17.1: Techniques for Studying the Secretory Pathway

• Assays that follow protein trafficking through a secretory pathway need to label a cohort and identify where the labeled proteins are located

• Radioactive amino acids can be used for pulse labeling a cohort of proteins that are newly made in the ER

• Microscopy can show the transport of fluorescently labeled proteins as they move along the secretory pathway

• Yeast contains many of the components needed for intracellular protein trafficking

• Biochemical dissection of the secretory pathway needs the intercompartmental protein transport powered by cell-free assays

17.2: Molecular Mechanisms of Vesicular Traffic

• Three transport vesicles:

  • COPI

  • COPII

  • Clathrin vesicles

• They are differentiated based on their protein coats and which routes they are mediating

• Polymerization of cytosolic coat proteins onto a parent/donor membrane is needed to form coated vesicles, and the bud of the vesicle eventually releases from the membrane as a complete vesicle

• Polymerization is controlled by GTP binding proteins which are a part of the GTPase superfamily

• The vesicles are disassembled by the hydrolysis of GTP bound to either ARF or Sar1 after they are released from the donor membrane

• Sorting signals found in the membrane interact with coat proteins along with luminal proteins of the donor organelles during vesicle budding, during cargo proteins into vesicles

• The viral envelope fuses around the endosomal membrane after the endocytosis of the enveloped animal virus occurs

17.3: Early Stages of Secretory Pathway

• Proteins are transported through the rough ER with the help of COPII, and COPI vesicles transport those proteins in the opposite direction

• COPII coats have three components:

  • Small GTP binding protein Sar1

  • A Sec23/Sec24 complex

  • Sec13/Sec31 complex

• These components bind to membrane cargo proteins that contain a di-acidic or any other form of soring signal located in their cytosolic region

• The cis-Golgi contains the membrane proteins needed for the formation of COPII vesicles, and COPI vesicles can retrieve them

• ER-resident proteins contain a KDEL sorting signal and this signal binding to a receptor protein in the cis-Golgi helps to get ER proteins needed for COPLI vesicles

17.4: Later Stages of the Secretory Pathway

• A branch point for soluble secreted proteins, lysosomal proteins, and some cell membrane proteins is the trans-Golgi network (TGN)

• Vesicles from the trans-Golgi network along with endocytic vesicles have a coat of AP (adapter protein) complexes

• The cis-Golgi modifies soluble enzymes that are meant to go into the lysosomes

• The concentration and storing of regulated proteins occurs in the secretory vesicles

• These proteins are stored until they receive a neural or hormonal signal to start exocytosis from the cell

17.5: Receptor-Mediated Endocytosis and the Sorting of Internalized Proteins

• Internalization can occur with some extracellular ligands which bind to cell surface receptors, and the clathrin-coated vesicles that the receptors are internalized into also contain AP2 complexes

• There are many sorting signals needed for the cell surface receptors:

  • Asn-Pro-X-Tyr

  • Tyr-X-X-oI

  • Leu-Leu

• Lysosomes are degraded once they are delivered through the endocytic pathway

• An endocytic pathway helps to transport Iron into the cell

• Lysosomes end incorporated into vesicles inside the endosome if they endocytose proteins meant for degradation

• Cellular components which mediate inward budding in the endosomal membrane can be used for the budding and pinching off of enveloped viruses in the cell

17.6: Synaptic Vesicle Function and Formation

• Nerve impulses transmission at chemical synapses depend on the exocytosis of neurotransmitter filled synaptic vesicles found in cells along with those empty vesicles regenerating

• Vesicles moving to the presynaptic membrane requires cytosolic proteins and a GTP- binding protein which stays connected to the vesicle membrane

• Endocytic budding rapidly regenerates synaptic vesicles which are coated in clathrin and this coating coming off helps the vesicles refill their neurotransmitter and move to the active zone, where they are once again put through docking and fusion