VL 34 (Michael Sauer)

Question 1

The endoplasmic reticulum (ER): gateway of the secretory pathway

Answer 1

• discovered 1945 in chicken embryo cells
• all eukaryotic cells
• intracellular compartment: lipid bilayer membrane enclosing an inner space (lumen)
• = network (“reticulum”) of interconnected membrane tubules, cisternae stretching across entire cytoplasm (“endoplasmic”)
• synthesis location of: membrane proteins, soluble cargo proteins, lipids → transported to other compartments/extracellular space along secretory pathway

Question 2

Adding a signal peptide to a cytosolic protein directs the protein into the ER:

Answer 2

• signal sequence on N-terminus required → protein send into ER-lumen
• often hydrophobic, ~ 15-25 aa

Question 3

Translation of membrane proteins:

Answer 3

• quality control mechanism recognizes unfolded proteins
  → chaperones released
  → misfolded protein response, if chaperones couldn ́t help
  → ERAD
  → retrotranslocation

Question 4

ER transport:

Answer 4

Question 5

GTPase Cycle:

Answer 5

Question 6

Coated protein complex II (COPII): assembly and function

Answer 6

• coats build up
  → outer coat attaches
  →membrane deformation (flat → round)
• SAR1 inactive state (cytosol)
• SAR1 activation by SEC12 → SAR1 active state by GDP-GTP exchange
• recruitment of SEC23-SEC24 heterodimer + cargo
• cluster
• recruitment of SEC13-SEC31 heterodimer → membrane
  shaped into vesicle → pinch off

Question 7

The earlyvsecretoryvpathwayvin plants

Answer 7

Question 8

Golgi apparatus:

Answer 8

• discovered by Italian physician Camillo Golgi (1897)
• synthesis of cell wall components (pectins, hemicellulose); cellulose synthesized at plasma membrane
• protein glycosylation continues (start in ER)

Question 9

The trans-Golgi network (TGN)

Answer 9

• derives from golgi through cistearnae maturation
• = tubular-vesicular organelle with pleimorphic shape (undefined, irregular shape)
• cistern dissolution at TGN site

Question 10

Exocytosis process:

Answer 10

Exocytosis (= secretion) needs v(esicle)-SNAREs and t(arget)-SNAREs for docking, fusion:

SNAREs
* form highly stable protein-protein interactions
→help to overcome energy barrier required for membrane fusion
* 57 different types in Arabidopsis
▪ belong to different classes
▪ each class has different members
▪ each member has a specific function in a specific transport route

• vSNARE binds t-SNARE * →vesicle pulled to
  membrane

Question 11

Exocytosis process:

Answer 11

Question 12

Molecular components of clathrin-mediated endocytosis:

Answer 12

Association of clathrin triskelions:
* 6 SU (3 heavy chains with 1 of each 2 different light chains)
* self-assembly → driving force for membrane invagination
Process
* AP2-accumulation on membrane surface
* clathrin-coat formation
* Amphiphysin and dynamin invagination of the membrane; cleavage (formation of a coated pits)
* uncoating by synaptojanin
* transport vesicle release for fusion with endosomes

Question 13

TPLATE Adaptor Complex Drives Clathrin-Mediated Endocytosis in Plants:

Answer 13

TPLATE:
* eight-core-component protein complex
* adaptor module for clathrin-mediated endocytosis → essential for plant growth
* recruited as dynamic foci at the plasma membrane preceding recruitment of AP2, clathrin, dynamin-related proteins
* reduced function of different complex components impaired internalization of
assorted endocytic cargoes