scholarly journals Interaction of the Betapapillomavirus E2 Tethering Protein with Mitotic Chromosomes

2009 ◽  
Vol 84 (1) ◽  
pp. 543-557 ◽  
Author(s):  
Vandana Sekhar ◽  
Shawna C. Reed ◽  
Alison A. McBride
Keyword(s):  
Amino Acid ◽  
Viral Genome ◽  
Sequence Similarity ◽  
Bovine Papillomavirus ◽  
Mitotic Chromosomes ◽  
E2 Protein ◽  
Amino Acid Peptide ◽  
Amino Acid Region ◽  
Papillomavirus Infection ◽  
Chromosome Binding

ABSTRACT During persistent papillomavirus infection, the viral E2 protein tethers the viral genome to the host cell chromosomes, ensuring maintenance and segregation of the viral genome during cell division. However, E2 proteins from different papillomaviruses interact with distinct chromosomal regions and targets. The tethering mechanism has been best characterized for bovine papillomavirus type 1 (BPV1), where the E2 protein tethers the viral genome to mitotic chromosomes in complex with the cellular bromodomain protein, Brd4. In contrast, the betapapillomavirus human papillomavirus type 8 (HPV8) E2 protein binds to the repeated ribosomal DNA genes that are found on the short arm of human acrocentric chromosomes. In this study, we show that a short 16-amino-acid peptide from the hinge region and the C-terminal DNA binding domain of HPV8 E2 are necessary and sufficient for interaction with mitotic chromosomes. This 16-amino-acid region contains an RXXS motif that is highly conserved among betapapillomaviruses, and both arginine 250 and serine 253 residues within this motif are required for mitotic chromosome binding. The HPV8 E2 proteins are highly phosphorylated, and serine 253 is a site of phosphorylation. The HPV8 E2 chromosome binding sequence also has sequence similarity with chromosome binding regions in the gammaherpesvirus EBNA and LANA tethering proteins.

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.

scholarly journals Dynamic Localization of the Human Papillomavirus Type 11 Origin Binding Protein E2 through Mitosis While in Association with the Spindle Apparatus

2006 ◽  
Vol 80 (10) ◽  
pp. 4792-4800 ◽  
Author(s):  
Luan D. Dao ◽  
Aaron Duffy ◽  
Brian A. Van Tine ◽  
Shwu-Yuan Wu ◽  
Cheng-Ming Chiang ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
Binding Protein ◽  
Cellular Protein ◽  
Bovine Papillomavirus ◽  
Mitotic Spindles ◽  
Mitotic Chromosomes ◽  
Mitotic Apparatus ◽  
Viral Origin ◽  
E2 Protein ◽  
Origin Binding Protein

ABSTRACT Papillomaviral DNA replicates as extrachromosomal plasmids in squamous epithelium. Viral DNA must segregate equitably into daughter cells to persist in dividing basal/parabasal cells. We have previously reported that the viral origin binding protein E2 of human papillomavirus types 11 (HPV-11), 16, and 18 colocalized with the mitotic spindles. In this study, we show the localization of the HPV-11 E2 protein to be dynamic. It colocalized with the mitotic spindles during prophase and metaphase. At anaphase, it began to migrate to the central spindle microtubules, where it remained through telophase and cytokinesis. It was additionally observed in the midbody at cytokinesis. A peptide spanning residues 285 to 308 in the carboxyl-terminal domain of HPV-11 E2 (E2C) is necessary and sufficient to confer localization on the mitotic spindles. This region is conserved in HPV-11, -16, and -18 and bovine papillomavirus type 4 (BPV-4) E2 and is also required for the respective E2C to colocalize with the mitotic spindles. The E2 protein of bovine papillomavirus type 1 is tethered to the mitotic chromosomes via the cellular protein Brd4. However, the HPV-11 E2 protein did not associate with Brd4 during mitosis. Lastly, a chimeric BPV-1 E2C containing the spindle localization domain from HPV-11 E2C gained the ability to localize to the mitotic spindles, whereas the reciprocal chimera lost the ability. We conclude that this region of HPV E2C is critical for localization with the mitotic apparatus, enabling the HPV DNA to sustain persistent infections.

The TIS11 primary response gene is a member of a gene family that encodes proteins with a highly conserved sequence containing an unusual Cys-His repeat

1991 ◽  
Vol 11 (3) ◽  
pp. 1754-1758
Author(s):  
B C Varnum ◽  
Q F Ma ◽  
T H Chi ◽  
B Fletcher ◽  
H R Herschman
Keyword(s):  
Amino Acid ◽  
Gene Family ◽  
Sequence Similarity ◽  
Primary Response ◽  
Response Gene ◽  
Pheochromocytoma Cells ◽  
Conserved Sequence ◽  
Amino Acid Region ◽  
Primary Response Gene ◽  
Acid Region

The TIS11 primary response gene is rapidly and transiently induced by both 12-O-tetradecanoylphorbol-13-acetate and growth factors. The predicted TIS11 protein contains a 6-amino-acid repeat, YKTELC. We cloned two additional cDNAs, TIS11b and TIS11d, that contain the YKTELC sequence. TIS11, TIS11b, and TIS11d proteins share a 67-amino-acid region of sequence similarity that includes the YKTELC repeat and two cysteine-histidine containing repeats. TIS11 gene family members are not coordinately expressed: (i) unlike TIS11, the TIS11b and TIS11d mRNAs are detectable in quiescent Swiss 3T3 cells and are not dramatically induced by 12-O-tetradecanoylphorbol-13-acetate; (ii) cycloheximide superinduction does not occur for TIS11b and TIS11d; and (iii) unlike TIS11, TIS11b expression is extinguished in PC12 pheochromocytoma cells.

scholarly journals Dimerization of the Papillomavirus E2 Protein Is Required for Efficient Mitotic Chromosome Association and Brd4 Binding

2008 ◽  
Vol 82 (15) ◽  
pp. 7298-7305 ◽  
Author(s):  
Juan Cardenas-Mora ◽  
Jonathan E. Spindler ◽  
Moon Kyoo Jang ◽  
Alison A. McBride
Keyword(s):  
Dna Binding ◽  
Fluorescent Protein ◽  
Binding Domain ◽  
Bovine Papillomavirus ◽  
Dna Binding Domain ◽  
Transactivation Domain ◽  
Mitotic Chromosomes ◽  
Dimerization Domain ◽  
E2 Protein ◽  
Binding Function

ABSTRACT The E2 proteins of several papillomaviruses link the viral genome to mitotic chromosomes to ensure retention and the efficient partitioning of genomes into daughter cells following cell division. Bovine papillomavirus type 1 E2 binds to chromosomes in a complex with Brd4, a cellular bromodomain protein. Interaction with Brd4 is also important for E2-mediated transcriptional regulation. The transactivation domain of E2 is crucial for interaction with the Brd4 protein; proteins lacking or mutated in this domain do not interact with Brd4. However, we found that the C-terminal DNA binding/dimerization domain of E2 is also required for efficient binding to Brd4. Mutations that eliminated the DNA binding function of E2 had no effect on the ability of E2 to interact with Brd4, but an E2 protein with a mutation that disrupted C-terminal dimerization bound Brd4 with greatly reduced efficiency. Furthermore, E2 proteins in which the C-terminal domains were replaced with the dimeric DNA binding domain of EBNA-1 or Gal4 bound efficiently to the Brd4 protein, but the replacement of the E2 C-terminal domain with a monomeric red fluorescent protein did not rescue efficient Brd4 binding. Thus, E2 bound to Brd4 most efficiently as a dimer. To prove this finding further, the E2 DNA binding domain was replaced with an FKBP12-derived domain in which dimerization was regulated by a bivalent ligand. This fusion protein bound Brd4 efficiently only in the presence of the ligand, confirming that a dimer of E2 was required. Correspondingly, E2 proteins that could dimerize were able to bind to mitotic chromosomes much more efficiently than monomeric E2 polypeptides.

scholarly journals Interaction of the Bovine Papillomavirus E2 Protein with Brd4 Tethers the Viral DNA to Host Mitotic Chromosomes

Cell ◽  
2004 ◽  
Vol 117 (3) ◽  
pp. 349-360 ◽  
Author(s):  
Jianxin You ◽  
Jennie L Croyle ◽  
Akiko Nishimura ◽  
Keiko Ozato ◽  
Peter M Howley
Keyword(s):  
Bovine Papillomavirus ◽  
Mitotic Chromosomes ◽  
Viral Dna ◽  
E2 Protein

scholarly journals Characterization of Cichopeptins, New Phytotoxic Cyclic Lipodepsipeptides Produced by Pseudomonas cichorii SF1-54 and Their Role in Bacterial Midrib Rot Disease of Lettuce

2015 ◽  
Vol 28 (9) ◽  
pp. 1009-1022 ◽  
Author(s):  
Chien-Jui Huang ◽  
Ellen Pauwelyn ◽  
Marc Ongena ◽  
Delphine Debois ◽  
Valerie Leclère ◽  
...  
Keyword(s):  
Amino Acid ◽  
Glycine Betaine ◽  
Sequence Similarity ◽  
Similar Rate ◽  
Peptide Sequence ◽  
Pseudomonas Cichorii ◽  
Cyclic Lipopeptides ◽  
Amino Acid Peptide ◽  
Fatty Acid Chain ◽  
Tomato Pathogen

The lettuce midrib rot pathogen Pseudomonas cichorii SF1-54 produces seven bioactive compounds with biosurfactant properties. Two compounds exhibited necrosis-inducing activity on chicory leaves. The structure of the two phytotoxic compounds, named cichopeptin A and B, was tentatively characterized. They are related cyclic lipopeptides composed of an unsaturated C12-fatty acid chain linked to the N-terminus of a 22–amino acid peptide moiety. Cichopeptin B differs from cichopeptin A only in the last C-terminal amino acid residue, which is probably Val instead of Leu/Ile. Based on peptide sequence similarity, cichopeptins are new cyclic lipopeptides related to corpeptin, produced by the tomato pathogen Pseudomonas corrugata. Production of cichopeptin is stimulated by glycine betaine but not by choline, an upstream precursor of glycine betaine. Furthermore, a gene cluster encoding cichopeptin synthethases, cipABCDEF, is responsible for cichopeptin biosynthesis. A cipA-deletion mutant exhibited significantly less virulence and rotten midribs than the parental strain upon spray inoculation on lettuce. However, the parental and mutant strains multiplied in lettuce leaves at a similar rate. These results demonstrate that cichopeptins contribute to virulence of P. cichorii SF1-54 on lettuce.

scholarly journals E1 Protein of Bovine Papillomavirus Type 1 Interferes with E2 Protein-Mediated Tethering of the Viral DNA to Mitotic Chromosomes

2002 ◽  
Vol 76 (7) ◽  
pp. 3440-3451 ◽  
Author(s):  
Christian Voitenleitner ◽  
Michael Botchan
Keyword(s):  
Negative Regulator ◽  
Bovine Papillomavirus ◽  
Wild Type ◽  
Mitotic Chromosomes ◽  
Viral Dna ◽  
Nuclear Retention ◽  
E2 Protein ◽  
Protein Levels ◽  
E1 Protein ◽  
Dividing Cells

ABSTRACT Eukaryotic viruses can maintain latency in dividing cells as extrachromosomal plasmids. It is therefore of vital importance for viruses to ensure nuclear retention and proper segregation of their viral DNA. The bovine papillomavirus (BPV) E2 enhancer protein plays a key role in these processes by tethering the viral DNA to the host cell chromosomes. Viral genomes that harbor phosphorylation mutations in the E2 gene are transformation defective, and for these mutant genomes, neither the viral DNA nor the E2 protein is detected on mitotic chromosomes, while other key functions of E2 in transcription and replication were wild type. Moreover, secondary mutations in both the E2 and E1 proteins lead to suppression of the phosphorylation mutant phenotype and resulted in reattachment of the viral DNA and the E2 protein onto mitotic chromosomes, suggesting that E1 also plays a role in viral genome partitioning. The E1 protein was cytologically always excluded from mitotic chromatin, either as a suppressor allele or as the wild type. In the absence of other viral proteins, an E2 protein containing alanine substitutions for phosphorylation substrates in the hinge region (E2-A4) was detected as wild-type on mitotic chromosomes. However, when wild-type E1 protein levels were increased in cells expressing either the A4 mutant E2 proteins or wild-type E2, the E2-A4 protein was much more sensitive to chromosomal dislocation than was the wild-type protein. In contrast, suppressor alleles of E1 were not capable of such abrogation of E2 binding (A4 or wild-type) to chromosomes. These results suggest that wild-type E1 can be a negative regulator of the chromosomal attachment of E2.

scholarly journals A Single Amino Acid Substitution Converts a Transmembrane Protein Activator of the Platelet-derived Growth Factor β Receptor into an Inhibitor

2013 ◽  
Vol 288 (38) ◽  
pp. 27273-27286 ◽  
Author(s):  
Lisa M. Petti ◽  
Kristina Talbert-Slagle ◽  
Megan L. Hochstrasser ◽  
Daniel DiMaio
Keyword(s):  
Amino Acid ◽  
Growth Factor ◽  
Transmembrane Domain ◽  
Transmembrane Protein ◽  
Sequence Similarity ◽  
Human Fibroblasts ◽  
Single Amino Acid ◽  
Bovine Papillomavirus ◽  
Independent Manner ◽  
Cultured Fibroblasts

Receptors for PDGF play an important role in cell proliferation and migration and have been implicated in certain cancers. The 44-amino acid E5 protein of bovine papillomavirus binds to and activates the PDGFβ receptor (PDGFβR), resulting in oncogenic transformation of cultured fibroblasts. Previously, we isolated an artificial 36-amino acid transmembrane protein, pTM36-4, which transforms cells because of its ability to activate the PDGFβR despite limited sequence similarity to E5. Here, we demonstrated complex formation between the PDGFβR and three pTM36-4 mutants: T21E, T21Q, and T21N. T21Q retained wild type transforming activity and activated the PDGFβR in a ligand-independent manner as a consequence of binding to the transmembrane domain of the PDGFβR, but T21E and T21N were severely defective. In fact, T21N substantially inhibited E5-induced PDGFβR activation and transformation in both mouse and human fibroblasts. T21N did not prevent E5 from binding to the receptor, and genetic evidence suggested that T21N and E5 bind to nonidentical sites in the transmembrane domain of the receptor. T21N also inhibited transformation and PDGFβR activation induced by v-Sis, a viral homologue of PDGF-BB, as well as PDGF-induced mitogenesis and signaling by preventing phosphorylation of the PDGFβR at particular tyrosine residues. These results demonstrated that T21N acts as a novel inhibitor of the PDGFβR and validated a new strategy for designing highly specific short transmembrane protein inhibitors of growth factor receptors and possibly other transmembrane proteins.

scholarly journals The Cellular DNA Helicase ChlR1 Regulates Chromatin and Nuclear Matrix Attachment of the Human Papillomavirus 16 E2 Protein and High-Copy-Number Viral Genome Establishment

2016 ◽  
Vol 91 (1) ◽  
Author(s):  
Leanne Harris ◽  
Laura McFarlane-Majeed ◽  
Karen Campos-León ◽  
Sally Roberts ◽  
Joanna L. Parish
Keyword(s):  
Life Cycle ◽  
Viral Genome ◽  
Copy Number ◽  
Dna Helicase ◽  
Bovine Papillomavirus ◽  
Wild Type ◽  
E2 Protein ◽  
Human Papillomavirus 16 ◽  
Host Interaction ◽  
Cellular Dna

ABSTRACT In papillomavirus infections, the viral genome is established as a double-stranded DNA episome. To segregate the episomes into daughter cells during mitosis, they are tethered to cellular chromatin by the viral E2 protein. We previously demonstrated that the E2 proteins of diverse papillomavirus types, including bovine papillomavirus (BPV) and human papillomavirus 16 (HPV16), associate with the cellular DNA helicase ChlR1. This virus-host interaction is important for the tethering of BPV E2 to mitotic chromatin and the stable maintenance of BPV episomes. The role of the association between E2 and ChlR1 in the HPV16 life cycle is unresolved. Here we show that an HPV16 E2 Y131A mutant (E2Y131A) had significantly reduced binding to ChlR1 but retained transcriptional activation and viral origin-dependent replication functions. Subcellular fractionation of keratinocytes expressing E2Y131A showed a marked change in the localization of the protein. Compared to that of wild-type E2 (E2WT), the chromatin-bound pool of E2Y131A was decreased, concomitant with an increase in nuclear matrix-associated protein. Cell cycle synchronization indicated that the shift in subcellular localization of E2Y131A occurred in mid-S phase. A similar alteration between the subcellular pools of the E2WT protein occurred upon ChlR1 silencing. Notably, in an HPV16 life cycle model in primary human keratinocytes, mutant E2Y131A genomes were established as episomes, but at a markedly lower copy number than that of wild-type HPV16 genomes, and they were not maintained upon cell passage. Our studies indicate that ChlR1 is an important regulator of the chromatin association of E2 and of the establishment and maintenance of HPV16 episomes. IMPORTANCE Infections with high-risk human papillomaviruses (HPVs) are a major cause of anogenital and oropharyngeal cancers. During infection, the circular DNA genome of HPV persists within the nucleus, independently of the host cell chromatin. Persistence of infection is a risk factor for cancer development and is partly achieved by the attachment of viral DNA to cellular chromatin during cell division. The HPV E2 protein plays a critical role in this tethering by binding simultaneously to the viral genome and to chromatin during mitosis. We previously showed that the cellular DNA helicase ChlR1 is required for loading of the bovine papillomavirus E2 protein onto chromatin during DNA synthesis. Here we identify a mutation in HPV16 E2 that abrogates interaction with ChlR1, and we show that ChlR1 regulates the chromatin association of HPV16 E2 and that this virus-host interaction is essential for viral episome maintenance.

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