Evaluation of Regulation of Late Gene Expression in Human Papillomavirus-infected Cervical Epithelia

Cervical cancer is second only to breast cancer as the most common cancer among women worldwide and is caused by Human Papillomavirus (HPV). The aim of this study was to evaluate the role of RNA-binding proteins in the expression of HPV in cervical cells and clinical samples. Firstly, an audit was performed on cervical biopsy samples taken by laser loop excision over 5 years in the National Maternity Hospital, Holles Street, Dublin, Ireland. The presence of HPV and HPV-16 was evaluated on fifty of these samples and the distribution of multiple RNA-binding proteins, including SR proteins and hnRNPs, was evaluated and related to differentiations (cytokeratins, syndecan) and proliferation-associated (PCNA, p161INK4a) biomarkers. The samples consisted of histologically normal cervical epithelium, HPV-induced low-grade and high-grade pre-malignant lesions and cervical cancers. Low expression of these proteins was detected in basal epithelial cells of normal cervix, with increased expression in intermediate layers, and a lack of expression in the upper superficial layers. Expression of all RNA-binding proteins increased in neoplastic lesions and highest expression was demonstrated in intermediate layers, and a lack of expression in the upper superficial layer. Expression of all RNA-binding proteins increased in neoplastic lesions and highest expression was demonstrated in cervical cancers. The expression profile of the RNA-binding proteins was similar to PCNA expression in normal and lesional cervical epithelia, but was not associated with HPV-16 status, p16INK4a or differentiation markers. The effect of a number of these proteins on transcription of the late HPV genes was evaluated in modified HeLa cell lines (pBEL and pBELM) by transfection studies, with effects evaluated using real-time PCR and Northern blot analyses. It was determined that adenoviral E4orf4, splicing proteins SRp30c and PTB as well as HPV-16 E2 can induce the expression of the late HPV genes and these were shown to alter the ratio of expression of HPV L1 mRNAs, demonstrating that these proteins may play a role in the differential expression of proteins during the HPV life-cycle

Antimicrobial Producing Bacteria Isolated From Petroleum-Laced Hypersaline Soil

Given the alternative functions of “antibiotics” as communication molecules or participants in metabolism, it seems probable that production might be influenced by factors such as nutrient availability, interactions with neighboring microbes, and/or colony or community structure and maturity. With this in mind, the present study aimed to broaden the scope of the search for novel antibiotics by experimenting with the following parameters: source of bacterial isolation, growth and assay media, and culturing techniques. Bacteria for this study were isolated from two categories of soil (petroleum-contaminated or uncontaminated) to compare diversity and antimicrobial activity. Compared to the uncontaminated soil, isolates of the petroleum-contaminated soil were as diverse and antimicrobial activity was as frequent. Antimicrobial assays were done on three different types of agar, including the standard Mueller-Hinton and two types of medium typically used for fungal growth, Yeast Peptone Dextrose (YPD) and Yeast Mold (YM). Compared to results on Mueller-Hinton, much more antimicrobial activity was seen when using YPD and YM. Finally, spent media assays were performed with pure and mixed cultures to determine if exposure to a target pathogen affects the production of antimicrobial substances by soil isolates. Those bacteria with activity against Pseudomonas aeruginosa in perpendicular streak tests were grown as mixed cultures with P. aeruginosa. In the case of Bacillus amyloliquefaciens and Pseudomonas marginalis, discs impregnated with concentrated spent media from these mixed cultures resulted in significant dose-dependent inhibition of P. aeruginosa. The same assay using pure cultures showed no inhibition

Identification of a 17-Nucleotide Splicing Enhancer in HPV-16 L1 that Counteracts the Effect of Multiple hnRNP A1-binding Splicing Silencers

Human papillomavirus type 16 (HPV-16) infections can in rare cases persist and cause lesions that may progress to cervical cancer. Cells in the lesions are not permissive for virus production, nor are cervical cancer cells. The intracellular environment is such that it prevents production of the highly immunogenic, viral structural proteins L1 and L2. One may speculate that inhibition of L1 and L2 expression is a prerequisite for persistence and cancer progression.We have therefore investigated how expression of HPV-16 L1 is regulated.We found that the only splice site in the HPV-16 late region, which is used to produce L1 mRNAs, is under control of a splicing enhancer located in the 17 nucleotides immediately downstream of the splice site. However, the function of this enhancer in cervical cancer cells is largely overshadowed by multiple splicing silencers in the late region which bind to hnRNP A1. High levels of hnRNP A1 therefore inhibit HPV-16 L1 expression. Immunohistological analysis of cervical epithelia revealed that hnRNPA1 is expressed primarily in the lower layers of the epithelium. hnRNPA1 is undetectable in terminally differentiated cells that can express HPV-16 late genes, which supports the conclusion that high levels of hnRNP A1 inhibit HPV-16 L1 expression

Adenovirus E4orf4 Induces HPV-16 Late L1 mRNA Production

The adenovirus E4orf4 protein regulates the switch from early to late gene expression during the adenoviral replication cycle. Here we report that overexpression of adenovirus E4orf4 induces human papillomavirus type 16 (HPV-16) late gene expression from subgenomic expression plasmids. E4orf4 specifically overcomes the negative effects of two splicing silencers at the two late HPV-16 splice sites SD3632 and SA5639. This results in the production of HPV-16 spliced L1 mRNAs. We show that the interaction of E4orf4 with protein phosphatase 2A (PP2A) is necessary for induction of HPV-16 late gene expression. Also an E4orf4 mutant that fails to bind the cellular splicing factor ASF/SF2 fails to induce L1 mRNA production. Collectively, these results suggest that dephosphorylation of SR proteins by E4orf4 activates HPV-16 late gene expression. Indeed, a mutant ASF/SF2 protein in which the RS-domain had been deleted could itself induce HPV-16 late gene expression, whereas wild type ASF/SF2 could not

Caspase-11 regulates the tumour suppressor function of STAT1 in a murine model of colitis-associated carcinogenesis

Murine inflammatory caspase-11 has an important role in intestinal epithelial inflammation and barrier function. Activation of the non-canonical inflammasome, mediated by caspase-11, serves as a regulatory pathway for the production of the proinflammatory cytokines IL-1β and IL-18, and has a key role in pyroptotic cell death. We have previously demonstrated a protective role for caspase-11 during dextran sulphate sodium (DSS)-induced colitis, however the importance of caspase-11 during colorectal tumour development remains unclear. Here, we show that Casp11−/− mice are highly susceptible to the azoxymethane (AOM)-DSS model of colitis-associated cancer (CAC), compared to their wild type (WT) littermates. We show that deficient IL-18 production occurs at initial inflammation stages of disease, and that IL-1β production is more significantly impaired in Casp11−/− colons during established CAC. We identify defective STAT1 activation in Casp11−/− colons during disease progression, and show that IL-1β signalling induces caspase-11 expression and STAT1 activation in primary murine macrophages and intestinal epithelial cells. These findings uncover an anti-tumour role for the caspase-11 and the non-canonical inflammasome during CAC, and suggest a critical role for caspase-11, linking IL-1β and STAT1 signalling pathways

Calnexin, an ER-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a leading cause of cancer mortality in the Western world and commonly treated with genotoxic chemotherapy. Stress in the endoplasmic reticulum (ER) was implicated to contribute to chemotherapeutic resistance. Hence, ER stress related protein may be of prognostic or therapeutic significance. METHODS: The expression levels of ER stress proteins calnexin, calreticulin, GRP78 and GRP94 were determined in n = 23 Stage II and III colon cancer fresh frozen tumour and matched normal tissue samples. Data were validated in a cohort of n = 11 rectal cancer patients treated with radiochemotherapy in the neoadjuvant setting. The calnexin gene was silenced using siRNA in HCT116 cells. RESULTS: There were no increased levels of ER stress proteins in tumour compared to matched normal tissue samples in Stage II or III CRC. However, increased calnexin protein levels were predictive of poor clinical outcome in the patient cohort. Data were validated in the rectal cancer cohort treated in the neoadjuvant setting. Calnexin gene-silencing significantly reduced cell survival and increased cancer cell susceptibility to 5FU chemotherapy. CONCLUSION: Increased tumour protein levels of calnexin may be of prognostic significance in CRC, and calnexin may represent a potential target for future therapies

Experiences of people with dementia and informal caregivers with post-diagnostic support: Data from the international COGNISANCE study

OBJECTIVES: The study aims to describe people with dementia and informal caregivers' respective experiences of support after diagnosis and compares these experiences. Additionally, we determine how people with dementia and informal caregivers who are satisfied with support differ from those dissatisfied. METHODS: A cross-sectional survey study in Australia, Canada, the Netherlands, Poland, and United Kingdom was carried out to examine people with dementia and informal caregivers experience with support (satisfaction with information, access to care, health literacy, and confidence in ability to live well with dementia). The separate surveys contained closed questions. Analysis consisted of descriptive statistics and Chi-square tests. RESULTS: Ninety people with dementia and 300 informal caregivers participated, and 69% of people with dementia and 67% of informal caregivers said support after diagnosis helped them deal more efficiently with their concerns. Up to one-third of people with dementia and informal caregivers were dissatisfied with information about management, prognosis, and strategies for living positively. Few people with dementia (22%) and informal caregivers (35%) received a care plan. People with dementia were more often satisfied with information, had more often confidence in their ability to live well with dementia, and were less often satisfied with access to care compared to informal caregivers. Informal caregivers who were satisfied with support were more satisfied with information and access to care compared to informal caregivers not satisfied with support. CONCLUSIONS: Experience of dementia support can be improved and people with dementia and informal caregiver differ in their experiences of support

Prognostic value of the 6-gene OncoMasTR test in hormone receptor–positive HER2-negative early-stage breast cancer: Comparative analysis with standard clinicopathological factors

Aim: The aim of the study was to assess the prognostic performance of a 6-gene molecular score (OncoMasTR Molecular Score [OMm]) and a composite risk score (OncoMasTR Risk Score [OM]) and to conduct a within-patient comparison against four routinely used molecular and clinicopathological risk assessment tools: Oncotype DX Recurrence Score, Ki67, Nottingham Prognostic Index and Clinical Risk Category, based on the modified Adjuvant! Online definition and three risk factors: patient age, tumour size and grade. Methods: Biospecimens and clinicopathological information for 404 Irish women also previously enrolled in the Trial Assigning Individualized Options for Treatment [Rx] were provided by 11 participating hospitals, as the primary objective of an independent translational study. Gene expression measured via RT-qPCR was used to calculate OMm and OM. The prognostic value for distant recurrence-free survival (DRFS) and invasive disease-free survival (IDFS) was assessed using Cox proportional hazards models and Kaplan-Meier analysis. All statistical tests were two-sided ones. Results: OMm and OM (both with likelihood ratio statistic [LRS] P Discussion: Both OncoMasTR scores were significantly prognostic for DRFS and IDFS and provided additional prognostic information to the molecular and clinicopathological risk factors/tools assessed. OM was also the most accurate risk classification tool for identifying DR. A concise 6-gene signature with superior risk stratification was shown to increase prognosis reliability, which may help clinicians optimise treatment decisions. Trial registration: ClinicalTrials.gov NCT02050750 NCT00310180.</p

Are there Social Spillovers in Consumers’ Security Assessments of Payment Instruments?

Even though security of payments has long been identified as an important aspect of the consumer payment experience, recent literature fails to appropriately assess the extent of social spillovers among payment users. We test for the existence and importance of such spillovers by analyzing whether social influence affects consumers’ perceptions of the security of payment instruments. Based on a 2008–2014 annual panel data survey of consumers, we find strong evidence of social spillovers in payment markets: others’ perceptions of security of payment instruments exert a positive influence on one’s own payment security perceptions. The significant and robust results imply that a consumer’s assessments of security converge to his peers’ average assessment: a 10 percent change in the divergence between one’s own security rating and peers’ average rating will result in a 7 percent change in one’s own rating in the next period. The results are robust to many specifications and do not change when we control for actual fraud or crime data. Our results indicate that spillovers rather than reflection appear to be the cause, although separating the two causes is very difficult (Manski 1993). In particular, the spillovers are stronger for people who experience an exogenous shock to security perception, people who have more social interactions, and younger consumers, who are more likely to be influenced by social media. We also examine the effects of social spillovers on payment behavior (that is, on decisions regarding payment adoption and use). Our results indicate that social spillovers have a rather limited impact on payment behavior, as others’ perceptions seem to affect one’s own payment behavior mainly indirectly through the effect on one’s own perceptions

Erratum to: Calnexin, an ER stress-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer

Erratum to: Calnexin, an ER stress-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer