scholarly journals Transmembrane topology of mammalian ORMDL proteins in the endoplasmic reticulum as revealed by the substituted cysteine accessibility method (SCAM™)

2019 ◽  
Vol 1867 (4) ◽  
pp. 382-395
Author(s):  
Deanna Davis ◽  
John Suemitsu ◽  
Binks Wattenberg
Keyword(s):  
Endoplasmic Reticulum ◽  
Transmembrane Topology ◽  
Substituted Cysteine Accessibility

scholarly journals Mechanism of Residence of Cytochrome B(5), a Tail-Anchored Protein, in the Endoplasmic Reticulum

2000 ◽  
Vol 148 (5) ◽  
pp. 899-914 ◽  
Author(s):  
Emanuela Pedrazzini ◽  
Antonello Villa ◽  
Renato Longhi ◽  
Alessandra Bulbarelli ◽  
Nica Borgese
Keyword(s):  
Endoplasmic Reticulum ◽  
Lipid Bilayer ◽  
Cytochrome B ◽  
Golgi Complex ◽  
Intracellular Trafficking ◽  
Secretory Pathway ◽  
Transmembrane Topology ◽  
Glycosylation Sites ◽  
Cytosolic Domain

Endoplasmic reticulum (ER) proteins maintain their residency by static retention, dynamic retrieval, or a combination of the two. Tail-anchored proteins that contain a cytosolic domain associated with the lipid bilayer via a hydrophobic stretch close to the COOH terminus are sorted within the secretory pathway by largely unknown mechanisms. Here, we have investigated the mode of insertion in the bilayer and the intracellular trafficking of cytochrome b(5) (b[5]), taken as a model for ER-resident tail-anchored proteins. We first demonstrated that b(5) can acquire a transmembrane topology posttranslationally, and then used two tagged versions of b(5), N-glyc and O-glyc b(5), containing potential N- and O-glycosylation sites, respectively, at the COOH-terminal lumenal extremity, to discriminate between retention and retrieval mechanisms. Whereas the N-linked oligosaccharide provided no evidence for retrieval from a downstream compartment, a more stringent assay based on carbohydrate acquisition by O-glyc b(5) showed that b(5) gains access to enzymes catalyzing the first steps of O-glycosylation. These results suggest that b(5) slowly recycles between the ER and the cis-Golgi complex and that dynamic retrieval as well as retention are involved in sorting of tail-anchored proteins.

scholarly journals Different Transmembrane Domains Associate with Distinct Endoplasmic Reticulum Components during Membrane Integration of a Polytopic Protein

2002 ◽  
Vol 13 (12) ◽  
pp. 4114-4129 ◽  
Author(s):  
Suzanna L. Meacock ◽  
Fabienne J.L. Lecomte ◽  
Samuel G. Crawshaw ◽  
Stephen High
Keyword(s):  
Endoplasmic Reticulum ◽  
Amino Acid ◽  
Polypeptide Chain ◽  
Transmembrane Domain ◽  
Insertion Site ◽  
Transmembrane Domains ◽  
Membrane Insertion ◽  
Relative Location ◽  
Transmembrane Topology ◽  
Nascent Chain

We have been studying the insertion of the seven transmembrane domain (TM) protein opsin to gain insights into how the multiple TMs of polytopic proteins are integrated at the endoplasmic reticulum (ER). We find that the ER components associated with the first and second TMs of the nascent opsin polypeptide chain are clearly distinct. The first TM (TM1) is adjacent to the α and β subunits of the Sec61 complex, and a novel component, a protein associated with the ER translocon of 10 kDa (PAT-10). The most striking characteristic of PAT-10 is that it remains adjacent to TM1 throughout the biogenesis and membrane integration of the full-length opsin polypeptide. TM2 is also found to be adjacent to Sec61α and Sec61β during its membrane integration. However, TM2 does not form any adducts with PAT-10; rather, a transient association with the TRAM protein is observed. We show that the association of PAT-10 with opsin TM1 does not require theN-glycosylation of the nascent chain and occurs irrespective of the amino acid sequence and transmembrane topology of TM1. We conclude that the precise makeup of the ER membrane insertion site can be distinct for the different transmembrane domains of a polytopic protein. We find that the environment of a particular TM can be influenced by both the “stage” of nascent chain biosynthesis reached, and the TM's relative location within the polypeptide.

scholarly journals An investigation of the transverse topology of bilirubin UDP-glucuronosyltransferase in rat hepatic endoplasmic reticulum

1989 ◽  
Vol 259 (2) ◽  
pp. 617-620 ◽  
Author(s):  
S R Shepherd ◽  
S J Baird ◽  
T Hallinan ◽  
B Burchell
Keyword(s):  
Endoplasmic Reticulum ◽  
Sodium Deoxycholate ◽  
Immunoblot Analysis ◽  
Proteinase K ◽  
Transmembrane Topology ◽  
Working Model ◽  
Hepatic Microsomes ◽  
Udp Glucuronosyltransferase ◽  
Cytoplasmic Side

Bilirubin UDP-glucuronosyltransferase (UDPGT) activity in sealed hepatic microsomes from clofibrate-treated rats was highly latent and was fully expressed by disruption of vesicles with detergents. Antibodies raised against purified bilirubin UDPGT were used to study the transmembrane orientation of the protein to provide a molecular understanding of the UDPGT latency. Immunoblot analysis of sealed microsomes, and microsomes after treatment with proteinases, showed that only a small portion of the protein resides on the cytoplasmic side of the microsomal vesicles. Treatment of microsomes with sodium deoxycholate allowed subtilisin and proteinase K to cleave the transferase, causing loss of activity and the release of smaller immunodetectable peptides. Treatment of the purified bilirubin UDPGT with peptide N-glycosidase F indicated that the enzyme was a glycoprotein. A working model of the transmembrane topology of bilirubin UDPGT is described.

scholarly journals A C-terminal amphipathic helix is necessary for the in vivo tubule-shaping function of a plant reticulon

2016 ◽  
Vol 113 (39) ◽  
pp. 10902-10907 ◽  
Author(s):  
Emily Breeze ◽  
Natasha Dzimitrowicz ◽  
Verena Kriechbaumer ◽  
Rhiannon Brooks ◽  
Stanley W. Botchway ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Structural Feature ◽  
Mechanism Of Action ◽  
Membrane Curvature ◽  
Amphipathic Helix ◽  
Transmembrane Topology ◽  
C Terminus ◽  
Er Membrane

Reticulons (RTNs) are a class of endoplasmic reticulum (ER) membrane proteins that are capable of maintaining high membrane curvature, thus helping shape the ER membrane into tubules. The mechanism of action of RTNs is hypothesized to be a combination of wedging, resulting from the transmembrane topology of their conserved reticulon homology domain, and scaffolding, arising from the ability of RTNs to form low-mobility homo-oligomers within the membrane. We studied the plant RTN isoform RTN13, which has previously been shown to locate to ER tubules and the edges of ER cisternae and to induce constrictions in ER tubules when overexpressed, and identified a region in the C terminus containing a putative amphipathic helix (APH). Here we show that deletion of this region or disruption of the hydrophobic face of the predicted helix abolishes the ability of RTN13 to induce constrictions of ER tubules in vivo. These mutants, however, still retain their ability to interact and form low-mobility oligomers in the ER membrane. Hence, our evidence indicates that the conserved APH is a key structural feature for RTN13 function in vivo, and we propose that RTN, like other membrane morphogens, rely on APHs for their function.

In Vitro Induction of Crystals of MT Protein by Vinblastine-Colcemid in Mouse Spermatids

1974 ◽  
Vol 32 ◽  
pp. 132-133
Author(s):  
John J. Wolosewick ◽  
John H. D. Bryan
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Endoplasmic Reticulum ◽  
Nuclear Ring ◽  
Rough Er ◽  
Distal Direction ◽  
In Vitro Induction

Early in spermiogenesis the manchette is rapidly assembled in a distal direction from the nuclear-ring-densities. The association of vesicles of smooth endoplasmic reticulum (SER) and the manchette microtubules (MTS) has been reported. In the mouse, osmophilic densities at the distal ends of the manchette are the organizing centers (MTOCS), and are associated with the SER. Rapid MT assembly and the lack of rough ER suggests that there is an existing pool of MT protein. Colcemid potentiates the reaction of vinblastine with tubulin and was used in this investigation to detect this protein.

Ultrastructure of the Parotid Gland of the Nine-Banded Armadillo

1977 ◽  
Vol 35 ◽  
pp. 640-641
Author(s):  
J. R. Ruby
Keyword(s):  
Endoplasmic Reticulum ◽  
Parotid Gland ◽  
Periodic Acid ◽  
Acinar Cells ◽  
Duct System ◽  
Parotid Glands ◽  
Dasypus Novemcinctus ◽  
Anterior Portion ◽  
Periodic Acid Schiff ◽  
Thin Sections

Parotid glands were obtained from five adult (four male and one female) armadillos (Dasypus novemcinctus) which were perfusion-fixed. The glands were located in a position similar to that of most mammals. They extended interiorly to the anterior portion of the submandibular gland.In the light microscope, it was noted that the acini were relatively small and stained strongly positive with the periodic acid-Schiff (PAS) and alcian blue techniques, confirming the earlier results of Shackleford (1). Based on these qualities and other structural criteria, these cells have been classified as seromucous (2). The duct system was well developed. There were numerous intercalated ducts and intralobular striated ducts. The striated duct cells contained large amounts of PAS-positive substance.Thin sections revealed that the acinar cells were pyramidal in shape and contained a basally placed, slightly flattened nucleus (Fig. 1). The rough endoplasmic reticulum was also at the base of the cell.

Organisation in the Structure of the Cytoplasmic Ground Substance

1978 ◽  
Vol 36 (3) ◽  
pp. 627-639
Author(s):  
K.R. Porter
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Complex ◽  
Random Distribution ◽  
Ground Substance ◽  
The Matrix ◽  
Cell Processes ◽  
Cytoplasmic Ground Substance

Most types of cells are known from their structure and overall form to possess a characteristic organization. In some instances this is evident in the non-random disposition of organelles and such system subunits as cisternae of the endoplasmic reticulum or the Golgi complex. In others it appears in the distribution and orientation of cytoplasmic fibrils. And in yet others the organization finds expression in the non-random distribution and orientation of microtubules, especially as found in highly anisometric cells and cell processes. The impression is unavoidable that in none of these cases is the organization achieved without the involvement of the cytoplasmic ground substance (CGS) or matrix. This impression is based on the fact that a matrix is present and that in all instances these formed structures, whether membranelimited or filamentous, are suspended in it. In some well-known instances, as in arrays of microtubules which make up axonemes and axostyles, the matrix resolves itself into bridges (and spokes) between the microtubules, bridges which are in some cases very regularly disposed and uniform in size (Mcintosh, 1973; Bloodgood and Miller, 1974; Warner and Satir, 1974).

Ultrastructural Changes Associated with Alcoholic Hyalin in Hepatocytes and Biliary Epithelial Cells

1971 ◽  
Vol 29 ◽  
pp. 572-573
Author(s):  
Odell T. Minick ◽  
Hidejiro Yokoo ◽  
Fawzia Batti
Keyword(s):  
Endoplasmic Reticulum ◽  
Epithelial Cells ◽  
Liver Biopsy ◽  
Alcoholic Hepatitis ◽  
Ultrastructural Changes ◽  
Main Mass ◽  
Biliary Epithelial Cells ◽  
Biopsy Specimens ◽  
Alcoholic Hyalin

To learn more of the nature and origin of alcoholic hyalin (AH), 15 liver biopsy specimens from patients with alcoholic hepatitis were studied in detail.AH was found not only in hepatocytes but also in ductular cells (Figs. 1 and 2), although in the latter location only rarely. The bulk of AH consisted of a randomly oriented network of closely packed filaments measuring about 150 Å in width. Bundles of filaments smaller in diameter (40-90 Å) were observed along the periphery of the main mass (Fig. 1), often surrounding it in a rim-like fashion. Fine filaments were also found close to the nucleus in both hepatocytes and biliary epithelial cells, the latter even though characteristic AH was not present (Figs. 3 and 4). Dispersed among the larger filaments were glycogen, RNA particles and profiles of endoplasmic reticulum. Dilated cisternae of endoplasmic reticulum were often conspicuous around the periphery of the AH mass. A limiting membrane was not observed.

Compound Symbiosis in Peridinium Balticum

1973 ◽  
Vol 31 ◽  
pp. 500-501
Author(s):  
R. N. Tomas
Keyword(s):  
Endoplasmic Reticulum ◽  
Nuclear Envelope ◽  
The Other ◽  
Exact Nature ◽  
Symbiotic Relationship

Peridinium balticum appears to be unusual among the dinoflagellates in that it possesses two DNA-containing structures as determined by histochemical techniques. Ultrastructurally, the two dissimilar nuclei are contained within different protoplasts; one of the nuclei is characteristically dinophycean in nature, while the other is characteristically eucaryotic. The chloroplasts observed within P. balticum are intrinsic to an eucaryotic photosynthetic endosymbiont and not to the dinoflagellate. These organelles are surrounded by outpocketings of endoplasmic reticulum which are continuous with the eucaryotic nuclear envelope and are characterized by thylakoids composed of three apposed lamellae. Girdle lamellae and membranebounded interlamellar pyrenoids are also present. Only the plasmalemma of the endosymbiont segregates its protoplast from that of the dinophycean cytoplasm. The exact nature of this symbiotic relationship is at present not known.

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