum (RER) to the Golgi apparatus.
- Identification of Cargo: Involves the identification and binding of cargo proteins, recruitment of adapter proteins, and establishment of outer coat proteins which assist in defining the structure of the forming vesicle.
Key Adapter Proteins:
- SAR1 GTP: An adapter protein that has an amphipathic helix, significant for its role in recruiting and embedding into the membrane.
- Transition from GDP to GTP: SAR1 initially exists as SAR1-GDP in the cytosol and undergoes a conformational change upon binding to SAR1 GEF, transitioning to the membrane with GTP bound.
  - Specific Activation: Only activated at the RER membrane where SAR1-GEF is present, providing specificity for COP II vesicle formation.
Recruitment Process:
- Once SAR1 is embedded, it recruits inner coat proteins Sec23 and Sec24, which function similarly to clathrin's adapter proteins.
- Outer Coat Proteins: Sec13 and Sec31 are recruited subsequently, culminating in the formation of a spherical vesicle.
- Further insight into naming conventions: Many proteins were named due to studies in yeast where mutations leading to secretion defects acquired the prefix "Sec".

Formation Flexibility: COP II vesicles can form in various shapes to accommodate larger cargo, such as collagen or large extracellular matrix proteins, which clathrin-coated vesicles cannot package efficiently.
- They are capable of assembling irregularly shaped vesicles vital for larger protein transport from the ER:
- Transport Process: Vesicles transport large proteins into the ER lumen for subsequent secretion.

Collagen Synthesis: Even large proteins like collagen carry an ER signal sequence that facilitates its entry into the RER, eventually binding to cargo receptors for vesicle packaging.
Cumulative Exam Focus: While the exam may not directly cover the extracellular matrix, it encourages recognition of relevant proteins as part of broader protein secretion pathways.

Dynamin Functionality: Similar to the formation process in clathrin-coated vesicles, COP II vesicles also require dynamin for the final detachment from the ER membrane.

Identifying the Target Membrane: Communication occurs between proteins on vesicles and their corresponding target membranes—vital for vesicle docking and fusion.
- Key Proteins: Various Rabs (GTPases similar to SAR1) shape the identity of membranes through differential presence at various cellular locations.
- Function of Rabs: Rabs are crucial for determining vesicle specificities, ensuring vesicles correctly dock with their intended membranes.
Rabs and Effectors: Each vesicle has Rab proteins that bind to specific Rab effector proteins on the target membrane, driving docked vesicle adhesion closer to the membrane.

SNARE Classification:
- V-SNAREs: Found on vesicles (e.g., presynaptically)
- T-SNAREs: Located on the target membrane (e.g., postsynaptically).
SNARE Complex Formation: Upon docking at the target membrane, v-SNAREs and t-SNAREs entwine, pulling membranes close enough to enable fusion:
- Water Molecule Clearance: The fusion process necessitates the removal of water molecules between the two membranes to favor membrane fusion.
- ATP Involvement: ATP is consumed to disassemble SNARE complexes post-fusion, facilitating new vesicle packaging efforts.

Membrane Identity: Specific regions in membranes exhibiting Rabs communicate the necessity of recruiting the correct integratory proteins for subsequent vesicle transport.
- Transitioning between different Rab domains exemplifies the dynamic change in membrane identity reliant upon Rabs throughout the cellular process.
Retrograde vs. Anterograde Transport: Different vesicular transport mechanisms are in place (COP II for anterograde transport from ER to Golgi; COP I for retrograde from Golgi to ER).

Golgi Apparatus Overview: The Golgi comprises distinct regions (cis, medial, trans) that facilitate glycosylation and other modifications to proteins arriving from the ER.
Enzymatic Modifications: Varying enzymes present in each compartment contribute to diverse post-translation alterations of proteins, primarily sugar modifications.
Resident Proteins: Proteins within the Golgi that stay anchored to perform specific roles include various glycosylation enzymes and Golgins which organize the Golgi apparatus and mediate vesicle transport.

Pathway Stages:
1. Proteins synthesized and processed in the RER
2. Transported to Golgi, processed further, sorted, and dispatched to the target membranes like the plasma membrane
3. Potential lysosomal and endosomal pathways also outlined, collectively encompassing the secretory system.

This comprehensive overview outlines the critical mechanisms involved in vesicle formation, targeting, and secretion through organelles of the endomembrane system, emphasizing key protein interactions, pathways, and cellular significance.