Unit 04 Pt3

Overview

  • Focus on vesicle fusion with target compartments.

  • Key compartments: Endoplasmic Reticulum (ER) and Golgi Apparatus.

Vesicle Fusion Mechanism

SNAREs in Vesicle Fusion

  • SNAREs (Soluble NSF Attachment Protein Receptors) facilitate vesicle fusion.

  • V-SNARE (vesicular SNARE) on transport vesicle and T-SNARE (target SNARE) on target membrane.

  • T-SNARE depicted in blue; V-SNARE in red.

  • They intertwine creating a helical structure to facilitate fusion.

Proximity Requirement

  • Requires 1 to 1.5 nm distance between membranes for fusion.

  • Close proximity allows lipid flow and excludes water, facilitating fusion.

Fusion Process Steps

  • Trans SNAREs utilize energy to bring membranes closer together.

  • Experimental observations show slow fusion when V-SNAREs mix with T-SNAREs in liposomes.

  • Inner membranes form a stalk facilitating lipid flow between them.

  • SNAREs interact with RAB effector proteins to promote fusion.

Delayed Fusion & Regulated Exocytosis

  • Delayed fusion noted in regulated exocytosis (e.g., neurotransmitter release).

  • Neurotransmitter release at synaptic junctions is crucial and regulated by SNAREs.

Pathological Implications of SNARE Proteins

Botulinum and Tetanus Toxins

  • Clostridium botulinum toxin destroys SNAREs, inhibiting neurotransmitter release.

  • Clostridium tetani also targets SNAREs affecting motor reflexes.

Resetting SNARE Proteins

  • Post-fusion: SNARE proteins must be reset for new transport cycles.

  • Requires ATP hydrolysis and proteins like NSF (N-ethylmaleimide-sensitive factor).

Biological Processes Requiring Membrane Fusion

  • Membrane fusion is crucial for various biological processes such as:

    • Fertilization

    • Muscle fiber formation

    • Viral entry into cells.

  • Viral entry involves virus fusing with cell membranes, and new virus particles budding off.

Golgi Apparatus Overview

  • Important site for carbohydrate synthesis and protein sorting.

  • Proteins transition from ER to Golgi via COPII-coated vesicles.

  • Proteins sorted and directed outwards via secretory vesicles.

Cargo Transport Mechanisms

Selectivity and Exit Signals

  • Research evolved understanding of selectivity via exit signals.

  • Proteins vary in packaging and exit rates based on folding efficiencies.

Chaperone Proteins

  • Chaperones like BiP help in protein folding and masking exit signals.

  • Misfolded proteins are degraded via proteasome.

Transport Vesicles and Retrograde Transport

  • Homotypic membrane fusion can occur allowing for vesicular tubular clusters (VTC).

  • These clusters transport cargo either to the plasma membrane or back to ER via COPI-coated vesicles (retrieval).

KDEL Retrieval Signal

  • KDEL sequence on soluble proteins aids in their retention in the ER.

  • KDEL receptors on the Golgi membranes facilitate retrieval of soluble ER proteins.

  • C-terminal KKXX signals characterize ER membrane resident proteins.

Structure of the Golgi Apparatus

  • Composed of flattened stacks of cisternae (10-20 stacks).

  • Has distinct cis face (near ER) and trans face (near plasma membrane).

Post-Translational Modifications

  • Cargo undergoes modifications while transitioning through the Golgi.

  • Enzymatic activities localized at different regions of the Golgi ensure proper processing.

  • Sorting of proteins is vital for their function and localization.