scholarly journals Long-Term Episomal Maintenance of Bovine Papillomavirus Type 1 Plasmids Is Determined by Attachment to Host Chromosomes, Which Is Mediated by the Viral E2 Protein and Its Binding Sites

1999 ◽  
Vol 73 (5) ◽  
pp. 4404-4412 ◽  
Author(s):  
Ivar Ilves ◽  
Sirje Kivi ◽  
Mart Ustav
Keyword(s):  
Host Cell ◽  
Binding Sites ◽  
Regulatory Region ◽  
Host Cells ◽  
Bovine Papillomavirus ◽  
E2 Protein ◽  
Multiple Binding Sites ◽  
Stable Maintenance ◽  
Necessary And Sufficient

ABSTRACT Papillomavirus genomes are stably maintained as extrachromosomal nuclear plasmids in dividing host cells. To address the mechanisms responsible for stable maintenance of virus, we examined nuclear compartmentalization of plasmids containing the full-length upstream regulatory region (URR) from the bovine papillomavirus type 1 (BPV1) genome. We found that these plasmids are tightly associated with the nuclear chromatin both in the stable cell lines that maintain episomal copies of the plasmids and in transiently transfected cells expressing the viral E1 and E2 proteins. Further analysis of viral factors revealed that the E2 protein in trans and its multiple binding sites in cis are both necessary and sufficient for the chromatin attachment of the plasmids. On the other hand, the BPV1 URR-dependent plasmid replication and chromatin attachment processes are clearly independent of each other. The ability of the plasmids to stably maintain episomes correlates clearly with their chromatin association function. These data suggest that viral E2 protein-mediated attachment of BPV1 genomes to the host cell chromatin could provide a mechanism for the coupling of viral genome multiplication and partitioning to the host cell cycle during viral latent infection.

scholarly journals Conditional Mutations in the Mitotic Chromosome Binding Function of the Bovine Papillomavirus Type 1 E2 Protein

2005 ◽  
Vol 79 (3) ◽  
pp. 1500-1509 ◽  
Author(s):  
Peng-Sheng Zheng ◽  
Jane Brokaw ◽  
Alison A. McBride
Keyword(s):  
Amino Acid ◽  
Bovine Papillomavirus ◽  
Transactivation Domain ◽  
Mitotic Chromosomes ◽  
E2 Protein ◽  
Binding Function ◽  
Acid Domain ◽  
Necessary And Sufficient ◽  
Amino Acid Domain

ABSTRACT The papillomavirus E2 protein is required for viral transcriptional regulation, DNA replication and genome segregation. We have previously shown that the E2 transactivator protein and BPV1 genomes are associated with mitotic chromosomes; E2 links the genomes to cellular chromosomes to ensure efficient segregation to daughter nuclei. The transactivation domain of the E2 protein is necessary and sufficient for association of the E2 protein with mitotic chromosomes. To determine which residues of this 200-amino-acid domain are important for chromosomal interaction, E2 proteins with amino acid substitutions in each conserved residue of the transactivation domain were tested for their ability to associate with mitotic chromosomes. Chromatin binding was assessed by using immunofluorescence on both spread and directly fixed mitotic chromosomes. E2 proteins defective in the transactivation and replication functions were unable to associate with chromosomes, and those that were competent in these functions were attached to mitotic chromosomes. However, several mutated proteins that were defective for chromosomal interaction could associate with chromosomes after treatment with agents that promote protein folding or when cells were incubated at lower temperatures. These results indicate that precise folding of the E2 transactivation domain is crucial for its interaction with mitotic chromosomes and that this association can be modulated.

Bix4 is activated directly by VegT and mediates endoderm formation in Xenopus development

Development ◽  
1999 ◽  
Vol 126 (19) ◽  
pp. 4193-4200 ◽  
Author(s):  
E.S. Casey ◽  
M. Tada ◽  
L. Fairclough ◽  
C.C. Wylie ◽  
J. Heasman ◽  
...  
Keyword(s):  
Binding Sites ◽  
Essential Role ◽  
Regulatory Region ◽  
Reporter Genes ◽  
Transcription Activator ◽  
T Box ◽  
Xenopus Embryos ◽  
Vegetal Pole ◽  
Endodermal Differentiation ◽  
Necessary And Sufficient

The maternal T-box gene VegT, whose transcripts are restricted to the vegetal hemisphere of the Xenopus embryo, plays an essential role in early development. Depletion of maternal VegT transcripts causes embryos to develop with no endoderm, while vegetal blastomeres lose the ability to induce mesoderm (Zhang, J., Houston, D. W., King, M. L., Payne, C., Wylie, C. and Heasman, J. (1998) Cell 94, 515–524). The targets of VegT, a transcription activator, must therefore include genes involved both in the specification of endoderm and in the production of mesoderm-inducing signals. We recently reported that the upstream regulatory region of the homeobox-containing gene Bix4 contains T-box binding sites. Here we show that expression of Bix4 requires maternal VegT and that two T-box binding sites are necessary and sufficient for mesodermal and endodermal expression of reporter genes driven by the Bix4 promoter in transgenic Xenopus embryos. Remarkably, a single T-box binding site is able to act as a mesoderm-specific enhancer when placed upstream of a minimal promoter. Finally, we show that Bix4 rescues the formation of endodermal markers in embryos in which VegT transcripts have been ablated but does not restore the ability of vegetal pole blastomeres to induce mesoderm. These results demonstrate that Bix4 acts directly downstream of VegT to specify endodermal differentiation in Xenopus embryos.

Analysis of the human CD10/neutral endopeptidase 24.11 promoter region: two separate regulatory elements

Blood ◽  
1995 ◽  
Vol 85 (11) ◽  
pp. 3199-3207 ◽  
Author(s):  
F Ishimaru ◽  
MA Shipp
Keyword(s):  
Binding Sites ◽  
Lymphoblastic Leukemia ◽  
Regulatory Region ◽  
Neutral Endopeptidase ◽  
Regulatory Elements ◽  
Control Element ◽  
Regulatory Regions ◽  
Endopeptidase 24.11

The cell surface zinc metalloproteinase CD10/neutral endopeptidase 24.11 (NEP) is expressed on normal and malignant lymphoid progenitors, granulocytes, and a variety of epithelial cells. To further define the tissue-specific and developmentally related expression of CD10/NEP, we have characterized two separate regulatory regions that control the transcription of 5′ alternatively spliced CD10/NEP transcripts. These type 1 and 2 CD10/NEP regulatory regions are both characterized by the presence of multiple transcription initiation sites and the absence of classic TATA boxes and consensus initiator elements. The purine-rich type 1 regulatory region, which includes 5′ UTR exon 1 sequence, is characterized by multiple putative PU.1 binding sites and consensus ets-binding motifs. In marked contrast, the GC-rich type 2 regulatory region contains multiple putative Sp1 binding sites, a potential consensus retinoblastoma control element (RCE), and an inverted CCAAT box. In the majority of tissues examined to date, type 2 CD10/NEP transcripts were more abundant; the abundance of type 1 transcripts was more variable, with the highest type 1 levels in fetal thymus and certain lymphoblastic leukemia cell lines.

scholarly journals Shigella applies molecular mimicry to subvert vinculin and invade host cells

2006 ◽  
Vol 175 (3) ◽  
pp. 465-475 ◽  
Author(s):  
Tina Izard ◽  
Guy Tran Van Nhieu ◽  
Philippe R.J. Bois
Keyword(s):  
Host Cell ◽  
Binding Sites ◽  
Type Iii Secretion ◽  
Molecular Mimicry ◽  
Shigella Flexneri ◽  
Cell Entry ◽  
Host Cells ◽  
General Mechanism ◽  
Host Cell Entry ◽  
Cell Spread

Shigella flexneri, the causative agent of bacillary dysentery, injects invasin proteins through a type III secretion apparatus upon contacting the host cell, which triggers pathogen internalization. The invasin IpaA is essential for S. flexneri pathogenesis and binds to the cytoskeletal protein vinculin to facilitate host cell entry. We report that IpaA harbors two vinculin-binding sites (VBSs) within its C-terminal domain that bind to and activate vinculin in a mutually exclusive fashion. Only the highest affinity C-terminal IpaA VBS is necessary for efficient entry and cell–cell spread of S. flexneri, whereas the lower affinity VBS appears to contribute to vinculin recruitment at entry foci of the pathogen. Finally, the crystal structures of vinculin in complex with the VBSs of IpaA reveal the mechanism by which IpaA subverts vinculin's functions, where S. flexneri utilizes a remarkable level of molecular mimicry of the talin–vinculin interaction to activate vinculin. Mimicry of vinculin's interactions may therefore be a general mechanism applied by pathogens to infect the host cell.

scholarly journals Identification of a Novel Nonstructural Protein, VP9, from White Spot Syndrome Virus: Its Structure Reveals a Ferredoxin Fold with Specific Metal Binding Sites

2006 ◽  
Vol 80 (21) ◽  
pp. 10419-10427 ◽  
Author(s):  
Yang Liu ◽  
Jinlu Wu ◽  
Jianxing Song ◽  
J. Sivaraman ◽  
Choy L. Hew
Keyword(s):  
Binding Sites ◽  
Metal Ion ◽  
White Spot Syndrome Virus ◽  
Nonstructural Protein ◽  
Divalent Metal ◽  
Full Length ◽  
Host Cells ◽  
White Spot ◽  
E2 Protein ◽  
White Spot Syndrome

ABSTRACT White spot syndrome virus (WSSV) is a major pathogen in shrimp aquaculture. VP9, a full-length protein of WSSV, encoded by open reading frame wsv230, was identified for the first time in the infected Penaeus monodon shrimp tissues, gill, and stomach as a novel, nonstructural protein by Western blotting, mass spectrometry, and immunoelectron microscopy. Real-time reverse transcription-PCR demonstrated that the transcription of VP9 started from the early to the late stage of WSSV infection as a major mRNA species. The structure of full-length VP9 was determined by both X-ray and nuclear magnetic resonance (NMR) techniques. It is the first structure to be reported for WSSV proteins. The crystal structure of VP9 revealed a ferredoxin fold with divalent metal ion binding sites. Cadmium sulfate was found to be essential for crystallization. The Cd2+ ions were bound between the monomer interfaces of the homodimer. Various divalent metal ions have been titrated against VP9, and their interactions were analyzed using NMR spectroscopy. The titration data indicated that VP9 binds with both Zn2+ and Cd2+. VP9 adopts a similar fold as the DNA binding domain of the papillomavirus E2 protein. Based on our present investigations, we hypothesize that VP9 might be involved in the transcriptional regulation of WSSV, a function similar to that of the E2 protein during papillomavirus infection of the host cells.

scholarly journals Ehrlichia chaffeensis TRP120 Binds a G+C-Rich Motif in Host Cell DNA and Exhibits Eukaryotic Transcriptional Activator Function

2011 ◽  
Vol 79 (11) ◽  
pp. 4370-4381 ◽  
Author(s):  
Bing Zhu ◽  
Jeeba A. Kuriakose ◽  
Tian Luo ◽  
Efren Ballesteros ◽  
Sharu Gupta ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Dna Binding ◽  
Host Cell ◽  
High Throughput ◽  
Tandem Repeat ◽  
Binding Sites ◽  
Host Gene ◽  
Host Cells ◽  
Ehrlichia Chaffeensis ◽  
Content Type

ABSTRACTEhrlichia chaffeensisis an obligately intracellular bacterium that modulates host cell gene transcription in the mononuclear phagocyte, but the host gene targets and mechanisms involved in transcriptional modulation are not well-defined. In this study, we identified a novel tandem repeat DNA-binding domain in theE. chaffeensis120-kDa tandem repeat protein (TRP120) that directly binds host cell DNA. TRP120 was observed by immunofluorescent microscopy in the nucleus ofE. chaffeensis-infected host cells and was detected in nuclear extracts by Western immunoblotting with TRP120-specific antibody. The TRP120 binding sites and associated host cell target genes were identified using high-throughput deep sequencing (Illumina) of immunoprecipitated DNA (chromatin immunoprecipitation and high-throughput DNA sequencing). Multiple em motif elicitation (MEME) analysis of the most highly enriched TRP120-bound sequences revealed a G+C-rich DNA motif, and recombinant TRP120 specifically bound synthetic oligonucleotides containing the motif. TRP120 target gene binding sites were mapped most frequently to intersecting regions (intron/exon; 49%) but were also identified in upstream regulatory regions (25%) and downstream locations (26%). Genes targeted by TRP120 were most frequently associated with transcriptional regulation, signal transduction, and apoptosis. TRP120 targeted inflammatory chemokine genes, CCL2, CCL20, and CXCL11, which were strongly upregulated duringE. chaffeensisinfection and were also upregulated by direct transfection with recombinant TRP120. This study reveals that TRP120 is a novel DNA-binding protein that is involved in a host gene transcriptional regulation strategy.

Chlorate: a reversible inhibitor of proteoglycan sulphation in Chlamydia trachomatis-infected cells

2004 ◽  
Vol 53 (2) ◽  
pp. 93-95 ◽  
Author(s):  
Sanaa Fadel ◽  
Adrian Eley
Keyword(s):  
Chlamydia Trachomatis ◽  
Host Cell ◽  
Chlamydial Infection ◽  
Heparan Sulphate ◽  
Sodium Chlorate ◽  
Inhibitory Effect ◽  
Host Cells ◽  
Reversible Inhibitor ◽  
Sulphated Glycosaminoglycans ◽  
Necessary And Sufficient

Sulphated glycosaminoglycans, such as heparan sulphate, have been shown to be essential for the infectivity of many organisms. The aims of this study were to verify the role of sulphated glycosaminoglycans in chlamydial infection and to investigate whether they are present on chlamydia or chlamydial host cells. The effect of undersulphation of host cells and chlamydial elementary bodies was examined using sodium chlorate. Also studied was whether any inhibitory effect was reversible. The results strongly suggest that Chlamydia trachomatis does not produce heparan sulphate and that heparan sulphate of the host cell is necessary and sufficient to mediate chlamydial infection. The essential role played by the sulphate constituents of the host-cell glycosaminoglycan in the infectivity of LGV serovars, and to a lesser extent of serovar E, was also confirmed.

scholarly journals Analysis of Chromatin Attachment and Partitioning Functions of Bovine Papillomavirus Type 1 E2 Protein

2004 ◽  
Vol 78 (4) ◽  
pp. 2100-2113 ◽  
Author(s):  
Aare Abroi ◽  
Ivar Ilves ◽  
Sirje Kivi ◽  
Mart Ustav
Keyword(s):  
Mutational Analysis ◽  
Point Mutations ◽  
Human Herpesvirus ◽  
Replication Initiation ◽  
Bovine Papillomavirus ◽  
Composite Surface ◽  
E2 Protein ◽  
Terminal Domain ◽  
Specific Receptors

ABSTRACT Recent studies have suggested that the tethering of viral genomes to host cell chromosomes could provide one of the ways to achieve their nuclear retention and partitioning during extrachromosomal maintenance in dividing cells. The data we present here provide firm evidence that the partitioning of the bovine papillomavirus type 1 (BPV1) genome is dependent on the chromatin attachment process mediated by viral E2 protein and its multiple binding sites. On the other hand, the attachment of E2 and the E2-mediated tethering of reporter plasmids to host chromosomes are not necessarily sufficient for efficient partitioning, suggesting that additional E2-dependent activities might be involved in the latter process. The activity of E2 protein in chromatin attachment and partitioning is more sensitive to the point mutations in the N-terminal domain than its transactivation and replication initiation functions. Therefore, at least part of the interactions of the E2 N-terminal domain with its targets during the chromatin attachment and partitioning processes are likely to involve specific receptors not involved in transactivation and replication activities of the protein. The mutational analysis also indicates that the binding of E2 to chromatin is not achieved through interaction of linear N-terminal subsequences of the E2 protein with putative receptors. Instead, the composite surface elements of the N-terminal domain build up the receptor-binding surface of E2. In this regard, the interaction of BPV1 E2 with its chromosomal targets clearly differs from the interactions of LANA1 protein from Kaposi's sarcoma-associated human herpesvirus and EBNA1 from Epstein-Barr virus with their specific receptors.

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