Introduction to Vesicular Transport

Introduction to Vesicular Transport

• Almost all proteins are synthesized in the cytosol
• Membrane-enclosed organelles import proteins by one of three mechanisms:
  • transported fully-folded through a pore
  • transported by protein translocators (generally as an unfolded polypeptide chain)
  • delivered by transport vesicles
• The extracellular space and each of the membrane-enclosed compartments (shaded gray) communicate via transport vesicles.
• These membranes are collectively considered the endomembrane system.
• Vesicles transport soluble proteins, membrane proteins, and lipids.
• The leaflet (inside) of the ER or Golgi is the same as the leaflet (outside) of the cell.
• The part of the protein facing the cytoplasm of the ER, Golgi, or vesicles will face the cytoplasm of the plasma membrane.

Basics of Vesicular Transport

• Vesicles bud off a donor compartment and fuse with a target compartment.
• Vesicles carry cargo from the donor compartment to the target compartment.
• Cargo can be soluble proteins, membrane proteins, and lipids.
• The assembly of a protein coat drives the budding process.
• Different coats are used for different steps in vesicle transport.
• COPII-mediated ER to Golgi vesicle traffic will be the primary focus.

COPII Coat Assembly

• The monomeric GTPAse Sar1 is responsible for the assembly of COPII coats at the ER membrane.
• The activity of Sar1 is spatially regulated by an ER-associated Sar1-GEF.
• GEF (guanine exchange factor) promotes the exchange of GDP for GTP.
• Sar1-GTP recruits COPII adaptor proteins.
• COPII adaptor proteins act to:
  • select cargo proteins to be packaged
  • initiate deformation of the ER membrane
  • recruit outer coat proteins which help form a bud

Cargo Selection for Transport

• EXIT SIGNALS are commonly an amino acid sequence on the protein.
• Both soluble and membrane-bound cargo proteins have exit signals.
• Membrane-bound proteins with exit signals are either:
  • destined to reside in the membrane of another organelle in the endomembrane system
  • act as cargo receptors for soluble proteins and are recycled back to the ER.
• The coat serves two main functions:
  • the inner coat serves to concentrate cargo
  • the outer coat, with its curved structure, deforms the membrane and shapes the vesicle
• Occasionally, resident ER proteins are trapped in the vesicle.

Vesicle Targeting and Fusion

• A sequential targeting system increases the probability of a vesicle fusing with the correct target.
• First targeting system: Specific Rab proteins on the surface of each vesicle are recognized by corresponding tethering proteins on the cytosolic surface of the target membrane.
• Second targeting system: SNAREs on the vesicle (v-SNAREs) interact with complementary SNAREs on the target membrane (t-SNAREs).
• Each transport vesicle-target membrane pair has unique Rab-tether and v-SNARE-t-SNARE pairs.
• SNARE proteins catalyze the fusion of the donor vesicle with the target membrane.
• Membranes must be brought within 1.5 nm of each other for lipids to flow from one bilayer to the other.
• Once fused with the target membrane, the cargo receptor releases soluble cargo.

Next Class

• ER exit and quality control
• the Golgi
• exocytosis

Introduction

• Most proteins are synthesized in the cytosol
• Proteins are trafficked to their specific compartment

Vesicular Transport within the Endomembrane System

• The endomembrane system includes all membrane-enclosed organelles
• Transport vesicles allow communication between these organelles
• Vesicles carry cargo including soluble proteins, membrane-bound proteins, and lipids

Membrane Topology

• The lumen of ER and Golgi is equivalent to the exterior of the cell
• The leaflet facing the cytoplasm of ER or Golgi corresponds to the cytoplasmic leaflet of the plasma membrane
• The part of a protein within the ER or Golgi lumen is equivalent to the part outside the cell
• This is due to the continuity of membrane leaflets as vesicles bud off from the donor, and fuse with the target, compartment

Vesicle Transport

• Vesicles must be selectively sorted to reach the correct target compartment
• Each compartment must have its own specific composition of proteins and lipids

The Basics of Vesicular Transport

• Vesicles bud off a donor compartment and fuse with a target compartment
• Coat proteins are involved in the budding process
• Different coat proteins are used for different transport pathways

COPII-mediated Transport

• The COPII coat mediates ER to Golgi transport
• Sar1, a monomeric GTPase, assembles the COPII coat at the ER membrane
• Sar1-GEF, an ER-associated protein, activates Sar1 by promoting GDP exchange for GTP
• Sar1-GTP recruits COPII adaptor proteins, responsible for selecting cargo, membrane deformation, and recruiting outer coat proteins

Cargo Selection for Transport

• Exit signals are used to select cargo proteins
• Both soluble and membrane-bound cargo proteins have exit signals
• Membrane-bound proteins with exit signals can either reside in another organelle’s membrane or act as cargo receptors

Function of the Coat

• The inner coat concentrates cargo proteins
• The outer coat, with its curved structure, deforms the membrane and shapes the vesicle

Targeting and Fusion

• Vesicles utilize a sequential targeting system with two levels
• The first targeting system involves Rab proteins and tethering proteins
• Rab proteins on the vesicle are recognized by corresponding tethering proteins on the target membrane
• The second targeting system involves SNAREs
• v-SNAREs on the vesicle interact with complementary t-SNAREs on the target membrane

Fusion Process

• SNARE proteins catalyze vesicle fusion with the target membrane
• They bring membranes within 1.5 nm to allow lipid flow and fusion

Cargo Receptor Release

• Once fused, cargo receptors release soluble cargo
• The mechanism is explained in the textbook

Next Class

• ER exit and quality control
• The Golgi apparatus
• Exocytosis

Explore the mechanisms and processes involved in vesicular transport within cells. This quiz covers the roles of membrane-enclosed organelles, the endomembrane system, and how vesicles facilitate the movement of proteins and lipids. Test your understanding of these essential cellular functions.