Endocytosis IV

Types of Endocytosis

Phagocytosis

• Description: Specialized endocytosis for engulfing large particles (≥500 nm).

• Cell Types: Macrophages, neutrophils, and dendritic cells.

• Mechanism:

  • Involves actin polymerization and receptor-mediated pathways.

  • Antibodies on target bind to Fc receptors on phagocytes, leading to pseudopod formation and cargo engulfment.

Autophagy

• Definition: A cellular degradation pathway for the turnover of cytoplasmic components.

• Types of Cargo: Long-lived proteins, protein aggregates, and defective organelles.

• Purpose:

  • Nutrient acquisition during starvation.

  • Removal of damaged organelles and protein aggregates.

  • Specific degradation of bacteria/infectious agents.

• Mechanisms:

  • Chaperone-mediated autophagy: Involves specific binding to chaperones and transport to lysosomes.

  • Microautophagy: Engulfment of cytoplasmic components directly by membrane invagination.

  • Macroautophagy: Formation of autophagosome encapsulating large structures.

Major Endocytic Components

• Plasma Membrane: Starting point for endocytosis.

• Cytoplasm: Where internalized material is processed.

• Rab Proteins: Essential for endosomal trafficking at different pH levels:

  • Rab5: Early endosomes (pH 6.0)

  • Rab7: Late endosomes (pH 5.0)

  • Rab9: Associated with TGN and lysosomes.

Receptor-Mediated Endocytosis Pathways

1. LDL, Insulin, Prolactin: Receptor recycled; ligand degraded.

2. EGF: Receptor down-regulated after binding; not recycled.

3. Diferric transferrin: Recycled back to the cell surface.

4. IgA and IgG: Transcytosis mechanism, coupling endocytosis and exocytosis.

5. Endothelial Transcytosis: Pathway involving albumin.

6. Phagocytosis: To be covered in detail.

Phagosome Maturation

• Steps:

  • Phagosomes fuse with early and late endosomes, gradually acquiring proteins.

  • Fusion with lysosomes transforms phagosome into phagolysosome for degradation.

• Escape Strategies by Pathogens:

  • Conversion of phagosome to autophagosome.

  • Inhibition of phagosome maturation.

Rab Proteins

• Usually have one or more fatty acids at C-terminus that allows insertion in membranes when in GTP state

• Typically, one Rab has many effectors that carry out different function son the same organelle

• Common functions:

  • Tethering proteins

    • EEA1 for Rab5

    • p115 for Rab7

  • Regulators of motor proteins

  • Regulators of lipid metabolism

• There are more than 60 rabs in humans, each unique to different compartments/organelles

  • Rab5 on early endosomes/phagosomes

    • Activated Rab5 recruits effectors for tethering and fusion between early endosomes

    • Rab5 recruits PI 3-kinase which produces PIP3

      • Phosphoinositides are found on endosome membranes as trafficking signals

  • Rab7 on late endosomes/phagosomes, autophagosomes, lysosomes

    • Contain PI(3,5)P2

• EEA-1 in early endosome-early endosome fusion

  • A long filamentous tethering protein

  • Assembled into homodimers

  • Bind to PI3P (FYVE domain), Rab5 (2 Rab binding domains)

  • EEA-1 and rabenosyn-5 form a complex that brings the membranes together

• Rab7 in late endosome regulation:

  • Rab7 requires 3 proteins for minus-end microtubule transport

    • RILP - recruits and connects late endosome to dynactin-dynein machinery

    • ORP1L - cholesterol sensor

      • If cholesterol is high, it recruits BIII spectrin, which recruits dynactin, which recruit dynein forminus-end movement

      • If cholesterol is low, ???, plus-end movement

    • HOPS - tethering protein

Endosome Maturation

• Early endosomes become late endosomes

• Maturation process:

  • Change in subcellular localization of endosome (motors)

  • Decrease in pH

  • Change in lipid composition (PI3P → PI(3,5)P2)

  • Closing off access to the recycling branch (retain vacuolar portion only)

  • Receive acidic hydrolases for lysosome function from TGN

  • Change in fusion machinery (tethers and SNAREs)

• Rab Conversion Process:

  • Activated Rab5 recruits Rab7 GEF

  • GEF catalyzes the exchange from GDP → GTP on Rab7

  • Activated Rab7 recruits TBC2 (Rab5 GAP) to catalyze the

    hydrolysis of GTP of Rab5

  • GDP bound Rab5 leaves the endosome membrane

Delivery to Lysosomes

• Rab7 is found on BOTH late endosomes and lysosomes along with the proper SNARE proteins to fuse

• Thus, late endosomes and lysosomes can be tethered together with the HOPS complex.

• Fusion of a late endosome with a lysosome produces a hybrid

  organelle with characteristics of both, sometimes referred to as an

  endolysosome

• As the material within is digested, the endolysosome becomes lysosome-like again.

Salmonella

• Salmonella is a pathogen that can enter digestive epithelial cells

• Salmonella arrests the late endosomes and prevents them from fusing with the lysosomes