Punctate Pattern & T2/ FLAIR Mismatch as findings in PML Related to Rituximab: Beyond Natalizumab-PML

Myr 1 is a widely distributed mammalian myosin I molecule related to brush border myosin 1. A second widely distributed myosin I molecule similar to myr 1 and brush border myosin I, called myr 2, has now been identified. Specific antibodies and expression of epitope-tagged molecules were used to determine the subcellular localization of myr 1 and myr 2 in NRK cells. Myr 1 was detected at the plasma membrane and was particularly enriched in cell protrusions like lamellipodia, membrane ruffles and filopodia. In dividing cells myr 1 localized to the cleavage furrow. Myr 2 was localized in a discrete punctate pattern in resting cells and in cells undergoing cytokinesis. In subcellular fractionation experiments myr 1 and myr 2 were both partly soluble and partly associated with smooth membranes of medium density. The tail domains of myosin I molecules have been proposed to interact with a receptor and thereby determine the subcellular localization. To test this hypothesis we expressed the tail domains of myr 1 and myr 2 that lack the F-actin-binding myosin head domain in NRK cells. These tail domains also partly copurified with smooth membranes of medium density and immunolocalized similar to the respective endogenous myosin I; however, they exhibited a lower affinity for membranes and an increased diffuse cytosolic localization. These results suggest that the tail domains of myr 1 and myr 2 are sufficient for subcellular targeting but that their head domains also contribute significantly to maintaining a proper subcellular localization.

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Reconstituted nuclei depleted of a vertebrate GLFG nuclear pore protein, p97, import but are defective in nuclear growth and replication.

The Journal of Cell Biology ◽

10.1083/jcb.128.5.721 ◽

1995 ◽

Vol 128 (5) ◽

pp. 721-736 ◽

Cited By ~ 97

Author(s):

M A Powers ◽

C Macaulay ◽

F R Masiarz ◽

D J Forbes

Keyword(s):

Protein Import ◽

Polyclonal Antiserum ◽

Peptide Sequence ◽

Nuclear Pore ◽

Chromosomal Dna ◽

Nuclear Pore Protein ◽

Egg Extracts ◽

Xenopus Egg ◽

Punctate Pattern

Xenopus egg extracts provide a powerful system for in vitro reconstitution of nuclei and analysis of nuclear transport. Such cell-free extracts contain three major N-acetylglucosaminylated proteins: p200, p97, and p60. Both p200 and p60 have been found to be components of the nuclear pore. Here, the role of p97 has been investigated. Xenopus p97 was isolated and antisera were raised and affinity purified. Immunolocalization experiments indicate that p97 is present in a punctate pattern on the nuclear envelope and also in the nuclear interior. Peptide sequence analysis reveals that p97 contains a GLFG motif which defines a family of yeast nuclear pore proteins, as well as a peptide that is identical at 11/15 amino acids to a specific member of the GLFG family, NUP116. An additional peptide is highly homologous to a second sequence found in NUP116 and other members of the yeast GLFG family. A monoclonal antibody to the GLFG domain cross-reacts with a major Xenopus protein of 97 kD and polyclonal antiserum to p97 recognizes the yeast GLFG nucleoporin family. The p97 antiserum was used to immunodeplete Xenopus egg cytosol and p97-deficient nuclei were reconstituted. The p97-depleted nuclei remained largely competent for nuclear protein import. However, in contrast to control nuclei, nuclei deficient in p97 fail to grow in size over time and do not replicate their chromosomal DNA. ssDNA replication in such extracts remains unaffected. Addition of the N-acetylglucosaminylated nuclear proteins of Xenopus or rat reverses these replication and growth defects. The possible role(s) of p97 in these nuclear functions is discussed.

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High Expression of Cathepsin D in Non-Hodgkin’s Lymphomas Negatively Impacts on Clinical Outcome

Disease Markers ◽

10.1155/2010/465040 ◽

2010 ◽

Vol 28 (3) ◽

pp. 167-183 ◽

Cited By ~ 4

Author(s):

Giuseppina Nicotra ◽

Federica Manfroi ◽

Carlo Follo ◽

Roberta Castino ◽

Nicola Fusco ◽

...

Keyword(s):

Clinical Outcome ◽

Tumor Cells ◽

Survival Probability ◽

Cathepsin D ◽

Rank Test ◽

Lysosomal Protease ◽

T Cell Lymphomas ◽

Kaplan Meier ◽

Punctate Pattern ◽

Hodgkin's Lymphomas

The lysosomal protease Cathepsin D (CD) has been implicated in the homeostasis of lymphatic tissues. We investigated whether the level of CD expression influences the progression and the clinical outcome in Non-Hodgkin’s Lymphomas (NHLs). The expression of CD was assessed by immunohistochemistry and immunofluorescence in biopsies of Diffuse Large B Cell Lymphomas (DLBCL, 35 cases), Follicular Lymphomas (FL, 9 cases of grade I-II plus 14 cases of grade IIIB), Chronic Lymphocytic Leukaemias (CLL, 17 cases) and Peripheral T-cell Lymphomas (PTCL, 5 cases). CD staining showed a cytoplasmic punctate pattern compatible with its lysosomal localization. Based on the level of CD expression and the proportion of positive cells, lymphomas were classified as ‘low expressing’ (< 20% of tumor cells) or ‘highly expressing’ (≥ 20% of tumor cells). Lymphomas highly expressing CD were associated with a worse stage (III-IV) at diagnosis (31/34 cases;p= 0.002) and with a poor clinical outcome (i.e., partial remission and death; 28/34 cases;p= 0.03). In the subgroup of aggressive/high grade of malignancy lymphomas (i.e., DLBCL, FL IIIB and PTCL), the Kaplan-Meier curve revealed a very low cumulative overall survival probability (~20% at 5 year) for patients bearing a NHL with > 40% CD-positive cells compared to that of patients bearing a NHL with < 20% CD-positive cells (~70% at 5 year). This correlation was statistically significant (log-rank test,p= 0.01). In Cox multivariate analysis CD failed to be a prognosticator independent of pathologic stage, though the hazard ratio confirmed the association of low expression with a better survival probability. These data indicate that the presence of a high percentage of CD-positive tumor cells negatively reflects on the progression of NHLs.

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Monoclonal antibodies identify a group of nuclear pore complex glycoproteins.

The Journal of Cell Biology ◽

10.1083/jcb.104.5.1143 ◽

1987 ◽

Vol 104 (5) ◽

pp. 1143-1156 ◽

Cited By ~ 314

Author(s):

C M Snow ◽

A Senior ◽

L Gerace

Keyword(s):

Monoclonal Antibodies ◽

Nuclear Envelope ◽

Nuclear Pore Complex ◽

Peptide Mapping ◽

Polyclonal Antibodies ◽

Integral Membrane Protein ◽

Nucleocytoplasmic Transport ◽

Nuclear Pore ◽

Nuclear Surface ◽

Punctate Pattern

Using monoclonal antibodies we identified a group of eight polypeptides of rat liver nuclear envelopes that have common epitopes. Most or all of these proteins are structurally distinct, as shown by tryptic peptide mapping and analysis with polyclonal antibodies. While these polypeptides are relatively tightly bound to nuclear membranes, only one is an integral membrane protein. The eight antigens cofractionate with the nuclear pore complex under various conditions of ionic strength and detergent. It can be seen by immunofluorescence microscopy that the monoclonal antibodies reacting with these antigens stain the nuclear surface of interphase cells in a finely punctate pattern. When the nuclear envelope is disassembled and subsequently reformed during mitosis, the proteins are reversibly dispersed throughout the cytoplasm in the form of minute foci. By EM immunogold localization on isolated nuclear envelopes, the monoclonal antibodies label exclusively the nuclear pore complex, at both its nucleoplasmic and cytoplasmic margins. Considered together, our biochemical and localization data indicate that the eight nuclear envelope polypeptides are pore complex components. As shown in the accompanying paper (Holt, G. D., C. M. Snow, A. Senior, R. S. Haltiwanger, L. Gerace, and G. W. Hart, J. Cell Biol., 104:1157-1164) these eight polypeptides contain a novel form of glycosylation, O-linked N-acetylglucosamine. The relative abundance and disposition of these O-linked glycoproteins in the pore complex are consistent with their having a role in nucleocytoplasmic transport.

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Punctate Pattern and Pemphigus

American Journal of Dermatopathology ◽

10.1097/dad.0000000000002047 ◽

2021 ◽

Vol Publish Ahead of Print ◽

Author(s):

Nafiseh Esmaili ◽

Kambiz Kamyab ◽

Parvaneh Hatami ◽

Shirin Behrouzifar ◽

Maryam Daneshpazhooh ◽

...

Keyword(s):

Punctate Pattern

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Application of a novel cell-permeable peptide-driven protein delivery in mouse blastocysts

Reproduction ◽

10.1530/rep-13-0203 ◽

2013 ◽

Vol 146 (2) ◽

pp. 145-153 ◽

Cited By ~ 4

Author(s):

Sojung Kwon ◽

Areum Kwak ◽

Hyejin Shin ◽

Soyoung Choi ◽

Soohyun Kim ◽

...

Keyword(s):

Fusion Protein ◽

Hela Cells ◽

Protein Delivery ◽

Cancer Cell Line ◽

Preimplantation Embryos ◽

Cervical Cancer Cell Line ◽

Cell Stage ◽

Cell Permeable Peptide ◽

Morula Stage ◽

Punctate Pattern

Cell-permeable peptides (CPPs) mediate the delivery of macromolecules into cells. However, whether CPPs are usable in mammalian oocytes and embryos for the modulation of protein expression has not been widely investigated. We have previously designed a novel 12-mer CPP from the conserved region of the human papillomavirus L1 capsid protein. In this study, we tested whether this peptide, LDP12, effectively delivers a protein cargo to mouse oocytes and preimplantation embryos. We prepared a LDP12–EGFP fusion protein having LDP12 as an N-terminal tag. This fusion protein readily enters HeLa cells, a cervical cancer cell line. The entry of LDP12–EGFP was partially blocked by amiloride, while cytochalasin D or methyl-β-cyclodextrin slightly increased the uptake. LDP12–EGFP shows efficient transduction in mouse blastocysts, but not in oocytes, two-cell-stage, or morula-stage-preimplantation embryos. LDP12-mediated delivery of EGFP–LC3, a widely used marker of autophagic activation, is successful in HeLa cells and mouse blastocysts, as it enters cells and exhibits a signature punctate pattern. The lipidation of EGFP–LC3 also normally occurs after transduction, suggesting that the transduced protein retains the functional characteristics. Collectively, we show that LDP12-driven protein delivery is a fast and convenient method applicable to mouse blastocysts and reproductive cancer cells.

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Effects of cytochalasin B on actin and myosin association with particle binding sites in mouse macrophages: implications with regard to the mechanism of action of the cytochalasins.

The Journal of Cell Biology ◽

10.1083/jcb.91.2.373 ◽

1981 ◽

Vol 91 (2) ◽

pp. 373-384 ◽

Cited By ~ 39

Author(s):

R G Painter ◽

J Whisenand ◽

A T McIntosh

Keyword(s):

Binding Sites ◽

Cytochalasin B ◽

Peritoneal Macrophages ◽

Immunofluorescent Staining ◽

Transmission Electron ◽

Mouse Macrophages ◽

Particle Binding ◽

Punctate Pattern ◽

Particle Ingestion ◽

Mouse Peritoneal Macrophages

The intracellular distribution of F-actin and myosin has been examined in mouse peritoneal macrophages by immunofluorescence microscopy. In resting, adherent cells, F-actin was distributed in a fine networklike pattern throughout the cytoplasm. Myosin, in contrast, was distributed in a punctate pattern. After treatment with cytochalasin B (CB), both proteins showed a coarse punctate pattern consistent with a condensation of protein around specific foci. After CB-pretreated cells were exposed to opsonized zymosan particles, immunofluorescent staining for F-actin and myosin showed an increased staining under particle binding sites. Transmission electron microscope (TEM) examination of whole-cell mounts of such preparations revealed a dense zone of filaments beneath the relatively electron-translucent zymosan particles. At sites where particles had detached during processing, these filament-rich areas were more clearly delineated. At such sites dense arrays of filaments that appeared more or less randomly oriented were apparent. The filaments could be decorated with heavy meromyosin, suggesting that they were composed, in part, of F-actin and were therefore identical to the structures giving rise to the immunofluorescence patterns. After viewing CB-treated preparations by whole-mount TEM, we examined the cells by scanning electron microscopy (SEM). Direct SEM comparison of the filament-rich zones seen by TEM showed that these structures resulted from the formation of short lamellipodial protrusions below the site of particle binding. Electron micrographs of thin-sectioned material established that these lamellipodial protrusions were densely packed with microfilaments that were in part associated with the cytoplasmic surface of the plasma membrane. The formation of particle-associated lamellipodia did not appear to represent merely a slower rate of ingestion in the presence of CB, because they formed within minutes of particle contact with the cell membrane and were not followed by particle ingestion even after a 1-h or longer incubation. Furthermore, their formation required cellular energy. These results suggest that cytochalasin B blocks phagocytosis of large particles by affecting the distances over which any putative actomyosin-mediated forces are generated.

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The odd one out: Arabidopsis reticulon20 has a role in lipid biosynthesis

10.1101/123679 ◽

2017 ◽

Author(s):

Verena Kriechbaumer ◽

Lilly Maneta-Peyret ◽

Stanley W Botchway ◽

Jessica Upson ◽

Louise Hughes ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Membrane Proteins ◽

Sterol Composition ◽

Lipid Biosynthesis ◽

Transmembrane Domains ◽

Eukaryotic Cells ◽

The Family ◽

Er Exit Sites ◽

Punctate Pattern ◽

Er Membrane

AbstractThe family of reticulon proteins has been shown to be involved in a variety of functions in eukaryotic cells including tubulation of the endoplasmic reticulum (ER), formation of cell plates and primary plasmodesmata. Reticulons are integral ER membrane proteins characterised by a reticulon homology domain comprising four transmembrane domains which results in the reticulons sitting in the membrane in a W-topology. Here we report on a subgroup of reticulons with an extended N-terminal domain and in particular on arabidopsis reticulon 20. We show that reticulon 20 is located in a unique punctate pattern on the ER membrane. Its closest homologue reticulon 19 labels the whole ER. We show that mutants in RTN20 or RTN19, respectively, display a significant change in sterol composition in the roots indicating a role in lipid biosynthesis or regulation. A third homologue in this family - 3BETAHSD/D1- is localised to ER exit sites resulting in an intriguing location difference for the three proteins.

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Polar asymmetry in the organization of the cortical cytokeratin system of Xenopus laevis oocytes and embryos

Development ◽

10.1242/dev.100.3.543 ◽

1987 ◽

Vol 100 (3) ◽

pp. 543-557 ◽

Cited By ~ 1

Author(s):

M.W. Klymkowsky ◽

L.A. Maynell ◽

A.G. Polson

Keyword(s):

Xenopus Laevis ◽

Mature Stage ◽

Xenopus Laevis Oocytes ◽

Blastula Stage ◽

Early Embryos ◽

Large Fibre ◽

Filament Bundles ◽

Punctate Pattern ◽

Cortical Cytoskeleton ◽

Cytokeratin Filaments

We have used whole-mount immunofluorescence microscopy of late-stage Xenopus laevis oocytes and early embryos to examine the organization of their cortical cytokeratin systems. In both mature oocytes and early embryos, there is a distinct animal-vegetal polarity in cytokeratin organization. In mature (stage-VI) oocytes, the cytokeratin filaments of the vegetal region form a unique, almost geodesic network; in the animal region, cytokeratin organization appears much more variable and irregular. In unfertilized, postgerminal vesicle breakdown eggs, the cortical cytokeratin system is disorganized throughout both animal and vegetal hemispheres. After fertilization, cytokeratin organization reappears first in a punctate pattern that is transformed into an array of oriented filaments. These cytokeratin filaments appear first in the vegetal hemisphere and are initially thin. Subsequently, they form bundles that grow thicker through the period of first to second cleavage, at which point large cytokeratin filament bundles form a loose, fishnet-like system that encompasses the vegetal portion of each blastomere. In the animal region, cytokeratin filaments do not appear to form large fibre networks, but rather appear to be organized into a system of fine filaments. The animal-vegetal polarity in cytokeratin organization persists until early blastula (stage 5); in later-stage embryos, both animal and vegetal blastomeres possess qualitatively similar cytokeratin filament systems. The entire process of cytokeratin reorganization in the egg is initiated by prick activation. These observations indicate that the cortical cytoskeleton of Xenopus oocytes and early embryos is both dynamic and asymmetric.

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A cytokeratin 8-like protein with plasminogen-binding activity is present on the external surfaces of hepatocytes, HepG2 cells and breast carcinoma cell lines

Journal of Cell Science ◽

10.1242/jcs.108.3.1071 ◽

1995 ◽

Vol 108 (3) ◽

pp. 1071-1082

Author(s):

T.A. Hembrough ◽

J. Vasudevan ◽

M.M. Allietta ◽

W.F. Glass ◽

S.L. Gonias

Keyword(s):

Breast Carcinoma ◽

Cell Surface ◽

Immunoelectron Microscopy ◽

Antigen Expression ◽

Rat Hepatocytes ◽

Binding Activity ◽

Carcinoma Cells ◽

Cellular Migration ◽

Punctate Pattern ◽

Breast Carcinoma Cell Lines

Plasminogen binding to cell surfaces may be important for tumor invasion and other processes that involve cellular migration. In this investigation, the principal plasminogen-binding protein was identified in the plasma membrane fraction of rat hepatocytes. The protein had an apparent mass of 59 kDa, was insoluble in a spectrum of detergents, and was identical to cytokeratin 8 (CK 8) as determined by sequence analysis of nine amino acids at the N terminus of two cyanogen bromide fragments. The 59 kDa protein bound CK 8-specific antibody in western blot analyses. These studies demonstrate that CK 8 or a CK 8-like protein binds plasminogen. Given this newly determined and potentially important CK 8 function, immunofluorescence and immunoelectron microscopy studies were performed to determine whether CK 8 may be present on the external surfaces of unpermeabilized, viable hepatocytes. All of the cells in each preparation were immunopositive with two separate CK 8-specific antibodies. A punctate pattern of immunofluorescence was detected on the cell surface with approximately even intensity from cell to cell. By immunoelectron microscopy, CK 8 was preferentially associated with microvilli. In order to determine whether other epithelial cells express cell-surface CK 8, immunofluorescence and immunoelectron microscopy studies were performed with HepG2 hepatocellular carcinoma cells and with BT20 and MCF-7 breast carcinoma cells. The pattern of antigen expression was equivalent with each cell type and comparable to that observed with hepatocytes. These studies support the hypothesis that CK 8 is associated with the external cell surface where it may express important proteinase receptor function.

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