The System of Internal Membranes

Overview of Eukaryotic Cells

• Internal compartments include:<br> - Nucleus <br> - Endosome <br> - Mitochondrion <br> - Lysosome <br> - Golgi apparatus <br> - Cytosol <br> - Endoplasmic reticulum (ER) <br> - Peroxisome <br> - Plasma membrane <br> - Membrane-bound polyribosomes <br> - Free ribosomes

Membrane Traffic

• Endocytosis: Import mediated by endocytic vesicles.

• Exocytosis: Export (secretion).

• Continuous exchange of materials via:<br> - Plasma membrane<br> - Small vesicles<br> - Endoplasmic reticulum<br> - Golgi apparatus<br> - Lysosomes

Eukaryotic Cell Nucleus

Structure

• Nuclear Envelope: Double membrane with an outer layer continuous with the rough ER.

• Perinuclear Space: The gap between the inner and outer membranes.

• Nuclear Pores: Facilitate both passive (for small proteins) and active (for larger proteins) transport.

Nuclear Lamina

• A network of lamin protein fibers providing structural support to the nucleus.<br>- Serves as attachment sites for chromatin.<br>- Destruction leads to nuclear collapse.<br>- Part of the cytoskeleton.

Nuclear Localization Signals (NLS)

• Definition: Amino acid sequences that allow proteins to enter the nucleus.

• Example: Seven amino acid stretch in SV40 T-antigen allows entry into the nucleus.

• Transport Mechanism:

  • Chimeric protein experiments (e.g., Pyruvate Kinase with NLS) used to assess NLS functionality.

  • Nuclear transport is energy-dependent and involves transport receptor proteins which recognize NLS.

Active Transport of Proteins

• Involves the binding of nuclear transport receptors to proteins containing NLS.

• Process includes:<br> - Translocation through nuclear pore<br> - Interaction with Ran-GTP which releases cargo inside the nucleus.

• Ran Cycle:<br> - Ran-GTP favors release of proteins in the nucleus, while Ran-GDP returns to nucleus.

Protein Synthesis in the Endoplasmic Reticulum

Types of Endoplasmic Reticulum

• Rough ER::

  • Site of protein synthesis; ribosome-associated.

  • Functions in folding, modification, and transport of proteins.

• Smooth ER:

  • Synthesizes lipids; no ribosomes.

Secreted Protein Synthesis

• All secreted proteins have an ER signal sequence for translocation.

• Signal Recognition Particle (SRP) recognizes ER signal and mediates docking on the ER membrane.

Golgi Apparatus

Function

• Protein modification, sorting, and packaging for secretion or delivery to organelles.<br>- Processes proteins received from ER and sorts them for export or internal use via vesicles.

Secretion Pathway

• Proteins from the rough ER are sent to the Golgi then directed to either:<br> - Secretory vesicles (out of the cell)<br> - Plasma membrane (membrane proteins)<br> - Lysosome (lysosomal proteins)

Exocytosis and Endocytosis

Exocytosis Types

• Constitutive Secretion: Unregulated transport present in all cells.

• Regulated Secretion: Occurs only in specialized cells, triggered by specific signals.

Endocytosis Mechanisms

• Receptor-mediated: Specific molecules (like LDL cholesterol) recognized and internalized by receptors.

• Phagocytosis: Ingestion of large particles or cells via pseudopodia.

• Autophagy: Self-digestion mechanism for maintaining cellular health.

Lysosomes

• Digestive organelles that degrade materials taken in by endocytosis or phagocytosis.<br>- Require an acidic internal environment (pH ~5) for enzyme function.

Receptor-mediated Endocytosis Example

• LDL is recognized by specific receptors, taken in, and delivered to lysosome for degradation.

Autophagy Process

• Elimination of old/cell components via engulfment and fusion with lysosomes.

• Four phases:

  1. Nucleation and extension of membrane.

  2. Closure of autophagosome.

  3. Fusion with lysosome.

  4. Digestion by lysosomal enzymes.

Ohsumi Experiment

• Demonstrated autophagy using yeast cells—showed accumulation of autophagosomes when degradation enzymes were lacking.