L12 types of vesciles (Tfk1)

• secretory pathway generally is: ER (protein synthesis) -> golgi (PTMs) -> vesicles into plasma membrane or endocome
• vesicles carry cargo to either go out of the cell or into it. the cargo is released via vesicle binding to the membrane of its target
• vesicles that go from ER to golgi are called COP-II vesicles (anterograde transport); vesicles from golgi back into ER is COP-I vesicles (retrograde transport); vesicles from exterior of cell to the interior are clathrin-coated vesicles (also golgi into lysosome or golgi into endosome)
• general vesicle formation is

  1. initiation event in membrane that leads to formation of vesicle
  2. coat formation, cytosolic adaptor proteins interact with initiator
  3. fission event where vesicle is detached from donor compartment
  4. vesicle uncoated
• for fission to occur, cargo is recognized by a receptor protein, which is recognized by an adaptor, which is recognized by a coat protein to create the coat
• COPI and COPII have a Ras GTPase where GTP = active conformation, GDP = inactive conformation. GAP (making inactive) and GEF (making active) proteins are needed for changing from active to inactive conformation
• process for creation of a COP-II vesicle (from Er to the Golgi)

  1. Sar 1 (a Ras GTPase) in the cytosol gets its amphipathic helix exposed when GEF changes GDP to GTP, making Sar1 part of the membrane facing the cytosol
  2. Sec23 recognizes Sar1 and Sec24 recognize the receptor bound to the cargo to be brought into the vesicle.
  3. Sec13/31 recognize the adaptor proteins (Sec23 and Sec24). they pinch the protein to create a vesicle and create a coat. (ATP-GTP independent)
  4. Sec23/24 will induce a change in Sar1 to make it no longer part of the membrane
• bulk transport/flow is when proteins in the ER are taken to the golgi and plasma membrane by default (some more efficient than others because of motifs) - done by COP-II vesicles
• ER exit signals on transmembrane proteins (on cytosolic side) will have one of the following motifs recognized by Sec23/24 adaptors - done by Cop-II vesicles

  • two phenylalanine at c-terminus
  • aspartic acid - x - glutamic acid within the sequence
• ER exit signals on lumenal proteins are recognized by receptor with two phenylalanine's in the c-terminus facing the cytosol - done by Cop-II vesicles
• COP-I vesicles move signals back to the ER. lumenal proteins for this pathway will have KDEL-COO- at c-terminus, recognized by TM KDEL receptor with KKxx motif at cytosolic c-terminus. TM proteins for this pathway have KKxx-COO- at c-terminus in cytosol (type 1 TM protein)or NH3+-MxxRR at cytosolic n-terminus in cytosol (type 2 TM protein) recognized by COP-I coat adaptors.
• formation for COP-I vesicle is the same as a COP-II vesicle.

  1. intiator is ARF1 (Ras GTPase) with now exposed amphipathic helix that makes it part of the membrane
  2. adaptors beta, delta and gamma subunit recognizes receptors on the cargo
  3. coat proteins are alpha and beta' subunits to pinch of membrane.
  4. beta, delta and gamma adaptors are GAPs for ATF1 which disassociate the coat
• clathrin coated vesicles do not need a Ras GTPase!! golgi to endosomes or plasma membrane to lysosome

  1. initiator is phosphatidyl inositol (PI) that is phosphorylated at positions 4,5 in plasma membrane or position 4 in the golgi
  2. adaptors AP1 and AP2 recognize PI and receptors attached to the cargo
  3. clathrin coat binds adaptors creating clathrin cage. dynamin (GTPase) ring closes to create vesicle in PM only.
  4. PI phosphatases weaken adaptor binding. chaperones auxilin (type of DNAJ) and HSC70 makes clathrin monomers uncoating the vesicle.
• in COPII and COPI there is a timer (GTP hydrolysis in Sar1 or ARF1 via adaptors (Sec23/24 or beta/delta/gamma) where the proteins stop interacting with adaptors, allowing for the uncoating of the vesicles.
• Protein quality control mechanisms in the ER are crucial as these proteins are sent to other areas of the cell
• The secretory pathway involves the trafficking of proteins across the cell, a complex mechanism with many vesicles moving in different directions
• The process from ER (protein synthesis) to golgi (many PTM) involves vesicles seeping into the plasma membrane or endosome, with vesicles formed in the plasma membrane bringing cargo into the interior of the cell, which can be degraded in the lysosome
• Different types of vesicles are involved in vesicle transport, such as COP-II vesicles going from ER to golgi, known as anterograde transport
• COP-I vesicles facilitate cargo from the golgi back into the ER, aiding in retrograde transport
• Clathrin-coated vesicles play a role in transporting cargo from the exterior of the cell into the interior, as well as from the golgi into the lysosome or endosome
• Vesicle formation involves initiation, coat formation, fission, and uncoating processes
• In the COP-II process, proteins are recognized by calnexin and calreticulin, with Sar1 (GTPase protein) playing a crucial role in vesicle formation
• ER exit (COP II) involves recognizing motifs like two phenylalanine at the c-terminus for transmembrane proteins
• ER retrieval (COP I vesicles) is guided by the KDEL motif, allowing proteins to return to the ER if sent to the golgi by mistake
• CCV fission requires dynamin, a GTPase, to separate the vesicle from the ER compartment
• Clathrin-coated vesicles are uncoated by chaperones like HSC70 and auxilin, allowing the vesicle to travel and fuse with the next compartment