Landmark discoveries in intracellular transport and secretion (original) (raw)
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Proteins involved in vesicular transport and membrane fusion
Current Opinion in Cell Biology, 1991
In the past year, new information about proteins involved in vesicular transport has been plentiful. Particularly noteworthy are the complementary findings that Secl7p is required for vesicle consumption in endoplasmic reticulum-to-Go@ transport in yeast and that an analogous activity in mammalian cells, termed SNAP, is required for transport from the cis to the medial cisternae of the Colgi apparatus.
Characteristics of endoplasmic reticulum-derived transport vesicles
The Journal of Cell Biology, 1994
We have isolated vesicles that mediate protein transport from the ER to Golgi membranes in perforated yeast. These vesicles, which form de novo during in vitro incubations, carry lumenal and membrane proteins that include core-glycosylated pro-~x-factor, Betl, Sec22, and Bosl, but not ER-resident Kar2 or Sec61 proteins. Thus, lumenal and membrane proteins in the ER are sorted prior to transport vesicle scission. Inhibition of Yptlp-function, which prevents newly formed vesicles from docking to cis-Golgi membranes, was used to block transport. Vesicles that accumulate are competent for fusion with cis-Golgi membranes, but not with ER membranes, and thus are functionally committed to vectorial transport. A 900-fold enrichment was developed using differential centrifugation and a series of velocity and equilibrium density gradients. Electron microscopic analysis shows a uniform population of 60 nm vesicles that lack peripheral protein coats. Quantitative Western blot analysis indicates that protein markers of cytosol and cellular membranes are depleted throughout the purification, whereas the synaptobrevin-like Betl, Sec22, and Bosl proteins are highly enriched. Uncoated E___RR-derived transport vesicles (ERV) contain twelve major proteins that associate tightly with the membrane. The ERV proteins may represent abundant cargo and additional targeting molecules.
Cargo Capture and Bulk Flow in the Early Secretory Pathway
Annual review of cell and developmental biology, 2016
Transport of newly synthesized proteins from the endoplasmic reticulum (ER) to the Golgi complex is highly selective. As a general rule, such transport is limited to soluble and membrane-associated secretory proteins that have reached properly folded and assembled conformations. To secure the efficiency, fidelity, and control of this crucial transport step, cells use a combination of mechanisms. The mechanisms are based on selective retention of proteins in the ER to prevent uptake into transport vesicles, on selective capture of proteins in COPII carrier vesicles, on inclusion of proteins in these vesicles by default as part of fluid and membrane bulk flow, and on selective retrieval of proteins from post-ER compartments by retrograde vesicle transport. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 32 is October 06, 2016. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.
Journal of Neurocytology, 2000
We have recently proposed a mechanism to describe secretion, a fundament process in all cells. That hypothesis, called porocytosis, embodies all available data, and encompasses both forms of secretion, i.e., vesicular and constitutive. The current accepted view of exocytotic secretion involves the physical fusion of vesicle-and plasma membranes. However, that hypothesized mechanism does not fit all available physiological data . Energetics of apposed lipid bilayers do not favor unfacilitated fusion. Calcium ion levels are elevated in microdomains at levels of 10 − 4 -10 − 3 M for 1 ms or less, with the calcium ions showing limited lateral mobility at the site of secretion (Llinas et al., 1992. We consider that calcium ions, whose mobility is restricted in space and time, establish "salt-bridges'' among adjacent lipid molecules, and establishes transient pores that span the vesicle and plasma membrane lipid bilayers; the lifetime of that transient pore being completely dependent on duration of sufficient calcium ion levels.
Discovery of the Cell Secretion Machinery
Journal of Biomedical Nanotechnology, 2007
Cellular secretion is fundamental to the very existence of an organism, regulating important physiological functions such as reproduction, digestion, energy production, growth, neurotransmission, hormone release, water and ion transport, etc., all required for the survival and maintenance of homeostasis within an organism. Understanding how cells secrete has therefore been of paramount importance, and only in the past decade, the molecular mechanism of the process has come to light with the discovery of the 'porosome,' the universal secretion machinery. Porosomes are supramolecular nanometer-size structures at the cell plasma membrane, where secretory vesicles fuse to release their contents such as neurotransmitters, hormones, or enzymes from the cell. Porosomes were first discovered at the plasma membrane of the digestive enzyme-secreting, live acinar cells of the exocrine pancreas, using atomic force microscopy. Immuno-atomic force microscopy, has conclusively demonstrated the release of digestive enzymes through the porosome opening in pancreatic acinar cells. Subsequent studies using electron microscopy, has further confirmed the presence of porosomes, and determined their function as the cells secretory apparatus. Studies have since determined the structure and dynamics of porosomes at nanometer resolution in live cells, their chemical composition, and their structural and functional reconstitution in artificial lipid membrane preparations. Porosomes have been determined to be the universal secretory machinery in cells, and besides pancreatic acinar cells, have been demonstrated in every secretory cell examined such as neurons, growth hormone cells of the pituitary gland, chromaffin cells, mast cells, and the insulin-secreting-cells of endocrine pancreas. The discovery of the porosome finally provides a molecular understanding of the secretory process in cells.
The Journal of Cell Biology, 1996
In order for secretion to progress, ERderived transport vesicles must target to, and fuse with the cis-Golgi compartment. These processes have been reconstituted using highly enriched membrane fractions and partially purified soluble components. The functionally active yeast Golgi membranes that have been purified are highly enriched in the cis-Golgi marker enzymes al,6 mannosyltransferase and GDPase. Fusion of transport vesicles with these membranes requires both GTP and ATP hydrolysis, and depends on cytosolic and peripheral membrane proteins. At least two protein fractions from yeast cytosol are required for the reconstitution of ER-derived vesicle fusion. Soluble fractions prepared from temperature-sensitive mutants revealed requirements for the Yptlp, Secl9p, Slylp, Sec7p, and Usol proteins. A model for the sequential involvement of these components in the targeting and fusion reaction is proposed.
Biogenesis of COPI-coated transport vesicles
FEBS Letters, 1997
Biosynthetic protein transport and sorting along the secretory pathway represents the last step in biosynthesis of a variety of proteins. Proteins destined for delivery to the cell surface are inserted cotranslationally into the endoplasmic reticulum (ER) and, after their correct folding, are transported out of the ER towards their final destinations. The successive compartments of the secretory pathway are connected by vesicular shuttles that mediate delivery of cargo. The formation of these carrier vesicles depends on the recruitment of cytosolic coat proteins that are thought to act as a mechanical device to shape a flattened donor membrane into a spherical vesicle. A general molecular machinery that mediates targeting and fusion of carrier vesicles has also been identified. This review is focused on COPI-coated vesicles that operate in protein transport within the early secretory pathway. Rather than representing a general overview of the role of COPI-coated vesicles, this mini-review will discuss mechanisms specifically related to the biogenesis of COPI-coated vesicles: (i) a possible role of phospholipase D in the formation of COPI-coated vesicles, (ii) a functional role of a novel family of transmembrane proteins, the p24 family, in the initiation of COPI assembly, and (iii) the direction COPI-coated vesicles may take within the early secretory pathway.