scholarly journals All aboard!: The secretory pathway

2003 ◽  
Vol 25 (3) ◽  
pp. 10-12
Author(s):  
Stephen High ◽  
Samuel G. Crawshaw
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Synthesis ◽  
Membrane Protein ◽  
Cell Surface ◽  
Secretory Pathway ◽  
Entry Point ◽  
Eukaryotic Cells ◽  
Secretory Cells ◽  
Intracellular Membrane ◽  
Biosynthetic Activity

The endoplasmic reticulum (ER) is a major subcellular feature of most eukaryotic cells, and in specialized secretory cells, like those of the pancreas, it densely packs most of the cell. It is the de facto entry point into the secretory pathway and one of its key functions is to provide an extensive intracellular membrane network that supports protein synthesis (Figure 1). This biosynthetic activity is highlighted by the large number of ribosomes that are bound tightly to much of its surface, and it is these ribosomes that synthesize the secretory and membrane protein cargoes that are ultimately destined for export from the ER to the cell surface via the Golgi complex1,2.

scholarly journals Two Endoplasmic Reticulum (ER) Membrane Proteins That Facilitate ER-to-Golgi Transport of Glycosylphosphatidylinositol-anchored Proteins

1999 ◽  
Vol 10 (4) ◽  
pp. 1043-1059 ◽  
Author(s):  
Wolfgang P. Barz ◽  
Peter Walter
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Surface ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Protein Translocation ◽  
Sequence Similarity ◽  
Surface Proteins ◽  
Golgi Transport ◽  
Er Membrane

Many eukaryotic cell surface proteins are anchored in the lipid bilayer through glycosylphosphatidylinositol (GPI). GPI anchors are covalently attached in the endoplasmic reticulum (ER). The modified proteins are then transported through the secretory pathway to the cell surface. We have identified two genes inSaccharomyces cerevisiae, LAG1 and a novel gene termed DGT1 (for “delayed GPI-anchored protein transport”), encoding structurally related proteins with multiple membrane-spanning domains. Both proteins are localized to the ER, as demonstrated by immunofluorescence microscopy. Deletion of either gene caused no detectable phenotype, whereas lag1Δ dgt1Δ cells displayed growth defects and a significant delay in ER-to-Golgi transport of GPI-anchored proteins, suggesting thatLAG1 and DGT1 encode functionally redundant or overlapping proteins. The rate of GPI anchor attachment was not affected, nor was the transport rate of several non–GPI-anchored proteins. Consistent with a role of Lag1p and Dgt1p in GPI-anchored protein transport, lag1Δ dgt1Δ cells deposit abnormal, multilayered cell walls. Both proteins have significant sequence similarity to TRAM, a mammalian membrane protein thought to be involved in protein translocation across the ER membrane. In vivo translocation studies, however, did not detect any defects in protein translocation in lag1Δ dgt1Δcells, suggesting that neither yeast gene plays a role in this process. Instead, we propose that Lag1p and Dgt1p facilitate efficient ER-to-Golgi transport of GPI-anchored proteins.

scholarly journals Divergent Regulation of Protein Synthesis in the Cytosol and Endoplasmic Reticulum Compartments of Mammalian Cells

2008 ◽  
Vol 19 (2) ◽  
pp. 623-632 ◽  
Author(s):  
Samuel B. Stephens ◽  
Christopher V. Nicchitta
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Synthesis ◽  
Steady State ◽  
Mammalian Cells ◽  
Signal Sequence ◽  
Integral Membrane Protein ◽  
Cellular Protein ◽  
Eukaryotic Cells ◽  
Global Protein Synthesis ◽  
Tightly Coupled

In eukaryotic cells, mRNAs encoding signal sequence-bearing proteins undergo translation-dependent trafficking to the endoplasmic reticulum (ER), thereby restricting secretory and integral membrane protein synthesis to the ER compartment. However, recent studies demonstrating that mRNAs encoding cytosolic/nucleoplasmic proteins are represented on ER-bound polyribosomes suggest a global role for the ER in cellular protein synthesis. Here, we examined the steady-state protein synthesis rates and compartmental distribution of newly synthesized proteins in the cytosol and ER compartments. We report that ER protein synthesis rates exceed cytosolic protein synthesis rates by 2.5- to 4-fold; yet, completed proteins accumulate to similar levels in the two compartments. These data suggest that a significant fraction of cytosolic proteins undergo synthesis on ER-bound ribosomes. The compartmental differences in steady-state protein synthesis rates correlated with a divergent regulation of the tRNA aminoacylation/deacylation cycle. In the cytosol, two pathways were observed to compete for aminoacyl-tRNAs—protein synthesis and aminoacyl-tRNA hydrolysis—whereas on the ER tRNA deacylation is tightly coupled to protein synthesis. These findings identify a role for the ER in global protein synthesis, and they suggest models where compartmentalization of the tRNA acylation/deacylation cycle contributes to the regulation of global protein synthesis rates.

scholarly journals Aberrant substrate engagement of the ER translocon triggers degradation by the Hrd1 ubiquitin ligase

2012 ◽  
Vol 197 (6) ◽  
pp. 761-773 ◽  
Author(s):  
Eric M. Rubenstein ◽  
Stefan G. Kreft ◽  
Wesley Greenblatt ◽  
Robert Swanson ◽  
Mark Hochstrasser
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Ubiquitin Ligase ◽  
Apolipoprotein B ◽  
Secretory Pathway ◽  
Protein A ◽  
Proteasomal Degradation ◽  
Membrane Localization ◽  
General Role

Little is known about quality control of proteins that aberrantly or persistently engage the endoplasmic reticulum (ER)-localized translocon en route to membrane localization or the secretory pathway. Hrd1 and Doa10, the primary ubiquitin ligases that function in ER-associated degradation (ERAD) in yeast, target distinct subsets of misfolded or otherwise abnormal proteins based primarily on degradation signal (degron) location. We report the surprising observation that fusing Deg1, a cytoplasmic degron normally recognized by Doa10, to the Sec62 membrane protein rendered the protein a Hrd1 substrate. Hrd1-dependent degradation occurred when Deg1-Sec62 aberrantly engaged the Sec61 translocon channel and underwent topological rearrangement. Mutations that prevent translocon engagement caused a reversion to Doa10-dependent degradation. Similarly, a variant of apolipoprotein B, a protein known to be cotranslocationally targeted for proteasomal degradation, was also a Hrd1 substrate. Hrd1 therefore likely plays a general role in targeting proteins that persistently associate with and potentially obstruct the translocon.

scholarly journals Differential effects of nitrated ricin and nitrated and dithionite-reduced ricin on protein-synthesis inhibition and transmembrane tramsport in eukaryotic cells

1980 ◽  
Vol 188 (3) ◽  
pp. 941-944 ◽  
Author(s):  
P N Dalrymple ◽  
L L Houston
Keyword(s):  
Protein Synthesis ◽  
Cell Surface ◽  
Indirect Immunofluorescence ◽  
Cultured Cells ◽  
Sodium Dithionite ◽  
Eukaryotic Cells ◽  
Protein Synthesis Inhibition ◽  
Inhibiting Protein ◽  
A Chain

Ricin, was nitrated with tetranitromethane and reduced with sodium dithionite. Of the 8.0 nitro groups incorporated, 3.2 were on the A chain and 5.1 were on the B chain. Nitrated ricin1 was somewhat less active than nitrated and reduced ricin1 in inhibiting protein synthesis in vitro, but both were highly inhibitory. However, the modified toxins were less than 1% as active as ricin in inhibiting protein synthesis in cultured cells. Indirect immunofluorescence assays demonstrated tha both modified toxins were specifically bound to the cell surface and could be displaced by galactose.

Ultrastructural changes accompany inhibition of proteoglycan synthesis in chondrocytes by Cyclofenil diphenol

1989 ◽  
Vol 92 (2) ◽  
pp. 271-280 ◽  
Author(s):  
C.A. Lancaster ◽  
P.R. Fryer ◽  
S. Griffiths ◽  
R.M. Mason
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Synthesis ◽  
Electron Density ◽  
Lipid Droplets ◽  
Secretory Pathway ◽  
Ultrastructural Changes ◽  
Proteoglycan Synthesis ◽  
Filamentous Material

Cyclofenil diphenol, a weak non-steroidal oestrogen, profoundly inhibits [35S]proteoglycan synthesis in cultures of Swarm chondrosarcoma chondrocytes under conditions in which protein synthesis is only marginally reduced. In the present experiments it was shown that after a 40-min treatment with Cyclofenil diphenol (90 micrograms ml-1) most of the normally abundant Golgi stacks in these cells disappeared and after 60 min they were absent. After 2–3 h treatment the cisternae of the endoplasmic reticulum (ER) were grossly distended and transformed into large ribosome-studded vesicles containing flocculent and filamentous material. These changes were dependent on the concentration of Cyclofenil and were fully reversible within 21 h of withdrawing the drug. The ultrastructural changes differed in some aspects if protein synthesis was blocked with cycloheximide for 15 min or 180 min before and during treatment with Cyclofenil. The Golgi disappeared but the ER cisternae, though distended, formed a continuous network and swollen ribosome-studded vesicles did not develop. However, non-membrane-bounded structures containing lipid droplets and material of low electron density developed in the cytoplasm under these conditions. The ultrastructural changes induced by Cyclofenil differ from those induced by monensin and diethylcarbamazine, suggesting that the drug acts at a different point in the secretory pathway for macromolecules.

scholarly journals The yeast p24 complex regulates GPI-anchored protein transport and quality control by monitoring anchor remodeling

2011 ◽  
Vol 22 (16) ◽  
pp. 2924-2936 ◽  
Author(s):  
Guillaume A. Castillon ◽  
Auxiliadora Aguilera-Romero ◽  
Javier Manzano-Lopez ◽  
Sharon Epstein ◽  
Kentaro Kajiwara ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Cell Surface ◽  
Intracellular Transport ◽  
Protein Transport ◽  
Eukaryotic Cells ◽  
Secretory Proteins ◽  
Dependent Manner ◽  
Gpi Anchor ◽  
The Status

Glycosylphosphatidylinositol (GPI)-anchored proteins are secretory proteins that are attached to the cell surface of eukaryotic cells by a glycolipid moiety. Once GPI anchoring has occurred in the lumen of the endoplasmic reticulum (ER), the structure of the lipid part on the GPI anchor undergoes a remodeling process prior to ER exit. In this study, we provide evidence suggesting that the yeast p24 complex, through binding specifically to GPI-anchored proteins in an anchor-dependent manner, plays a dual role in their selective trafficking. First, the p24 complex promotes efficient ER exit of remodeled GPI-anchored proteins after concentration by connecting them with the COPII coat and thus facilitates their incorporation into vesicles. Second, it retrieves escaped, unremodeled GPI-anchored proteins from the Golgi to the ER in COPI vesicles. Therefore the p24 complex, by sensing the status of the GPI anchor, regulates GPI-anchored protein intracellular transport and coordinates this with correct anchor remodeling.

scholarly journals CARTS biogenesis requires VAP–lipid transfer protein complexes functioning at the endoplasmic reticulum–Golgi interface

2015 ◽  
Vol 26 (25) ◽  
pp. 4686-4699 ◽  
Author(s):  
Yuichi Wakana ◽  
Richika Kotake ◽  
Nanako Oyama ◽  
Motohide Murate ◽  
Toshihide Kobayashi ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Cell Surface ◽  
Lipid Transfer Protein ◽  
Protein Complexes ◽  
Lipid Transfer ◽  
Trans Golgi Network ◽  
Transfer Protein ◽  
Membrane Contact Sites ◽  
Golgi Network

Vesicle-associated membrane protein–associated protein (VAP) is an endoplasmic reticulum (ER)-resident integral membrane protein that controls a nonvesicular mode of ceramide and cholesterol transfer from the ER to the Golgi complex by interacting with ceramide transfer protein and oxysterol-binding protein (OSBP), respectively. We report that VAP and its interacting proteins are required for the processing and secretion of pancreatic adenocarcinoma up-regulated factor, whose transport from the trans-Golgi network (TGN) to the cell surface is mediated by transport carriers called “carriers of the trans-Golgi network to the cell surface” (CARTS). In VAP-depleted cells, diacylglycerol level at the TGN was decreased and CARTS formation was impaired. We found that VAP forms a complex with not only OSBP but also Sac1 phosphoinositide phosphatase at specialized ER subdomains that are closely apposed to the trans-Golgi/TGN, most likely reflecting membrane contact sites. Immobilization of ER–Golgi contacts dramatically reduced CARTS production, indicating that association–dissociation dynamics of the two membranes are important. On the basis of these findings, we propose that the ER–Golgi contacts play a pivotal role in lipid metabolism to control the biogenesis of transport carriers from the TGN.

scholarly journals Folding and Insertion of Transmembrane Helices at the ER

2021 ◽  
Vol 22 (23) ◽  
pp. 12778
Author(s):  
Paul Whitley ◽  
Brayan Grau ◽  
James C. Gumbart ◽  
Luis Martínez-Gil ◽  
Ismael Mingarro
Keyword(s):  
Amino Acids ◽  
Endoplasmic Reticulum ◽  
Entry Point ◽  
Eukaryotic Cells ◽  
Helical Conformation ◽  
Aqueous Environment ◽  
Transmembrane Helices ◽  
Endomembrane System ◽  
Membrane Bilayer ◽  
Er Membrane

In eukaryotic cells, the endoplasmic reticulum (ER) is the entry point for newly synthesized proteins that are subsequently distributed to organelles of the endomembrane system. Some of these proteins are completely translocated into the lumen of the ER while others integrate stretches of amino acids into the greasy 30 Å wide interior of the ER membrane bilayer. It is generally accepted that to exist in this non-aqueous environment the majority of membrane integrated amino acids are primarily non-polar/hydrophobic and adopt an α-helical conformation. These stretches are typically around 20 amino acids long and are known as transmembrane (TM) helices. In this review, we will consider how transmembrane helices achieve membrane integration. We will address questions such as: Where do the stretches of amino acids fold into a helical conformation? What is/are the route/routes that these stretches take from synthesis at the ribosome to integration through the ER translocon? How do these stretches ‘know’ to integrate and in which orientation? How do marginally hydrophobic stretches of amino acids integrate and survive as transmembrane helices?

scholarly journals Enhanced ER proteostasis and temperature differentially impact the mutational tolerance of influenza hemagglutinin

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Angela M Phillips ◽  
Michael B Doud ◽  
Luna O Gonzalez ◽  
Vincent L Butty ◽  
Yu-Shan Lin ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Experimental Evidence ◽  
Secretory Pathway ◽  
Immune Escape ◽  
Structural Elements ◽  
Chemical Methods ◽  
Viral Membrane ◽  
Influenza Hemagglutinin ◽  
And Function

We systematically and quantitatively evaluate whether endoplasmic reticulum (ER) proteostasis factors impact the mutational tolerance of secretory pathway proteins. We focus on influenza hemaggluttinin (HA), a viral membrane protein that folds in the host’s ER via a complex pathway. By integrating chemical methods to modulate ER proteostasis with deep mutational scanning to assess mutational tolerance, we discover that upregulation of ER proteostasis factors broadly enhances HA mutational tolerance across diverse structural elements. Remarkably, this proteostasis network-enhanced mutational tolerance occurs at the same sites where mutational tolerance is most reduced by propagation at fever-like temperature. These findings have important implications for influenza evolution, because influenza immune escape is contingent on HA possessing sufficient mutational tolerance to evade antibodies while maintaining the capacity to fold and function. More broadly, this work provides the first experimental evidence that ER proteostasis mechanisms define the mutational tolerance and, therefore, the evolution of secretory pathway proteins.

scholarly journals How many lives does CLIMP-63 have?

2015 ◽  
Vol 43 (2) ◽  
pp. 222-228 ◽  
Author(s):  
Patrick A. Sandoz ◽  
F. Gisou van der Goot
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Cell Surface ◽  
Transmembrane Protein ◽  
The Other ◽  
Type Ii ◽  
Post Translational Modifications ◽  
Cytosolic Domain ◽  
In Trans

In 1995, in the Biochemical Society Transactions, Mundy published the first review on CLIMP-63 (cytoskeleton-linking membrane protein 63) or CKPA4 (cytoskeleton-associated protein 4), initially just p63 [1]. Here we review the following 20 years of research on this still mysterious protein. CLIMP-63 is a type II transmembrane protein, the cytosolic domain of which has the capacity to bind microtubules whereas the luminal domain can form homo-oligomeric complexes, not only with neighbouring molecules but also, in trans, with CLIMP-63 molecules on the other side of the endoplasmic reticulum (ER) lumen, thus promoting the formation of ER sheets. CLIMP-63 however also appears to have a life at the cell surface where it acts as a ligand-activated receptor. The still rudimentary information of how CLIMP-63 fulfills these different roles, what these are exactly and how post-translational modifications control them, will be discussed.

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