Regulation of Human Papillomavirus Type 16 Late Protein Expression During Epithelial Differentiation

Human papillomaviruses type 16 (HPV-16) is one of a number of HPV types specifically associated with the development of carcinoma in the anogenital tract and HPV-16 is the most commonly found subtype in cervical carcinomas worldwide. The early HPV proteins are expressed throughout epithelial differentiation, but expression of the late proteins is tightly linked to the differentiation status of the host cell, being found only in cells in the process of keratinisation in the granular layer of stratified epithelia. For HPV-16, a 79 nt negative regulatory element (NRE), overlapping the 3' end of the LI stop codon and extending into the late 3' untranslated region, is an important post-transcriptional regulator of viral late gene expression. Unsurprisingly, the HPV-16 NRE has previously been shown to bind a range of RNA processing factors in vitro, including the auxiliary splicing factor U2AF65, the polyadenylation factor CstF-64, and the elav-Wke HuR protein. I carried out saturating site-directed mutagenesis of the HPV-16 NRE and found that no individual short sequences are responsible for the activity of this inhibitory element, or its ability to interact with cellular factors. I have used an affinity-chromatography based RNA binding assay to show that the NRE can also bind the splicing-related (SR) protein, ASF/SF2, a member of the Sm protein family, and the U1A subunit of UlsnRNP. Furthermore, using a novel monolayer culture differentiation system and organotypic rafts to generate undifferentiated and differentiating populations of HPV-16 positive epithelial cell lines, I have shown by Western blot analysis that the level of expression of the NRE-binding proteins is differentially regulated, and is influenced by the status of the viral genome in the infected cell. I suggest that the inhibitory activity of the HPV-16 NRE is overcome as a result of the upregulation of cellular factors that bind the NRE

The alternative splicing factor hnRNP A1 is up-regulated during virus-infected epithelial cell differentiation and binds the human papillomavirus type 16 late regulatory element

AbstractHuman papillomavirus type 16 (HPV16) infects anogenital epithelia and is the etiological agent of cervical cancer. We showed previously that HPV16 infection regulates the key splicing/alternative splicing factor SF2/ASF and that virus late transcripts are extensively alternatively spliced. hnRNP A1 is the antagonistic counterpart of SF2/ASF in alternative splicing. We show here that hnRNP A1 is also up-regulated during differentiation of virus-infected epithelial cells in monolayer and organotypic raft culture. Taken together with our previous data on SF2/ASF, this comprises the first report of HPV-mediated regulation of expression of two functionally related cellular proteins during epithelial differentiation. Further, using electrophoretic mobility shift assays and UV crosslinking we demonstrate that hnRNP A1 binds the HPV16 late regulatory element (LRE) in differentiated HPV16 infected cells. The LRE has been shown to be important in temporally controlling virus late gene expression during epithelial differentiation. We suggest that increased levels of these cellular RNA processing factors facilitate appropriate alternative splicing necessary for production of virus late transcripts in differentiated epithelial cells

Qatar Islamic Archaeology and Heritage Project: End of Season Report : 2011-2012

International audienc

Progress Towards a Multi-Modal Capsule Endoscopy Device Featuring Microultrasound Imaging

Current clinical standards for endoscopy in the gastrointestinal (GI) tract combine high definition optics and ultrasound imaging to view the lumen superficially and through its thickness. However, these instruments are limited to the length of an endoscope and the only clinically available, autonomous devices able to travel the full length of the GI tract easily offer only video capsule endoscopy (VCE). Our work seeks to overcome this limitation with a device (“Sonopill”) for multimodal capsule endoscopy, providing optical and microultrasound (μUS) imaging and supporting sensors1. μUS transducers have been developed with multiple piezoelectric materials operating across a range of centre frequencies to study viability in the GI tract. Because of the combined constraints of μUS imaging and the low power / heat tolerance of autonomous devices, a hybrid approach has been taken to the transducer design, with separate transmit and receive arrays allowing multiple manufacturing approaches to maximise system efficiency. To explore these approaches fully, prototype devices have been developed with PVDF, high-frequency PZT and PMN-PT composites, and piezoelectric micromachined ultrasonic transducer arrays. Test capsules have been developed using 3D printing to investigate issues including power consumption, heat generation / dissipation, acoustic coupling, signal strength and capsule integrity. Because of the high functional density of the electronics in our proposed system, application specific integrated circuits (ASICs) have been developed to realise the ultrasound transmit and receive circuitry along with white-light and autofluorescence imaging with single-photon avalanche detectors (SPADs). The ultrasound ASIC has been developed and the SPAD electronics and optical subsystem have been validated experimentally. The functionality of various transducer materials has been examined as a function of frequency and ultrasound transducers have been developed to operate at centre frequencies in the range 15 - 50 MHz. Ex vivo testing of porcine tissue has been performed, generating images of interest to the clinical community, demonstrating the viability of the Sonopill concept

In-Vivo Evaluation of Microultrasound and Thermometric Capsule Endoscopes

Clinical endoscopy and colonoscopy are commonly used to investigate and diagnose disorders in the upper gastrointestinal tract and colon respectively. However, examination of the anatomically remote small bowel with conventional endoscopy is challenging. This and advances in miniaturization led to the development of video capsule endoscopy (VCE) to allow small bowel examination in a non-invasive manner. Available since 2001, current capsule endoscopes are limited to viewing the mucosal surface only due to their reliance on optical imaging. To overcome this limitation with submucosal imaging, work is under way to implement microultrasound (μUS) imaging in the same form as VCE devices. This paper describes two prototype capsules, termed Sonocap and Thermocap, which were developed respectively to assess the quality of μUS imaging and the maximum power consumption that can be tolerated for such a system. The capsules were tested in vivo in the oesophagus and small bowel of porcine models. Results are presented in the form of μUS B-scans and safe temperature readings observed up to 100 mW in both biological regions. These results demonstrate that acoustic coupling and μUS imaging can be achieved in vivo in the lumen of the bowel and the maximum power consumption that is possible for miniature μUS systems

Refractive-index sensing with ultra-thin plasmonic nanotubes

We study the refractive-index sensing properties of plasmonic nanotubes with a dielectric core and ultra-thin metal shell. The few-nm thin metal shell is described by both the usual Drude model and the nonlocal hydrodynamic model to investigate the effects of nonlocality. We derive an analytical expression for the extinction cross section and show how sensing of the refractive index of the surrounding medium and the figure-of-merit are affected by the shape and size of the nanotubes. Comparison with other localized surface plasmon resonance sensors reveals that the nanotube exhibits superior sensitivity and comparable figure-of-merit

Hydrogeomorphology of the Hyporheic Zone: Stream Solute and Fine Particle Interactions With a Dynamic Streambed

Hyporheic flow in streams has typically been studied separately from geomorphic processes. We investigated interactions between bed mobility and dynamic hyporheic storage of solutes and fine particles in a sand-bed stream before, during, and after a flood. A conservatively transported solute tracer (bromide) and a fine particles tracer (5 μm latex particles), a surrogate for fine particulate organic matter, were co-injected during base flow. The tracers were differentially stored, with fine particles penetrating more shallowly in hyporheic flow and retained more efficiently due to the high rate of particle filtration in bed sediment compared to solute. Tracer injections lasted 3.5 h after which we released a small flood from an upstream dam one hour later. Due to shallower storage in the bed, fine particles were rapidly entrained during the rising limb of the flood hydrograph. Rather than being flushed by the flood, we observed that solutes were stored longer due to expansion of hyporheic flow paths beneath the temporarily enlarged bedforms. Three important timescales determined the fate of solutes and fine particles: (1) flood duration, (2) relaxation time of flood-enlarged bedforms back to base flow dimensions, and (3) resulting adjustments and lag times of hyporheic flow. Recurrent transitions between these timescales explain why we observed a peak accumulation of natural particulate organic matter between 2 and 4 cm deep in the bed, i.e., below the scour layer of mobile bedforms but above the maximum depth of particle filtration in hyporheic flow paths. Thus, physical interactions between bed mobility and hyporheic transport influence how organic matter is stored in the bed and how long it is retained, which affects decomposition rate and metabolism of this southeastern Coastal Plain stream. In summary we found that dynamic interactions between hyporheic flow, bed mobility, and flow variation had strong but differential influences on base flow retention and flood mobilization of solutes and fine particulates. These hydrogeomorphic relationships have implications for microbial respiration of organic matter, carbon and nutrient cycling, and fate of contaminants in streams

Tapering Analysis of Airways with Bronchiectasis

Bronchiectasis is the permanent dilation of airways. Patients with the disease can suffer recurrent exacerbations, reducing their quality of life. The gold standard to diagnose and monitor bronchiectasis is accomplished by inspection of chest computed tomography (CT) scans. A clinician examines the broncho-arterial ratio to determine if an airway is brochiectatic. The visual analysis assumes the blood vessel diameter remains constant, although this assumption is disputed in the literature. We propose a simple measurement of tapering along the airways to diagnose and monitor bronchiectasis. To this end, we constructed a pipeline to measure the cross-sectional area along the airways at contiguous intervals, starting from the carina to the most distal point observable. Using a phantom with calibrated 3D printed structures, the precision and accuracy of our algorithm extends to the sub voxel level. The tapering measurement is robust to bifurcations along the airway and was applied to chest CT images acquired in clinical practice. The result is a statistical difference in tapering rate between airways with bronchiectasis and controls

Environmental effects on water intake and water intake prediction in growing beef cattle

Water is an essential nutrient, but there are few recent studies that evaluate how much water individual beef cattle consume and how environmental factors affect an individual’s water intake (WI). Most studies have focused on WI of whole pens rather than WI of individual animals. Thus, the objective of this study was to evaluate the impact of environmental parameters on individual-animal WI across different seasons and develop prediction equations to estimate WI, including within different environments and management protocols. Individual daily feed intake and WI records were collected on 579 crossbred steers for a 70-d period following a 21-d acclimation period for feed and water bunk training. Steers were fed in 5 separate groups over a 3-yr period from May 2014 to March 2017. Individual weights were collected every 14 d and weather data were retrieved from the Oklahoma Mesonet’s Stillwater station. Differences in WI as a percent of body weight (WI%) were analyzed accounting for average temperature (TAVG), relative humidity (HAVG), solar radiation (SRAD), and wind speed (WSPD). Seasonal (summer vs. winter) and management differences (ad libitum vs. slick bunk) were examined. Regression analysis was utilized to generate 5 WI prediction equations (overall, summer, winter, slick, and ad libitum). There were significant (P \u3c 0.05) differences in WI between all groups when no environmental parameters were included in the model. Although performance was more similar after accounting for all differences in weather variables, significant (P \u3c 0.05) seasonal and feed management differences were still observed for WI%, but were less than 0.75% of steer body weight. The best linear predictors of daily WI (DWI) were dry mater intake (DMI), metabolic body weights (MWTS), TAVG, SRAD, HAVG, and WSPD. Slight differences in the coefficient of determinations for the various models were observed for the summer (0.34), winter (0.39), ad libitum (0.385), slick bunk (0.41), and overall models (0.40). Based on the moderate R2 values for the WI prediction equations, individual DWI can be predicted with reasonable accuracy based on the environmental conditions that are present, MWTS, and DMI consumed, but substantial variation exists in individual animal WI that is not accounted for by these models