scholarly journals A Fifteen-Amino-Acid Peptide Inhibits Human Papillomavirus E1-E2 Interaction and Human Papillomavirus DNA Replication In Vitro

1998 ◽  
Vol 72 (10) ◽  
pp. 8166-8173 ◽  
Author(s):  
Hiroaki Kasukawa ◽  
Peter M. Howley ◽  
John D. Benson
Keyword(s):  
Human Papillomavirus ◽  
Amino Acid ◽  
Dna Replication ◽  
Transcriptional Activation ◽  
Hpv 16 ◽  
Hpv Dna ◽  
Amino Acid Peptide ◽  
In Vitro Replication ◽  
Papillomavirus Dna

ABSTRACT Mutation of the conserved glutamic acid residue at position 39 of human papillomavirus type 16 (HPV-16) E2 to alanine (E39A) disrupts its E1 interaction activity and its replication function in transient replication assays but does not affect E2 transcriptional activation. This E39A mutation also disrupts replication activity of HPV-16 E2 in HPV-16 in vitro DNA replication. On this basis, we designed 23- and 15-amino-acid peptides derived from HPV-16 E2 sequences flanking the E39 residue and tested the ability of these peptides to inhibit interaction between HPV-16 E1 and E2 in vitro. The inhibitory activity of these peptides was specific, since analogous peptides in which alanine was substituted for the E39 residue did not inhibit interaction. The 15-amino-acid peptide E2N-WP15 was the smallest peptide tested that effectively inhibited HPV-16 E1-E2 interaction. This peptide also inhibited in vitro replication of HPV-16 DNA. The efficacy of E2N-WP15 was not exclusive to HPV-16: this peptide also inhibited interaction of HPV-11 E1 with the E2 proteins of both HPV-11 and HPV-16 and inhibited in vitro replication with these same combinations of E1 and E2 proteins. These results provide further evidence that E1-E2 interaction is required for papillomavirus DNA replication and constitute the first demonstration that inhibition of this interaction is sufficient to prevent HPV DNA replication in vitro.

scholarly journals Transcriptional Activation of the Telomerase hTERT Gene by Human Papillomavirus Type 16 E6 Oncoprotein

2001 ◽  
Vol 75 (9) ◽  
pp. 4467-4472 ◽  
Author(s):  
Tim Veldman ◽  
Izumi Horikawa ◽  
J. Carl Barrett ◽  
Richard Schlegel
Keyword(s):  
Human Papillomavirus ◽  
Transcriptional Activation ◽  
Transcriptional Control ◽  
Regulatory Region ◽  
Rnase Protection ◽  
Hpv 16 ◽  
Htert Gene ◽  
Human Papillomavirus Type 16 ◽  
E6 Gene

ABSTRACT The E6 and E7 oncogenes of human papillomavirus type 16 (HPV-16) are sufficient for the immortalization of human genital keratinocytes in vitro. The products of these viral genes associate with p53 and pRb tumor suppressor proteins, respectively, and interfere with their normal growth-regulatory functions. The HPV-16 E6 protein has also been shown to increase the telomerase enzyme activity in primary epithelial cells by an unknown mechanism. We report here that a study using reverse transcription-PCR and RNase protection assays in transduced primary human foreskin keratinocytes (HFKs) shows that the E6 gene (but not the E7 gene) increases telomerase hTERT gene transcription coordinately with E6-induced telomerase activity. In these same cells, the E6 gene induces a 6.5-fold increase in the activity of a 1,165-bp 5′ promoter/regulatory region of the hTERT gene, and this induction is attributable to a minimal 251-bp sequence (−211 to +40). Furthermore, there is a 35-bp region (+5 to +40) within this minimal E6-responsive promoter that is responsible for 60% of E6 activity. Although the minimal hTERT promoter contains Myc-responsive E-box elements and recent studies have suggested a role for Myc protein in hTERT transcriptional control, we found no alterations in the abundance of either c-Myc or c-Mad in E6-transduced HFKs, suggesting that there are other or additional transcription factors critical for regulating hTERT expression.

scholarly journals Identification of Peptides That Inhibit the DNA Binding, trans-Activator, and DNA Replication Functions of the Human Papillomavirus Type 11 E2 Protein

2004 ◽  
Vol 78 (5) ◽  
pp. 2637-2641 ◽  
Author(s):  
Su-Jun Deng ◽  
Kenneth H. Pearce ◽  
Eric P. Dixon ◽  
Kelly A. Hartley ◽  
Thomas B. Stanley ◽  
...  
Keyword(s):  
Cell Culture ◽  
Human Papillomavirus ◽  
Dna Replication ◽  
Transcriptional Activation ◽  
Synthetic Peptides ◽  
Dna Interaction ◽  
Peptide Library ◽  
Random Peptide Library ◽  
Random Peptide

ABSTRACT Peptide antagonists of the human papillomavirus type 11 (HPV-11) E2-DNA association were identified using a filamentous bacteriophage random peptide library. Synthetic peptides antagonized the E2-DNA interaction, effectively blocked E2-mediated transcriptional activation of a reporter gene in cell culture, and inhibited E1-E2-mediated HPV-11 DNA replication in vitro. These peptides may prove to be useful tools for characterizing E2 function and for exploring the effectiveness of E2-inhibitor-based treatments for HPV-associated diseases.

scholarly journals The Inhibitory Action of P56 on Select Functions of E1 Mediates Interferon's Effect on Human Papillomavirus DNA Replication

2010 ◽  
Vol 84 (24) ◽  
pp. 13036-13039 ◽  
Author(s):  
Paramananda Saikia ◽  
Volker Fensterl ◽  
Ganes C. Sen
Keyword(s):  
Human Papillomavirus ◽  
Dna Replication ◽  
Dna Binding ◽  
Inhibitory Action ◽  
Dna Helicase ◽  
Single Amino Acid ◽  
Viral Dna ◽  
Viral Dna Replication ◽  
Hpv Dna ◽  
Papillomavirus Dna

ABSTRACT The interferon (IFN)-induced protein P56 inhibits human papillomavirus (HPV) DNA replication by binding to HPV E1, which has several distinct functions in initiating viral DNA replication. Here, we determined that P56 inhibited HPV type 18 (HPV18) E1's DNA helicase activity, E2 binding, and HPV Ori sequence-specific DNA binding but not nonspecific DNA binding. We observed that deletion of a single amino acid, F399, produced an E1 mutant that could not bind P56. This E1 mutant retained its ability to support Ori DNA replication, but this activity was not inhibited by IFN, demonstrating that P56 is the principal executor of the anti-HPV action of IFN.

scholarly journals Human TATA Binding Protein Inhibits Human Papillomavirus Type 11 DNA Replication by Antagonizing E1-E2 Protein Complex Formation on the Viral Origin of Replication

2002 ◽  
Vol 76 (10) ◽  
pp. 5014-5023 ◽  
Author(s):  
Kelly A. Hartley ◽  
Kenneth A. Alexander
Keyword(s):  
Transcription Factors ◽  
Human Papillomavirus ◽  
Dna Replication ◽  
Dna Binding ◽  
Tata Binding Protein ◽  
Keratinocyte Differentiation ◽  
Origin Of Replication ◽  
Viral Origin ◽  
Hpv Dna

ABSTRACT The human papillomavirus (HPV) protein E2 possesses dual roles in the viral life cycle. By interacting directly with host transcription factors in basal keratinocytes, E2 promotes viral transcription. As keratinocyte differentiation progresses, E2 associates with the viral helicase, E1, to activate vegetative viral DNA replication. How E2's major role switches from transcription to replication during keratinocyte differentiation is not understood, but the presence of a TATA site near the viral origin of replication led us to hypothesize that TATA-binding protein (TBP) could affect HPV replication. Here we show that the C-terminal domain of TBP (TBPc) is a potent inhibitor of E2-stimulated HPV DNA replication in vitro (50% inhibitory concentration = 0.56 nM). Increasing the E1 concentration could not overcome TBPc inhibition in replication assays, indicating that TBPc is a noncompetitive inhibitor of E1 binding. While direct E2-TBPc association could be demonstrated, this interaction could not fully account for the mechanism of TBPc-mediated inhibition of viral replication. Because E2 supports sequence-specific binding of E1 to the viral ori, we proposed that TBPc antagonizes E1-ori association indirectly through inhibition of E2-DNA binding. Indeed, TBPc potently antagonized E2 binding to DNA in the absence (Ki = 0.5 ± 0.1 nM) and presence (Ki = 0.6 ± 0.3 nM) of E1. Since E2 and TBPc cannot be coadjacent on viral sequences, direct DNA-binding competition between TBPc and E2 was responsible for replication inhibition. Given the ability of TBPc to inhibit HPV DNA replication in vitro and data indicating that TBPc antagonized E2-ori association, we propose that transcription factors regulate HPV DNA replication as well as viral transcription.

scholarly journals Functional interaction of a novel cellular protein with the papillomavirus E2 transactivation domain.

1997 ◽  
Vol 17 (12) ◽  
pp. 7208-7219 ◽  
Author(s):  
D E Breiding ◽  
F Sverdrup ◽  
M J Grossel ◽  
N Moscufo ◽  
W Boonchai ◽  
...  
Keyword(s):  
Amino Acid ◽  
Dna Replication ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Point Mutations ◽  
Cellular Protein ◽  
Bovine Papillomavirus ◽  
Transactivation Domain

The transactivation domain (AD) of bovine papillomavirus type 1 E2 stimulates gene expression and DNA replication. To identify cellular proteins that interact with this 215-amino-acid domain, we used a transactivation-defective mutant as bait in the yeast two-hybrid screen. In vitro and in vivo results demonstrate that the cDNA of one plasmid isolated in this screen encodes a 37-kDa nuclear protein that specifically binds to an 82-amino-acid segment within the E2 AD. Mutants with point mutations within this E2 domain were isolated based on their inability to interact with AMF-1 and were found to be unable to stimulate transcription. These mutants also exhibited defects in viral DNA replication yet retained binding to the viral E1 replication initiator protein. Overexpression of AMF-1 stimulated transactivation by both wild-type E2 and a LexA fusion to the E2 AD, indicating that AMF-1 is a positive effector of the AD of E2. We conclude that interaction with AMF-1 is necessary for the transcriptional activation function of the E2 AD in mammalian cells.

scholarly journals Pseudoepitheliomatous keratotic and micaceous balanitis: low-risk human papilloma virus detection in two further cases

2020 ◽  
pp. 095646242096194
Author(s):  
Marialuisa Corbeddu ◽  
Luca Pilloni ◽  
Roberta Satta ◽  
Laura Atzori ◽  
Franco Rongioletti
Keyword(s):  
Human Papillomavirus ◽  
Verrucous Carcinoma ◽  
Virus Detection ◽  
Low Risk ◽  
Hpv Testing ◽  
Papilloma Virus ◽  
Hpv Dna ◽  
Proliferative Markers ◽  
Male Patients ◽  
Papillomavirus Dna

We report two cases of histologically documented pseudoepitheliomatous keratotic and micaceous balanitis in middle-aged male patients, which showed positivity for low-risk serotype human papillomavirus DNA. To our knowledge, only one other case has been documented. Further immunohistochemical proliferative markers were performed and compared to literature findings in penile epithelial proliferations. Evolution to invasive verrucous carcinoma has been associated with absence of HPV DNA. Thus, if confirmed by further studies, HPV testing should be included in pseudoepitheliomatous keratotic and micaceous balanitis assessment to address prognosis, and management.

scholarly journals Inhibition of transcription starting from bacteriophage lambda pR promoter during the stringent response in Escherichia coli: implications for lambda DNA replication.

1995 ◽  
Vol 42 (2) ◽  
pp. 233-239 ◽  
Author(s):  
A Szalewska-Pałasz ◽  
G Wegrzyn
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Dna Replication ◽  
Transcriptional Activation ◽  
Plasmid Dna ◽  
Bacteriophage Lambda ◽  
Stringent Response ◽  
Lacz Gene ◽  
Relative Level ◽  
Lambda Dna

Replication of lambda plasmid DNA is halted in amino acid-starved wild type (stringent) strains whereas it proceeds in relA (relaxed) mutants. The only transcription which could be important in lambda plasmid DNA replication in amino acid-starved Escherichia coli cells is that starting from the pR promoter. Using a fusion which consists of the lacZ gene under the control of bacteriophage lambda pR promoter we found that transcription starting from this promoter was inhibited during the stringent, but not the relaxed, response in E. coli. We confirmed our conclusion by estimating the relative level of the pR transcript by RNA-DNA hybridization. We propose that decreased transcription from the pR promoter which serves as transcriptional activation of ori lambda is responsible for inhibition of lambda plasmid replication during the stringent response. The results presented in this paper, combined with our recent findings (published elsewhere), indicate that the transcriptional activation of ori lambda may be a main regulatory process controlling lambda DNA replication not only during the relaxed response but also in normal growth conditions.

scholarly journals Incorporation of Cervista Human Papillomavirus 16/18 Assay Into Algorithms for Classifying Human Papillomavirus Status in Formalin-Fixed, Paraffin-Embedded Head and Neck Squamous Carcinoma Specimens

2018 ◽  
Vol 143 (3) ◽  
pp. 356-361
Author(s):  
Ming Guo ◽  
Abha Khanna ◽  
Jianping Wang ◽  
Michelle D. Williams ◽  
Neda Kalhor ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Human Papillomavirus ◽  
Ffpe Tissue ◽  
Hpv 16 ◽  
Hpv Dna ◽  
Formalin Fixed Paraffin ◽  
Hpv Status ◽  
Formalin Fixed Paraffin Embedded ◽  
Hpv Assays ◽  
Formalin Fixed

Context.— Human papillomavirus (HPV) DNA in situ hybridization (ISH) assay and p16 immunohistochemistry (IHC) are used to determine high-risk HPV status in formalin-fixed, paraffin-embedded (FFPE) tissues in oropharyngeal squamous cell carcinoma (SCC). Although high sensitivity and specificity for HPV can be obtained by combined p16 IHC and HPV DNA ISH, the occasional discrepancy between these assays has prompted evaluation of Cervista HPV assays in FFPE tissue from patients with oropharyngeal SCC. Objective.— To compare the efficacy of Cervista HPV 16/18 and Cervista HPV HR assay to that of HPV DNA ISH assay and p16 IHC in FFPE tissue in head and neck squamous cell carcinoma of oropharyngeal origin. Design.— Archived FFPE tissue from 84 patients with SCC of oropharyngeal origin and available HPV DNA ISH and p16 IHC test results were tested with the Cervista HPV 16/18 assay and further verified by polymerase chain reaction (PCR)–based HPV16/18 genotyping tests in cases with discrepancy. Results.— Of the 84 specimens, 75% (63 of 84) were positive and 16% (13 of 84) had discrepant or equivocal findings by p16 IHC and HPV DNA ISH testing. Use of Cervista HPV assays, either to clarify discrepant/equivocal findings or as confirmation after initial p16 IHC/HPV DNA ISH tests, identified 81% (68 of 84) of HPV-positive cases without equivocal HPV results. Five of 13 cases with discrepancy or equivocal HPV DNA ISH results tested positive for HPV16 or HPV18 by Cervista HPV 16/18 assay, which was further confirmed by PCR-based HPV 16/18 genotyping. Conclusions.— The Cervista HPV assays are a reasonable alternative to HPV DNA ISH in determining HPV status in FFPE tissue specimens from patients with oropharyngeal SCC.

scholarly journals Identification of amino acids essential for DNA binding and dimerization in p67SRF: implications for a novel DNA-binding motif

1993 ◽  
Vol 13 (1) ◽  
pp. 123-132
Author(s):  
A D Sharrocks ◽  
H Gille ◽  
P E Shaw
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Serum Response Factor ◽  
Alpha Helix ◽  
Site Directed Mutagenesis ◽  
Binding Motif ◽  
Amino Acid Peptide ◽  
Serum Response

The serum response factor (p67SRF) binds to a palindromic sequence in the c-fos serum response element (SRE). A second protein, p62TCF binds in conjunction with p67SRF to form a ternary complex, and it is through this complex that growth factor-induced transcriptional activation of c-fos is thought to take place. A 90-amino-acid peptide, coreSRF, is capable for dimerizing, binding DNA, and recruiting p62TCF. By using extensive site-directed mutagenesis we have investigated the role of individual coreSRF amino acids in DNA binding. Mutant phenotypes were defined by gel retardation and cross-linking analyses. Our results have identified residues essential for either DNA binding or dimerization. Three essential basic amino acids whose conservative mutation severely reduced DNA binding were identified. Evidence which is consistent with these residues being on the face of a DNA binding alpha-helix is presented. A phenylalanine residue and a hexameric hydrophobic box are identified as essential for dimerization. The amino acid phasing is consistent with the dimerization interface being presented as a continuous region on a beta-strand. A putative second alpha-helix acts as a linker between these two regions. This study indicates that p67SRF is a member of a protein family which, in common with many DNA binding proteins, utilize an alpha-helix for DNA binding. However, this alpha-helix is contained within a novel domain structure.

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