Cellular binding partners of the human papillomavirus E6 protein

The high-risk strains of human papillomavirus (HR-HPV) are known to be causative agents of cervical cancer and have recently also been implicated in cancers of the oropharynx. E6 is a potent oncogene of HR-HPVs, and its role in the progression to malignancy has been and continues to be explored. E6 is known to interact with and subsequently inactivate numerous cellular proteins pivotal in the mediation of apoptosis, transcription of tumor suppressor genes, maintenance of epithelial organization, and control of cell proliferation. Binding of E6 to these proteins cumulatively contributes to the oncogenic potential of HPV. This paper provides an overview of these cellular protein partners of HR-E6, the motifs known to mediate oncoprotein binding, and the agents that have the potential to interfere with E6 expression and activity and thus prevent the subsequent progression to oncogenesis

New Human Papilloma Virus E2 Transcription Factor Mimics: A Tripyrrole-Peptide Conjugate with Tight and Specific DNA-Recognition

BACKGROUND: Human papillomavirus (HPV) is the main causative agent of cervical cancer, particularly high risk strains such us HPV-16, -18 and -31. The viral encoded E2 protein acts as a transcriptional modulator and exerts a key role in viral DNA replication. Thus, E2 constitutes an attractive target for developing antiviral agents. E2 is a homodimeric protein that interacts with the DNA target through an α-helix of each monomer. However, a peptide corresponding to the DNA recognition helix of HPV-16 E2 binds DNA with lower affinity than its full-length DNA binding domain. Therefore, in an attempt to promote the DNA binding of the isolated peptide, we have designed a conjugate compound of the E2 α-helix peptide and a derivative of the antibiotic distamycin, which involves simultaneous minor- and major-groove interactions. METHODOLOGY/PRINCIPAL FINDINGS: An E2 α-helix peptide-distamycin conjugate was designed and synthesized. It was characterized by NMR and CD spectroscopy, and its DNA binding properties were investigated by CD, DNA melting and gel shift experiments. The coupling of E2 peptide with distamycin does not affect its structural properties. The conjugate improves significantly the affinity of the peptide for specific DNA. In addition, stoichiometric amounts of specific DNA increase meaningfully the helical population of the peptide. The conjugate enhances the DNA binding constant 50-fold, maintaining its specificity. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that peptide-distamycin conjugates are a promising tool to obtain compounds that bind the E2 target DNA-sequences with remarkable affinity and suggest that a bipartite major/minor groove binding scaffold can be a useful approach for therapeutic treatment of HPV infection

Insulin Gene Expression Is Regulated by DNA Methylation

BACKGROUND:Insulin is a critical component of metabolic control, and as such, insulin gene expression has been the focus of extensive study. DNA sequences that regulate transcription of the insulin gene and the majority of regulatory factors have already been identified. However, only recently have other components of insulin gene expression been investigated, and in this study we examine the role of DNA methylation in the regulation of mouse and human insulin gene expression. METHODOLOGY/PRINCIPAL FINDINGS:Genomic DNA samples from several tissues were bisulfite-treated and sequenced which revealed that cytosine-guanosine dinucleotide (CpG) sites in both the mouse Ins2 and human INS promoters are uniquely demethylated in insulin-producing pancreatic beta cells. Methylation of these CpG sites suppressed insulin promoter-driven reporter gene activity by almost 90% and specific methylation of the CpG site in the cAMP responsive element (CRE) in the promoter alone suppressed insulin promoter activity by 50%. Methylation did not directly inhibit factor binding to the CRE in vitro, but inhibited ATF2 and CREB binding in vivo and conversely increased the binding of methyl CpG binding protein 2 (MeCP2). Examination of the Ins2 gene in mouse embryonic stem cell cultures revealed that it is fully methylated and becomes demethylated as the cells differentiate into insulin-expressing cells in vitro. CONCLUSIONS/SIGNIFICANCE:Our findings suggest that insulin promoter CpG demethylation may play a crucial role in beta cell maturation and tissue-specific insulin gene expression

Association between hMLH1 hypermethylation and JC virus (JCV) infection in human colorectal cancer (CRC)

Incorporation of viral DNA may interfere with the normal sequence of human DNA bases on the genetic level or cause secondary epigenetic changes such as gene promoter methylation or histone acetylation. Colorectal cancer (CRC) is the second leading cause of cancer mortality in the USA. Chromosomal instability (CIN) was established as the key mechanism in cancer development. Later, it was found that CRC results not only from the progressive accumulation of genetic alterations but also from epigenetic changes. JC virus (JCV) is a candidate etiologic factor in sporadic CRC. It may act by stabilizing β-catenin, facilitating its entrance to the cell nucleus, initialing proliferation and cancer development. Diploid CRC cell lines transfected with JCV-containing plasmids developed CIN. This result provides direct experimental evidence for the ability of JCV T-Ag to induce CIN in the genome of colonic epithelial cells. The association of CRC hMLH1 methylation and tumor positivity for JCV was recently documented. JC virus T-Ag DNA sequences were found in 77% of CRCs and are associated with promoter methylation of multiple genes. hMLH1 was methylated in 25 out of 80 CRC patients positive for T-Ag (31%) in comparison with only one out of 11 T-Ag negative cases (9%). Thus, JCV can mediate both CIN and aberrant methylation in CRC. Like other viruses, chronic infection with JCV may induce CRC by different mechanisms which should be further investigated. Thus, gene promoter methylation induced by JCV may be an important process in CRC and the polyp-carcinoma sequence

Epigenetic Repression of RARRES1 Is Mediated by Methylation of a Proximal Promoter and a Loss of CTCF Binding

The cis-acting promoter element responsible for epigenetic silencing of retinoic acid receptor responder 1 (RARRES1) by methylation is unclear. Likewise, how aberrant methylation interplays effectors and thus affects breast neoplastic features remains largely unknown.We first compared methylation occurring at the sequences (-664~+420) flanking the RARRES1 promoter in primary breast carcinomas to that in adjacent benign tissues. Surprisingly, tumor cores displayed significantly elevated methylation occurring solely at the upstream region (-664~-86), while the downstream element (-85~+420) proximal to the transcriptional start site (+1) remained largely unchanged. Yet, hypermethylation at the former did not result in appreciable silencing effect. In contrast, the proximal sequence displayed full promoter activity and methylation of which remarkably silenced RARRES1 transcription. This phenomenon was recapitulated in breast cancer cell lines, in which methylation at the proximal region strikingly coincided with downregulation. We also discovered that CTCF occupancy was enriched at the unmethylayed promoter bound with transcription-active histone markings. Furthermore, knocking-down CTCF expression hampered RARRES1 expression, suggesting CTCF positively regulated RARRES1 transcription presumably by binding to unmethylated promoter poised at transcription-ready state. Moreover, RARRES1 restoration not only impeded cell invasion but also promoted death induced by chemotherapeutic agents, denoting its tumor suppressive effect. Its role of attenuating invasion agreed with data generated from clinical specimens revealing that RARRES1 was generally downregulated in metastatic lymph nodes compared to the tumor cores.This report delineated silencing of RARRES1 by hypermethylation is occurring at a proximal promoter element and is associated with a loss of binding to CTCF, an activator for RARRES1 expression. We also revealed the tumor suppressive roles exerted by RARRES1 in part by promoting breast epithelial cell death and by impeding cell invasion that is an important property for metastatic spread

Lipid Peroxidation After Intracortical Injection of Ferric Chloride Increases the Incidence of Seizures in Young Rats

Clinical studies have shown that the incidence of early posttraumatic seizures ishigher in children than in adults and it has been proposed that iron-induced lipidperoxidation has an important role in the development of epileptogenic foci. In this study,we examined some of the hypothesized reasons for the difference in the incidence ofearly posttraumatic seizures between young and adult rats. Twelve young and twelveadult rats were randomized into 4 groups. Group 1 and 2 were control groups, eachcomprising of 6 young rats and 6 adult rats respectively and were given intracorticalinjections of normal saline. Group 3 and 4 were injury groups, again comprising 6 youngrats and 6 adult rats respectively and were given intracortical injections of FeCl3. All ratswere observed for 6 hours post injection for the occurrence of seizures and were thenkilled. The injected hemispheres were extirpated and tested for malondialdehyde (MDA)level and superoxide dismutase (SOD) activity as indices of oxidative damage. Resultsshowed that seizures were observed only in Group 3. Increased MDA level and decreasedSOD activity were observed in Group 3 (ANOVA, p<0.001). Increased MDA levels anddecreased SOD activity were significantly higher in rats with seizures (Group 3) than inthose without seizures (independent t-test, p<0.001). We conclude was that differentlevels of lipid peroxidation induced by intracortical ferric chloride injection may accountfor the different seizure incidence between young and adult rat

Free flap head and neck reconstruction in the elderly: What is the impact on quality of life?

Morphofunctional reconstruction is a pivotal aspect in the surgery of head and neck neoplasms: Nowadays, microvascular free flap surgery represents the gold standard. In choosing the surgical technique, the effects on residual quality of life, especially in elderly people, usually considered more fragile and so often excluded from microsurgical procedures, must be taken into account. This multicentre study evaluated the quality of life index in patients more than 75 years of age and who underwent to head and neck microsurgical reconstruction. Data from patients aged > 75 years at the time of major head and neck reconstruction conducted with free flaps between 1 January 2005 and 30 June 2015 were analysed retrospectively. We administered the Italian version of Quality of Life questionnaire SF-36, at least 24 months after surgery. Results were compared to those for the general Italian population of the same age. We enrolled 39 patients with an average age of 80.6 years. The results did not differ significantly from the reference population. The international literature has already shown that chronologic age is not a valid parameter to determine the surgical treatment modality. Even considering the quality of residual life, our study supports the indication for free-flap reconstruction of head and neck defects in the elderly, confirming its effectiveness in this population

Motion of wave fronts in semiconductor superlattices

An analysis of wave front motion in weakly coupled doped semiconductor superlattices is presented. If a dimensionless doping is sufficiently large, the superlattice behaves as a discrete system presenting front propagation failure and the wave fronts can be described near the threshold currents J i (iϭ1,2) at which they depin and move. The wave front velocity scales with current as ͉JϪJ i ͉ 1/2 . If the dimensionless doping is low enough, the superlattice behaves as a continuum system and wave fronts are essentially shock waves whose velocity obeys an equal area rule

Apparatus for measuring a sorbate dispersed in a fluid stream

A sensitive, miniature apparatus was designed for measuring low concentrations of a sorbate dispersed in a fluid stream. The device consists of an elongated body having a surface capable of sorbing an amount of the sorbate proportional to the concentration in the fluid stream and propagating acoustic energy along its length. The acoustic energy is converted to an electrical output signal corresponding to the concentration of sorbate in the fluid stream. The device can be designed to exhibit high sensitivity to extremely small amounts of sorbate dispersed in a fluid stream and to exhibit low sensitivity to large amounts of sorbate. Another advantage is that the apparatus may be formed in a microminiature size and at a low cost using bath microfabrication technology