An Endoplasmic Reticulum Retrieval Signal Partitions Human Foamy Virus Maturation to Intracytoplasmic Membranes

ABSTRACT Among all retroviruses, foamy viruses (FVs) are unique in that they regularly mature at intracytoplasmic membranes. The envelope glycoprotein of FV encodes an endoplasmic reticulum (ER) retrieval signal, the dilysine motif (KKXX), that functions to localize the human FV (HFV) glycoprotein to the ER. This study analyzed the function of the dilysine motif in the context of infectious molecular clones of HFV that encoded mutations in the dilysine motif. Electron microscopy (EM) demonstrated virion budding both intracytoplasmically and at the plasma membrane for the wild-type and mutant viruses. Additionally, mutant viruses retained their infectivity, but viruses lacking the dilysine signal budded at the plasma membrane to a greater extent than did wild-type viruses. Interestingly, this relative increase in budding across the plasma membrane did not increase the overall release of viral particles into cell culture media as measured by protein levels in viral pellets or infectious virus titers. We conclude that the dilysine motif of HFV imposes a partial restriction on the site of viral maturation but is not necessary for viral infectivity.

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Ultrastructure and development of urediospore ornamentation in Melampsora lini

Canadian Journal of Botany ◽

10.1139/b71-291 ◽

1971 ◽

Vol 49 (12) ◽

pp. 2067-2073 ◽

Cited By ~ 26

Author(s):

L. J. Littlefield ◽

C. E. Bracker

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Outer Surface ◽

Spore Wall ◽

Wall Material ◽

Wild Type ◽

Melampsora Lini ◽

Inner Surface ◽

External Position ◽

Basal Portion

The urediospores of Melampsora lini (Ehrenb.) Lev. are echinulate, with spines ca. 1 μ long over their surface. The spines are electron-transparent, conical projections, with their basal portion embedded in the electron-dense spore wall. The entire spore, including the spines, is covered by a wrinkled pellicle ca. 150–200 Å thick. The spore wall consists of three recognizable layers in addition to the pellicle. Spines form initially as small deposits at the inner surface of the spore wall adjacent to the plasma membrane. Endoplasmic reticulum occurs close to the plasma membrane in localized areas near the base of spines. During development, the spore wall thickens, and the spines increase in size. Centripetal growth of the wall encases the spines in the wall material. The spines progressively assume a more external position in the spore wall and finally reside at the outer surface of the wall. A mutant strain with finely verrucose spores was compared to the wild type. The warts on the surface of the mutant spores are rounded, electron-dense structures ca. 0.2–0.4 μ high, in contrast to spines of the wild type. Their initiation near the inner surface of the spore wall and their eventual placement on the outer surface of the spore are similar to that of spines. The wall is thinner in mutant spores than in wild-type spores.

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Retention of adenovirus E19 glycoprotein in the endoplasmic reticulum is essential to its ability to block antigen presentation.

Journal of Experimental Medicine ◽

10.1084/jem.174.6.1629 ◽

1991 ◽

Vol 174 (6) ◽

pp. 1629-1637 ◽

Cited By ~ 69

Author(s):

J H Cox ◽

J R Bennink ◽

J W Yewdell

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Antigen Presentation ◽

Viral Proteins ◽

Genetically Engineered ◽

Class I ◽

Wild Type ◽

Histocompatibility Complex ◽

Infected Cells ◽

Lumenal Domain

The E3/19K glycoprotein of adenovirus functions to diminish recognition of adenovirus-infected cells by major histocompatibility complex class I-restricted cytotoxic T lymphocytes (CTLs) by binding intracellular class I molecules and preventing them from reaching the plasma membrane. In the present study we have characterized the nature of the interaction between E3/19K and the H-2Kd (Kd) molecule. An E3/19K molecule genetically engineered to terminate six residues from its normal COOH terminus (delta E19), was found to associate with Kd in a manner indistinguishable from wild-type E3/19K. Unlike E3/19K, however, delta E19 was transported through the Golgi complex to the plasma membrane, where it could be detected biochemically and immunocytochemically using a monoclonal antibody specific for the lumenal domain of E3/19K. Importantly, delta E19 also differed from E3/19K in being unable to prevent the presentation of Kd-restricted viral proteins to CTLs. This is unlikely to be due to delta E19 having a lower avidity for Kd than E3/19K, since delta E19 was able to compete with E3/19K for Kd binding, both physically, and functionally in nullifying the E3/19K blockade of antigen presentation. These findings indicate that the ability of E3/19K to block antigen presentation is due solely to its ability to retain newly synthesized class I molecules in the endoplasmic reticulum.

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Defective targeting of hemojuvelin to plasma membrane is a common pathogenetic mechanism in juvenile hemochromatosis

Blood ◽

10.1182/blood-2006-08-041004 ◽

2007 ◽

Vol 109 (10) ◽

pp. 4503-4510 ◽

Cited By ~ 59

Author(s):

Laura Silvestri ◽

Alessia Pagani ◽

Claudia Fazi ◽

Gianmario Gerardi ◽

Sonia Levi ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Iron Supplementation ◽

Proteolytic Processing ◽

Dual Function ◽

Wild Type ◽

Pathogenetic Mechanism ◽

Gpi Anchor ◽

Juvenile Hemochromatosis ◽

Deficient Mice

Abstract Hemojuvelin (HJV) positively modulates the iron regulator hepcidin, and its mutations are the major cause of juvenile hemochromatosis (JH), a recessive disease leading to iron overload. Defective HJV reduces hepcidin up-regulation both in humans and in Hjv-deficient mice. To investigate the JH pathogenesis and the functional properties of human HJV we studied the biosynthesis and maturation of 6 HJV pathogenic mutants in HeLa and HepG2 cells. We show that proteolytic processing is defective in mutants F170S, W191C, and G320V, but not in G99V and C119F. Moreover, we show that mutants G99V and C119F are targeted to the cell surface, while F170S, W191C, G320V, and R326X (lacking the glycosilphosphatidylinositol [GPI] anchor) are mainly retained in the endoplasmic reticulum, although all mutants are released as soluble forms (s-HJV) in a proportion that is modulated by iron supplementation. Membrane HJV (m-HJV) is mainly composed of the cleaved protein, and its level is increased by iron in wild-type (WT) mice but not in the mutants. Altogether, the data demonstrate that the loss of HJV membrane export is central to the pathogenesis of JH, and that HJV cleavage is essential for the export. The results support a dual function for s- and m-HJV in iron deficiency and overload, respectively.

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KISS1R Intracellular Trafficking and Degradation: Effect of the Arg386Pro Disease-Associated Mutation

Endocrinology ◽

10.1210/en.2010-0903 ◽

2011 ◽

Vol 152 (4) ◽

pp. 1616-1626 ◽

Cited By ~ 32

Author(s):

Suzy D. C. Bianco ◽

Lauren Vandepas ◽

Mayrin Correa-Medina ◽

Balázs Gereben ◽

Abir Mukherjee ◽

...

Keyword(s):

Plasma Membrane ◽

Intracellular Trafficking ◽

Proteasome Inhibitor ◽

Central Precocious Puberty ◽

Confocal Imaging ◽

Time Dependent ◽

Relative Distribution ◽

Wild Type ◽

Protein Levels ◽

Degradation Effect

Abstract The goal of this study was to investigate how the Arg386Pro mutation prolongs KiSS-1 receptor (KISS1R) responsiveness to kisspeptin, contributing to human central precocious puberty. Confocal imaging showed colocalization of wild-type (WT) KISS1R with a membrane marker, which persisted for up to 5 h of stimulation. Conversely, no colocalization with a lysosome marker was detected. Also, overnight treatment with a lysosome inhibitor did not affect WT KISS1R protein, whereas overnight treatment with a proteasome inhibitor increased protein levels by 24-fold. WT and Arg386Pro KISS1R showed time-dependent internalization upon stimulation. However, both receptors were recycled back to the membrane. The Arg386Pro mutation did not affect the relative distribution of KISS1R in membrane and internalized fractions when compared to WT KISS1R for up to 120 min of stimulation, demonstrating that this mutation does not affect KISS1R trafficking rate. Nonetheless, total Arg386Pro KISS1R was substantially increased compared with WT after 120 min of kisspeptin stimulation. This net increase was eliminated by blockade of detection of recycled receptors, demonstrating that recycled receptors account for the increased responsiveness of this mutant to kisspeptin. We therefore conclude the following: 1) WT KISS1R is degraded by proteasomes rather than lysosomes; 2) WT and Arg386Pro KISS1R are internalized upon stimulation, but most of the internalized receptors are recycled back to the membrane rather than degraded; 3) the Arg386Pro mutation does not affect the rate of KISS1R trafficking—instead, it prolongs responsiveness to kisspeptin by decreasing KISS1R degradation, resulting in the net increase on mutant receptor recycled back to the plasma membrane.

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A 3D analysis of yeast ER structure reveals how ER domains are organized by membrane curvature

The Journal of Cell Biology ◽

10.1083/jcb.201011039 ◽

2011 ◽

Vol 193 (2) ◽

pp. 333-346 ◽

Cited By ~ 214

Author(s):

Matt West ◽

Nesia Zurek ◽

Andreas Hoenger ◽

Gia K. Voeltz

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Electron Tomography ◽

Three Dimensional ◽

Membrane Curvature ◽

3D Analysis ◽

Wild Type ◽

Nanometer Resolution ◽

Ribosome Density ◽

Multiple Stages

We analyzed the structure of yeast endoplasmic reticulum (ER) during six sequential stages of budding by electron tomography to reveal a three-dimensional portrait of ER organization during inheritance at a nanometer resolution. We have determined the distribution, dimensions, and ribosome densities of structurally distinct but continuous ER domains during multiple stages of budding with and without the tubule-shaping proteins, reticulons (Rtns) and Yop1. In wild-type cells, the peripheral ER contains cytoplasmic cisternae, many tubules, and a large plasma membrane (PM)–associated ER domain that consists of both tubules and fenestrated cisternae. In the absence of Rtn/Yop1, all three domains lose membrane curvature, ER ribosome density changes, and the amount of PM-associated ER increases dramatically. Deletion of Rtns/Yop1 does not, however, prevent bloated ER tubules from being pulled from the mother cisterna into the bud and strongly suggests that Rtns/Yop1 stabilize/maintain rather than generate membrane curvature at all peripheral ER domains in yeast.

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Cytochrome P-450 3A13 and endothelin jointly mediate ductus arteriosus constriction to oxygen in mice

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00907.2010 ◽

2011 ◽

Vol 300 (3) ◽

pp. H892-H901 ◽

Cited By ~ 22

Author(s):

Barbara Baragatti ◽

Enrica Ciofini ◽

Francesca Scebba ◽

Debora Angeloni ◽

Daria Sodini ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Immunofluorescence Microscopy ◽

Oxygen Sensing ◽

Ductus Arteriosus ◽

Wild Type ◽

Phasic Response ◽

Sensing System ◽

Cytochrome P 450

The fetal ductus arteriosus (DA) contracts to oxygen, and this feature, maturing through gestation, is considered important for its closure at birth. We have previously obtained evidence of the involvement of cytochrome P-450, possibly of the 3A subfamily (CYP3A), in oxygen sensing and have also identified endothelin (ET)-1 as the attendant effector for the contraction. Here, we examined comparatively wild-type (WT) and CYP3A-null ( Cyp3a−/−) mice for direct validation of this concept. We found that the CYP3A subfamily is represented only by CYP3A13 in the WT DA. CYP3A13 was also detected in the DA by immunofluorescence microscopy, being primarily colocalized with the endoplasmic reticulum in both endothelial and muscle cells. However, a distinct signal was also evident in the plasma membrane. Isolated DAs from term WT animals developed a sustained contraction to oxygen with transient contractions superimposed. Conversely, no tonic response occurred in Cyp3a−/− DAs, whereas the phasic response persisted unabated. Oxygen did not contract the preterm WT DA but caused a full-fledged contraction after retinoic acid (RA) treatment. RA also promoted an oxygen contraction in the Cyp3a −/− DA. However, responses of RA-treated WT and Cyp3a−/− mice differed in that only the former abated with ET-1 suppression. This implies the existence of an alternative target for RA responsible for the oxygen-induced contraction in the absence of CYP3A13. In vivo, the DA was constricted in WT and Cyp3a−/− newborns, although with a tendency to be less narrowed in the mutant. We conclude that oxygen acts primarily through the complex CYP3A13 (sensor)/ET-1 (effector) and, in an accessory way, directly onto ET-1. However, even in the absence of CYP3A13, the DA may close postnatally thanks to the contribution of ET-1 and the likely involvement of compensating mechanism(s) identifiable with an alternative oxygen-sensing system and/or the withdrawal of relaxing influence(s) operating prenatally.

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The Fine Structure of Streptomyces coelicolor

The Journal of Cell Biology ◽

10.1083/jcb.7.3.479 ◽

1960 ◽

Vol 7 (3) ◽

pp. 479-487 ◽

Cited By ~ 62

Author(s):

Audrey M. Glauert ◽

David A. Hopwood

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Fine Structure ◽

Streptomyces Coelicolor ◽

Nuclear Material ◽

Tubular Structures ◽

Thin Sections ◽

Cytoplasmic Structure ◽

Intracytoplasmic Membranes ◽

Membranous Component

Colonies and spore suspensions of Streptomyces coelicolor were fixed by the method of Kellenberger, Ryter, and Séchaud (1958) and embedded in methacrylate or araldite. Thin sections were cut with an A. F. Huxley microtome and examined in a Siemens' Elmiskop I. At all stages of development the hyphae of Streptomyces coelicolor have an extensive membranous component in the cytoplasm. The membranes are continuous with the plasma membrane and have a variety of configurations at different places in the hyphae. Tubular structures, vesicles, and parallel stacks of membranes are seen. In some areas concentric layers of membranes form whorled structures which are particularly frequent in the region of developing cross-walls and within maturing spores. In the spores membranous structures often lie embedded in the nuclear material. In disintegrating hyphae the intracytoplasmic membranes round off into small vesicles and remain when the rest of the cytoplasmic structure has gone. In the absence of typical mitochondria and other cytoplasmic membranous structures it is possible that the membranous component of the cytoplasm of Streptomyces coelicolor may perform the functions of the endoplasmic reticulum and/or the mitochondria of higher cells.

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Remodeling of ER–plasma membrane contact sites but not STIM1 phosphorylation inhibits Ca2+influx in mitosis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1821399116 ◽

2019 ◽

Vol 116 (21) ◽

pp. 10392-10401 ◽

Cited By ~ 6

Author(s):

Fang Yu ◽

Satanay Z. Hubrack ◽

Sumita Chakraborty ◽

Lu Sun ◽

Ethel Alcantara-Adap ◽

...

Keyword(s):

Cell Cycle ◽

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Cell Cycle Progression ◽

Mechanistic Explanation ◽

Cycle Progression ◽

Protein Levels ◽

Membrane Contact Sites ◽

Contact Sites ◽

Membrane Contact

Store-operated Ca2+entry (SOCE), mediated by the endoplasmic reticulum (ER) Ca2+sensor stromal interaction molecule 1 (STIM1) and the plasma membrane (PM) channel Orai1, is inhibited during mitosis. STIM1 phosphorylation has been suggested to mediate this inhibition, but it is unclear whether additional pathways are involved. Here, we demonstrate using various approaches, including a nonphosphorylatable STIM1 knock-in mouse, that STIM1 phosphorylation is not required for SOCE inhibition in mitosis. Rather, multiple pathways converge to inhibit Ca2+influx in mitosis. STIM1 interacts with the cochaperone BAG3 and localizes to autophagosomes in mitosis, and STIM1 protein levels are reduced. The density of ER–PM contact sites (CSs) is also dramatically reduced in mitosis, thus physically preventing STIM1 and Orai1 from interacting to activate SOCE. Our findings provide insights into ER–PM CS remodeling during mitosis and a mechanistic explanation of the inhibition of Ca2+influx that is required for cell cycle progression.

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An NH2-terminal deleted plasma membrane H+-ATPase is a dominant negative mutant and is sequestered in endoplasmic reticulum derived structures

Biochemistry and Cell Biology ◽

10.1139/o99-071 ◽

2000 ◽

Vol 78 (1) ◽

pp. 51-58 ◽

Cited By ~ 8

Author(s):

Claudio Akio Masuda ◽

Mónica Montero-Lomelí

Keyword(s):

Endoplasmic Reticulum ◽

Plasma Membrane ◽

Immunofluorescence Microscopy ◽

Dominant Negative ◽

Dominant Negative Mutant ◽

Wild Type ◽

Internal Peptide ◽

Limited Trypsinolysis ◽

Wild Type Yeast ◽

Negative Mutant

The NH2-terminus of the plasma membrane H+-ATPase is one of the least conserved segments of this protein among fungi. We constructed and expressed a mutant H+-ATPase from Saccharomyces cerevisiae deleted at an internal peptide within the cytoplasmic NH2-terminus (D44-F116). When the enzyme was subjected to limited trypsinolysis it was digested more rapidly than wild type H+-ATPase. Membrane fractionation experiments and immunofluorescence microscopy, using antibodies against H+-ATPase showed that the mutant ATPase is retained in the endoplasmic reticulum. The pattern observed in the immunofluorescence microscopy resembled structures similar to Russell bodies (modifications of the endoplasmic reticulum membranes) recently described in yeast. When the wild type H+-ATPase was co-expressed with the mutant, wild type H+-ATPase was also retained in the endoplasmic reticulum. Co-expression of both ATPases in a wild type yeast strain was lethal, demonstrating that this is a dominant negative mutant.

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Nonapoptotic Death of Saccharomyces cerevisiae Cells That Is Stimulated by Hsp90 and Inhibited by Calcineurin and Cmk2 in Response to Endoplasmic Reticulum Stresses

Eukaryotic Cell ◽

10.1128/ec.00291-08 ◽

2008 ◽

Vol 7 (12) ◽

pp. 2037-2051 ◽

Cited By ~ 40

Author(s):

Drew D. Dudgeon ◽

Nannan Zhang ◽

Olufisayo O. Ositelu ◽

Hyemin Kim ◽

Kyle W. Cunningham

Keyword(s):

Saccharomyces Cerevisiae ◽

Endoplasmic Reticulum ◽

Mammalian Cells ◽

Culture Media ◽

Wild Type ◽

Dependent Protein Kinase ◽

Dependent Protein ◽

Synthetic Media ◽

Apoptosis And Necrosis ◽

Cells Death

ABSTRACT Endoplasmic reticulum (ER) stress can trigger apoptosis and necrosis in many types of mammalian cells. Previous studies in yeast found little or no cell death in response to the ER stressor tunicamycin, but a recent study suggested widespread apoptosis-like death. Here we show that wild-type laboratory Saccharomyces cerevisiae cells responding to tunicamycin die by nonapoptotic mechanisms in low-osmolyte culture media and survive for long periods of time in standard synthetic media. Survival requires calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, but none of its known targets. The Ca2+/calmodulin-dependent protein kinase Cmk2 was identified as an indirect target of calcineurin that suppresses death of calcineurin-deficient cells. Death of Cmk2- and/or calcineurin-deficient S. cerevisiae cells was preceded by accumulation of reactive oxygen species but was not associated with hallmarks of apoptosis and was not dependent on Mca1, Aif1, Nuc1, or other factors implicated in apoptosis-like death. Cmk2 and calcineurin also independently suppressed the death of S. cerevisiae cells responding to dithiothreitol or miconazole, a common azole-class antifungal drug. Though inhibitors of Hsp90 have been shown to diminish calcineurin signaling in S. cerevisiae and to synergistically inhibit growth in combination with azoles, they did not stimulate death of S. cerevisiae cells in combination with miconazole or tunicamycin, and instead they prevented the death of calcineurin- and Cmk2-deficient cells. These findings reveal a novel prodeath role for Hsp90 and antideath roles for calcineurin and Cmk2 that extend the life span of S. cerevisiae cells responding to both natural and clinical antifungal compounds.

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