Exocytosis and Endocytosis PDF

MODULE 16 Exocytosis versus Endocytosis endocytosis: brings molecules and other substances into the cell; associated with the formation and budding of vesicles from the plasma membrane exocytosis: the exit of molecules from the cell; associated with the fusion of vesicles originating from intr...

MODULE 16 Exocytosis versus Endocytosis endocytosis: brings molecules and other substances into the cell; associated with the formation and budding of vesicles from the plasma membrane exocytosis: the exit of molecules from the cell; associated with the fusion of vesicles originating from intracellular organelles with the plasma membrane Introduction to Exocytosis transport of molecules out of the cells through the fusion of vesicles with the plasma membrane three classes of proteins from TGN to (1) lysosomes, (2) secretory vesicles, or (3) immediate delivery to the cell surface constitutive and regulated (as in specialized secreting cells) in unpolarized cells, soluble proteins from the Golgi are carried by constitutive pathway to the cell surface, unless these are retained as ER or Golgi residents, or sorted for the lysosomes or regulated secretion Steps of Exocytosis four steps for constitutive exocytosis, five for regulated exocytosis: trafficking: vesicle transport to the cell membrane on microtubules tethering: vesicle’s contact with the cell membrane docking: the vesicle and cell membranes attach; the two phospholipid bilayers merge priming: only in regulated exocytosis fusion: the vesicle membrane fully fuses with the cell membrane, and the result is the release of vesicle contents into the extracellular space Coated Vesicles vesicle budding is mediated by protein coats, structures that cycle on and off membranes coats have adaptor proteins that recognize cargo molecules and cage/coat proteins assembling a lattice on the membranes to collect adaptor-cargo complexes the coat deforms membranes into buds and selects the cargo MODULE 16 coat proteins are recruited from the cytosol by Arf1/Sar1 small GTPases three types of coats: clathrin-coated vesicles mediate transport between the Golgi and plasma membrane; COPI- (retrograde) and COPII-coated vesicles mediate transport between the ER and Golgi Formation of Secretory Vesicles some hormones, neuropeptides, and hydrolytic enzymes are synthesized as inactive precursors; the proteolysis to active forms begins in the TGN and continues in the vesicles and extracellular fluid after secretion for proteins synthesized as pre-pro-proteins, the pre-peptide is the ER signal peptide cleaved in rER some signaling molecules are polyproteins that contain multiple copies of the same amino acid sequence; some peptides are synthesized as parts of a single polyprotein, a precursor for multiple products Sorting Signals polarized epithelial cells: proteins are sorted in the TGN to vesicles for different plasma membrane domains proteins for the basolateral membrane contain specific signals recognized by coat proteins the apical plasma membrane is enriched in glycosphingolipids and GPI-anchored proteins (that associate with glycosphingolipids in lipid rafts in the TGN) Coat Assembly GTPases ADP-ribosylation factor (ARF) GTPases: small GTPases, activated by guanine nucleotide exchange factors (GEFs); inactivated by GTPase-activating proteins (GAPs) ARFs recruit coat proteins, lipid-modifying enzymes, scaffold proteins Sar1 regulates anterograde transport of proteins (ER → Golgi) through COPII vesicles ARF1 regulates retrograde transport (Golgi → ER) through COPI vesicles; it also regulates recruitment of clathrin Pinching-off and Uncoating dynamin binds to a forming bud on the membrane and assembles into a ring around the neck of the bud MODULE 16 the ring recruits other proteins that together with dynamin destabilize the membrane once the vesicle is released from the membrane, the clathrin coat is lost an uncoating ATPase peels off the coat Transport, Tethering, Docking vesicles traffic: along microtubules with kinesin/dynein motors; or myosin motors along the actin network transport vesicles have surface markers identifying their origin/cargo target membranes have receptors for vesicle markers; the recognition step is controlled by SNAREs and Rab GTPases Rab effectors accomplish the tethering/docking of appropriate vesicles to the target membranes Fusion at docking: proteins on the two lipid bilayers interact for fusion, water is displaced from the surface of the two membranes SNAREs are transmembrane proteins; there are vesicle membrane SNAREs (v-SNAREs) and target membrane SNAREs (t-SNAREs) when a v-SNARE interacts with a t-SNARE, the helical domains of one wrap around the helical domains of the other to form trans-SNARE complexes; this provides energy to pull the membranes together and squeeze water from the interface Release of Secretory Vesicle Cargo for regulated exocytosis, after docking, the secretory vesicles need to be primed to fuse and release their contents the fusion and release of cargo is triggered by a signal; the signal could be a chemical messenger (e.g., a hormone leading to intracellular signals such as cytosolic Ca²⁺) at nerve terminals, the signal is an action potential causing a Ca²⁺ influx; Ca²⁺ binding to sensors triggers vesicles fusion with the plasma membrane and release of cargo to the extracellular space MODULE 16 Putting It Together proteins are secreted by exocytosis in a constitutive or regulated fashion; secretory vesicles bud from the TGN; secretory proteins condense during the formation and maturation of secretory vesicles there are four steps in constitutive exocytosis and five in regulated exocytosis; in regulated exocytosis, in addition to trafficking, tethering, docking, and fusion, there is priming (i.e., molecules are stored in vesicles until a signal is received) in polarized cells, the transport from the TGN to the plasma membrane selectively delivers different sets of molecules to the different domains of the plasma membrane transport vesicles bud from coated regions of the donor membrane; the coat helps to collect specific cargo and drives the formation of the vesicle; the process is regulated by small GTPases (e.g., Sar1, ARF) the docking/tethering requires the small GTPases Rab that bind to specific effectors, and the fusion requires v-SNAREs and t-SNAREs to form trans-SNARE complexes How to Kill Cancer Cells degranulation: release of antimicrobial cytotoxic or other molecules from secretory vesicles utilized by granulocytes and mast cells also used by lymphocytes such as natural killer (NK) cells and cytotoxic T cells to destroy invading microorganisms cytotoxic T cells and NK cells release molecules like perforin and granzymes by a process of directed exocytosis to kill infected target cells (or cancer cells)