A monoclonal antibody recognizes a human nuclear protein resembling Xenopus oocyte nucleoplasmin

1987 ◽  
Vol 87 (5) ◽  
pp. 713-722
Author(s):  
J. Lord ◽  
G. Brown ◽  
C. Bunce ◽  
P. Nathan
Keyword(s):  
Monoclonal Antibody ◽  
Xenopus Oocytes ◽  
Nuclear Protein ◽  
Chinese Hamster ◽  
Nucleosome Assembly ◽  
3T3 Cells ◽  
Nucleosome Assembly Protein ◽  
Subunit Molecular Weight ◽  
Interphase Cells ◽  
A Subunit

The monoclonal antibody 10BG2 (IgG3) was derived from a mouse immunized with human pre B cells. In immunofluorescence studies the antibody revealed a human nuclear-associated determinant, which in interphase cells was entirely restricted to the nucleus. In metaphase cells 10BG2 antigen was detected throughout the cytoplasm with intensified staining at the periphery of chromosomes. 10BG2 antibody stained all human normal and transformed cells tested. In contrast, the antibody did not stain mouse 3T3 cells or Chinese hamster ovary (CHO) cells. Electrophoresis under denaturing and non-denaturing conditions revealed the 10BG2 antigen to be a 130 X 10(3) to 140 X 10(3) Mr protein with a subunit molecular weight of 29.5 X 10(3). This suggests the protein is at least a tetramer. On two-dimensional gels the 10BG2 antigen had a streaked appearance and separated into two isoelectric variants (pI5.2, 5.4). The protein was also shown to be phosphorylated and thermostable, and remained in solution at pH 3.6. 10BG2 antigen was highly soluble in aqueous buffers and co-migrated on non-denaturing gels with the nucleosome-assembly protein, nucleoplasmin, purified from Xenopus oocytes. The similarity of 10BG2 antigen to nucleoplasmin is discussed.

The analysis of 40 kDa nuclear protein, p40, in interphase cells and mitotic cells

1993 ◽  
Vol 106 (3) ◽  
pp. 741-748 ◽  
Author(s):  
Y. Kaneda ◽  
K. Kinoshita ◽  
M. Sato ◽  
K. Tanaka ◽  
Y. Kaneda
Keyword(s):  
Monoclonal Antibody ◽  
Nuclear Envelope ◽  
Nuclear Protein ◽  
Dnase I ◽  
High Salt ◽  
The Other ◽  
Interphase Nuclei ◽  
Interphase Cells ◽  
Detergent Treatment ◽  
Sequential Treatments

We previously reported that the monoclonal antibody M108 recognized a 40 kDa protein both in the nucleus and the cytoplasm. This nuclear 40 kDa antigen was located in the nuclear envelope in interphase cells and in the perichromosomal region during mitosis. Now, we have analyzed this nuclear 40 kDa protein (p40) further, through morphological and biochemical approaches. At the beginning of mitosis, the perinuclear p40 detached from the nuclear envelope and moved to surround the condensing chromatin, while in the late stage of mitosis, the perichromosomal p40 moved back to the reassembled nuclear envelope. Most of the perichromosomal p40 on the metaphase chromosome was solubilized only by DNase I treatment, not by either high salt or detergent treatment. On the other hand, the perinuclear p40 was not solubilized by DNase1 alone, or high salt detergent alone. Sequential treatments with DNase I and high salt detergent were required to extract p40 in interphase nuclei. These results suggest that p40 was associated both with the nuclear envelope and chromatin DNA in interphase nuclei, while it bound only to chromatin DNA in mitosis.

Colocalization of a high molecular mass phosphoprotein of the nuclear matrix (p255) with spliceosomes

1995 ◽  
Vol 108 (5) ◽  
pp. 1873-1882 ◽  
Author(s):  
S. Bisotto ◽  
P. Lauriault ◽  
M. Duval ◽  
M. Vincent
Keyword(s):  
Monoclonal Antibody ◽  
Nuclear Matrix ◽  
Nuclear Protein ◽  
High Ionic Strength ◽  
High Molecular Mass ◽  
Splicing Factor ◽  
Nuclear Speckles ◽  
Ionic Detergents ◽  
Interphase Cells ◽  
Nuclear Structures

It was previously demonstrated that monoclonal antibody CC-3 binds to a phosphorylation dependent epitope present on a 255 kDa nuclear protein (p255). We show here that in interphase cells, p255 distributes to typical nuclear speckles that correspond to the localization of spliceosome components as revealed by antibodies to the m3G cap of snRNAs or to the non-snRNP splicing factor SC-35. Immunofluorescence and immunoblot studies indicated that p255 is resistant to extraction with non-ionic detergents, nucleases and high ionic strength buffers and may thus be defined biochemically as a nuclear matrix phosphoprotein. To determine the nature of the association of p255 with the nuclear structure, its distribution was studied at different stages of the cell cycle and after the cells were treated with nucleases or heat shocked. We found that the antigen diffused into the cytoplasm during metaphase but was reorganized into cytoplasmic speckles during anaphase-telophase transition, where it colocalized with SC-35. Nuclear matrix preparations that were digested with DNases and RNases showed that interphasic p255 still localized to nuclear speckles even though snRNA and snRNP antigens were removed. Heat-shocked cells labelled with monoclonal antibody CC-3 exhibited more rounded and less interconnected speckles, identical to those decorated by anti-SC-35 antibody under such conditions. These results indicate that p255 and SC-35 are present in the same nuclear structures, to which they are more tightly bound than the snRNP antigens. They further suggest that both proteins are implicated in spliceosome assembly or attachment.

scholarly journals Evaluation of Fab and F(ab′)2 Fragments and Isotype Variants of a Recombinant Human Monoclonal Antibody against Shiga Toxin 2

2010 ◽  
Vol 78 (3) ◽  
pp. 1376-1382 ◽  
Author(s):  
Donna E. Akiyoshi ◽  
Abhineet S. Sheoran ◽  
Curtis M. Rich ◽  
L. Richard ◽  
Susan Chapman-Bonofiglio ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Shiga Toxin ◽  
Chinese Hamster Ovary Cells ◽  
Human Monoclonal Antibody ◽  
Chinese Hamster ◽  
The Body ◽  
Neutralization Activity ◽  
Shiga Toxin 2

ABSTRACT 5C12 HuMAb is a human monoclonal antibody against the A subunit of Shiga toxin 2 (Stx2). We have previously shown that 5C12 HuMAb effectively neutralizes the cytotoxic effects of this toxin by redirecting its transport within the cell and also by neutralizing the toxin's ability to inhibit protein synthesis. The 5C12 HuMAb and its recombinant IgG1 version protect mice at a dose of 0.6 μg against a lethal challenge of Stx2. The contribution of the Fc region to this observed neutralization activity of the 5C12 antibody against Stx2 was investigated in this study. Using recombinant DNA technology, 5C12 isotype variants (IgG1, IgG2, IgG3, and IgG4) and antibody fragments [Fab, F(ab′)2] were expressed in Chinese hamster ovary cells and evaluated in vitro and in vivo. All four 5C12 isotype variants showed protection in vitro, with the IgG3 and IgG4 variants showing the highest protection in vivo. The Fab and F(ab′)2 fragments also showed protection in vitro but no protection in the mouse toxicity model. Similar results were obtained for a second HuMAb (5H8) against the B subunit of Stx2. The data suggest the importance of the Fc region for neutralization activity, but it is not clear if this is related to the stability of the full-length antibody or if the Fc region is required for effective elimination of the toxin from the body.

scholarly journals Tumor Suppressor NM23-H1 Is a Granzyme A-Activated DNase during CTL-Mediated Apoptosis, and the Nucleosome Assembly Protein SET Is Its Inhibitor

Cell ◽  
2003 ◽  
Vol 112 (5) ◽  
pp. 659-672 ◽  
Author(s):  
Zusen Fan ◽  
Paul J. Beresford ◽  
David Y. Oh ◽  
Dong Zhang ◽  
Judy Lieberman
Keyword(s):  
Tumor Suppressor ◽  
Nucleosome Assembly ◽  
Nucleosome Assembly Protein ◽  
Granzyme A

scholarly journals Crystal Structure of Malaria Parasite Nucleosome Assembly Protein

2009 ◽  
Vol 284 (15) ◽  
pp. 10076-10087 ◽  
Author(s):  
Jasmita Gill ◽  
Manickam Yogavel ◽  
Anuj Kumar ◽  
Hassan Belrhali ◽  
S. K. Jain ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Malaria Parasite ◽  
Nucleosome Assembly ◽  
Nucleosome Assembly Protein

scholarly journals A possible role for a mammalian facilitative hexose transporter in the development of resistance to drugs.

1991 ◽  
Vol 11 (7) ◽  
pp. 3407-3418 ◽  
Author(s):  
J C Vera ◽  
G R Castillo ◽  
O M Rosen
Keyword(s):  
Xenopus Oocytes ◽  
Glucose Transporter ◽  
Cytochalasin B ◽  
Chinese Hamster ◽  
Multidrug Resistant ◽  
Xenopus Laevis Oocytes ◽  
Hexose Transporter ◽  
P Glycoprotein ◽  
Glut1 Protein ◽  
Facilitative Glucose Transporter

We show that D- but not L-hexoses modulate the accumulation of radioactive vinblastine in injected Xenopus laevis oocytes expressing the murine Mdr1b P-glycoprotein. We also show that X. laevis oocytes injected with RNA encoding the rat erythroid/brain glucose transport protein (GLUT1) and expressing the corresponding functional transporter exhibit a lower accumulation of [3H]vinblastine and show a greater capacity to extrude the drug than do control oocytes not expressing the rat GLUT1 protein. Cytochalasin B and phloretin, two inhibitors of the mammalian facilitative glucose transporters, can overcome the reduced drug accumulation conferred by expression of the rat GLUT1 protein in Xenopus oocytes but have no significant effect on the accumulation of drug by Xenopus oocytes expressing the mouse Mdr1b P-glycoprotein. These drugs also increase the accumulation of [3H]vinblastine in multidrug-resistant Chinese hamster ovary cells. Cytochalasin E, an analog of cytochalasin B that does not affect the activity of the facilitative glucose transporter, has no effect on the accumulation of vinblastine by multidrug-resistant Chinese hamster cells or by oocytes expressing either the mouse Mdr1b P-glycoprotein or the GLUT1 protein. In all three cases, the drug verapamil produces a profound effect on the cellular accumulation of vinblastine. Interestingly, although immunological analysis indicated the presence of massive amounts of P-glycoprotein in the multidrug-resistant cells, immunological and functional studies revealed only a minor increase in the expression of a hexose transporter-like protein in resistant versus drug-sensitive cells. Taken together, these results suggest the participation of the mammalian facilitative glucose transporter in the development of drug resistance.

scholarly journals Isolation of monoclonal antibodies specific for products of avian oncogene myb.

1984 ◽  
Vol 4 (12) ◽  
pp. 2843-2850 ◽  
Author(s):  
G I Evan ◽  
G K Lewis ◽  
J M Bishop
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Nuclear Protein ◽  
Gene Products ◽  
Viral Oncogenes ◽  
Myb Gene

We isolated a series of monoclonal antibodies which were raised against a bacterially expressed protein, bp37v-myb, and coded for by part of the avian v-myb gene. These monoclonal antibodies recognized a range of antigenic specificities on bp37v-myb, and this was reflected in their differing specificities for the gene products of the v-myb, c-myb, and E26 viral oncogenes. One monoclonal antibody recognized, in addition to the v-myb and c-myb gene products, a conserved nuclear protein found in all tested cells. We describe the characterization of these monoclonal antibodies.

scholarly journals Zygotic nucleosome assembly protein–like 1 has a specific, non–cell autonomous role in hematopoiesis

Blood ◽  
2005 ◽  
Vol 106 (2) ◽  
pp. 514-520 ◽  
Author(s):  
Anita Abu-Daya ◽  
Wendy M. Steer ◽  
Alexandra F. Trollope ◽  
Christine E. Friedeberg ◽  
Roger K. Patient ◽  
...  
Keyword(s):  
Chromatin Remodeling ◽  
Neural Cells ◽  
Nucleosome Assembly ◽  
Globin Mrna ◽  
Blood Island ◽  
Nucleosome Assembly Protein ◽  
Hematopoietic Lineage ◽  
Core Histones ◽  
Hematopoietic Precursor ◽  
Assembly Proteins

Abstract Nucleosome assembly proteins (NAPs) bind core histones, facilitate chromatin remodeling, and can act as transcriptional coactivators. We previously described the isolation of a Xenopus NAP1-like (xNAP1L) cDNA, which encodes a member of this protein family. Its zygotic expression is restricted to neural cells, the outer cells of the ventral blood island (VBIs), and the ectoderm overlying the blood precursors. Here, we report that depletion of zygotic xNAP1L in embryos produces no obvious morphologic phenotype, but ablates α-globin mRNA expression in the VBIs. Transcript levels of the hematopoietic precursor genes SCL and Xaml (Runx-1) are also reduced in the VBIs. SCL expression can be rescued by injection of xNAP1L mRNA into the ectoderm, showing that the effect of xNAP1L can be non–cell autonomous. Fli1 and Hex, genes expressed in hemangioblasts but subsequently endothelial markers, were unaffected, suggesting that xNAP1L is required for the hematopoietic lineage specifically. Our data are consistent with a requirement for xNAP1L upstream of SCL, and injection of SCL mRNA into xNAP1L-depleted embryos rescues α-globin expression. Thus, xNAP1L, which belongs to a family of proteins previously believed to have general roles, has a specific function in hematopoiesis.

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