Week 3 A Intracellular Trafficking

Intracellular Trafficking Overview

• Focuses on:

  • Protein targeting

  • Vesicle trafficking

  • Secretion

Key Topics to Consider

• Compartmentalisation in Multicellular Organisms

  • Intracellular compartmentalisation is essential for cellular function and specialization.

• Targeting Proteins

  • Proteins must be directed to the correct cellular compartments for proper function.

• Mechanisms of Transport within Cells

  • Various mechanisms facilitate the movement of proteins and other molecules within cellular compartments.

Cellular Structures and Components (Animal Cell)

• Key Structures:

  • Microtubules: Structural components that maintain the cell's shape.

  • Centrosome & Centrioles: Involved in cell division.

  • Nucleus: Contains chromatin (DNA)

    • Nuclear Pores: Allow transport of materials in and out of the nucleus.

  • Cytosol: Fluid component of the cytoplasm.

  • Peroxisomes, Ribosomes, Lysosomes, Mitochondria: Play roles in metabolism, protein synthesis, and cell respiration.

  • Golgi Apparatus: Involved in modifying, sorting, and packaging proteins.

  • Endoplasmic Reticulum (ER): Includes rough ER (with ribosomes) and smooth ER.

  • Plasma Membrane: Encases the cell.

  • Extracellular Matrix: Provides structural and biochemical support to surrounding cells.

Intracellular Compartment Volumes

• Volume Distribution Estimates Per Cell:

  • Cytosol: 50-60%

  • Mitochondria: 20%

  • Rough ER: 10%

  • Smooth ER: 6%

  • Golgi: 6%

  • Nucleus: 1%

  • Peroxisomes, Lysosomes, & Endosomes: ~1% each.

Nobel Prize in Physiology or Medicine - 2013

• Jointly awarded to:

  • James E. Rothman

  • Randy W. Schekman

  • Thomas C. Südhof

• Work Focus: Discovered the machinery regulating vesicle traffic, essential for cellular transport systems.

Transport Processes in Cells

• Cell as a Factory:

  • Produces and exports molecules such as insulin (in blood) and neurotransmitters (between nerve cells).

• Vesicle Transport:

  • Small packages called vesicles transport molecules to their specific destinations.

Contributions of the Nobel Laureates

• Randy Schekman:

  • Identified genes necessary for vesicle traffic.

• James Rothman:

  • Unraveled the protein machinery enabling vesicle targeting.

• Thomas Südhof:

  • Discovered how signals instruct vesicles to release their contents precisely.

Genetic Control of Vesicular Traffic (Randy Schekman)

• Studied yeast: Developed a genetic screen to identify defects in vesicle transport.

• Findings led to the identification of genes responsible for cellular transport processes and provided insights into genetic regulation.

Maturation of Secretory Proteins in Yeast

• Five Steps Defined by Yeast Mutants:

  • Transport from:

    1. Rough ER to the Golgi

    2. Golgi to secretory vesicles

    3. Secretory vesicles to cell surface

  • Each step affected by specific classes of Sec genes resulting in distinct outcomes.

Microscopy Analysis of Trafficking Mutations in Yeast

• Visualized effects of temperature-sensitive trafficking mutations via electron microscopy.

• Illustrates impact of mutation on vesicle transport at different temperatures.

Key References for Further Reading

1. Novick, P., Schekman, R. (1979). Proc Natl Acad Sci USA.

2. Kaiser, C.A., Schekman, R. (1990). Cell.

• Recommended for a deeper understanding of experimental elucidation related to vesicle trafficking (not essential reading).

Endocytic and Secretory Pathways

• Clathrin & COPII Transport Mechanisms:

  • Clathrin-coated pits function in endocytosis.

  • COPII vesicles facilitate transport from the ER to the Golgi.

Sorting Signals and Vesicle Types (Table 17-6)**

• Signal Sequences:

  • Direct specific proteins to their corresponding transport vesicles (e.g., KDEL, Mannose 6-phosphate).

  • Each sequence defines the type of protein and the corresponding vesicle for transport.

Brefeldin A

• A fungal metabolite that blocks transport from ER to Golgi, used to study the mechanism of secretion.

Vesicle Fusion and SNARE Proteins

• SNARE Complex:

  • Involves V-SNARE (on transport vesicles) and T-SNARE (on target membranes) for vesicle docking and fusion.

• Rab Proteins:

  • Control the specificity of vesicle fusion, enhancing the accuracy of transport processes.

Rab GTP-binding Proteins

• Monomeric GTPases: Over 30 family members with varied functions depending on the organelle location (e.g., early endosomes, Golgi complex).

Lipid Influence on Membrane Dynamics

• Phosphoinositides in Phagocytosis and Cellular Compartmentalization:

  • Different PI lipids are involved in specifying cellular compartments.

Conclusion on Transport Mechanisms

• Understanding intracellular traffic is crucial for comprehending cellular functions, including secretion, membrane dynamics, and metabolic processes.