aCGH.Spline--an R package for aCGH dye bias normalization.

MOTIVATION: The careful normalization of array-based comparative genomic hybridization (aCGH) data is of critical importance for the accurate detection of copy number changes. The difference in labelling affinity between the two fluorophores used in aCGH-usually Cy5 and Cy3-can be observed as a bias within the intensity distributions. If left unchecked, this bias is likely to skew data interpretation during downstream analysis and lead to an increased number of false discoveries. RESULTS: In this study, we have developed aCGH.Spline, a natural cubic spline interpolation method followed by linear interpolation of outlier values, which is able to remove a large portion of the dye bias from large aCGH datasets in a quick and efficient manner. CONCLUSIONS: We have shown that removing this bias and reducing the experimental noise has a strong positive impact on the ability to detect accurately both copy number variation (CNV) and copy number alterations (CNA).

Passive control of quorum sensing: prevention of Pseudomonas aeruginosa biofilm formation by imprinted polymers.

Here we present the first molecular imprinted polymer (MIP) that is able to attenuate the biofilm formation of the opportunistic human pathogen Pseudomonas aeruginosa through specific sequestration of its signal molecule N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C(12)-AHL). The MIP was rationally designed using computational modeling, and its capacity and specificity and that of a corresponding blank polymer toward signal molecule of P. aeruginosa (3-oxo-C(12)-AHL) and its analogue were tested. The biofilm formation in the presence of polymers and without polymers was studied using scanning confocal laser microscopy. Staining with crystal violet dye was used for the quantification of the biofilm formation. A significant reduction of the biofilm growth was observed in the presence of MIP (>80%), which was superior to that of the resin prepared without template, which showed a reduction of 40% in comparison with biofilm, which was grown without polymer addition. It was shown that 3-oxo-C(12)-AHL-specific MIP prevented the development of quorum-sensing-controlled phenotypes (in this case, biofilm formation) from being up-regulated. The developed MIP could be considered as a new tool for the elimination of life-threatening infections in a multitude of practical applications; it could, for example, be grafted on the surface of medical devices such as catheters and lenses, be a component of paints, or be used as a wound adsorbent.

CancerEST: a web-based tool for automatic meta-analysis of public EST data.

The identification of cancer-restricted biomarkers is fundamental to the development of novel cancer therapies and diagnostic tools. The construction of comprehensive profiles to define tissue- and cancer-specific gene expression has been central to this. To this end, the exploitation of the current wealth of 'omic'-scale databases can be facilitated by automated approaches, allowing researchers to directly address specific biological questions. Here we present CancerEST, a user-friendly and intuitive web-based tool for the automated identification of candidate cancer markers/targets, for examining tissue specificity as well as for integrated expression profiling. CancerEST operates by means of constructing and meta-analyzing expressed sequence tag (EST) profiles of user-supplied gene sets across an EST database supporting 36 tissue types. Using a validation data set from the literature, we show the functionality and utility of CancerEST. DATABASE URL: http://www.cancerest.org.uk.

Identification of a class of human cancer germline genes with transcriptional silencing refractory to the hypomethylating drug 5-aza-2'-deoxycytidine.

Bona fide germline genes have expression restricted to the germ cells of the gonads. Testis-specific germline development-associated genes can become activated in cancer cells and can potentially drive the oncogenic process and serve as therapeutic/biomarker targets; such germline genes are referred to as cancer/testis genes. Many cancer/testis genes are silenced via hypermethylation of CpG islands in their associated transcriptional control regions and become activated upon treatment with DNA hypomethylating agents; such hypomethylation-induced activation of cancer/testis genes provides a potential combination approach to augment immunotherapeutics. Thus, understanding cancer/testis gene regulation is of increasing clinical importance. Previously studied cancer/testis gene activation has focused on X chromosome encoded cancer/testis genes. Here we find that a sub-set of non-X encoded cancer/testis genes are silenced in non-germline cells via a mechanism that is refractory to epigenetic dysregulation, including treatment with the hypomethylating agent 5-aza-2'-deoxycytidine and the histone deacetylase inhibitor tricostatin A. These findings formally indicate that there is a sub-group of the clinically important cancer/testis genes that are unlikely to be activated in clinical therapeutic approaches using hypomethylating agents and it indicates a unique transcriptional silencing mechanism for germline genes in non-germline cells that might provide a target mechanism for new clinical therapies.

Detection and imaging the expression of the trans-membrane protein CD44 in RT112 cells by use of enzyme-labeled antibodies and SECM.

The deposition of human RT112 cells in a patterned fashion onto glass substrates and subsequent imaging of the expression of the trans-membrane protein CD44 have been studied using scanning electrochemical microscopy (SECM). Patterns of RT112 cells derived from a transitional cell carcinoma of the bladder could be deposited on amino-modified glass substrates by cytospinning. These were then treated with horseradish peroxidase (HRP) labeled secondary antibodies to the trans-membrane protein CD44. Expression of CD44 protein by the cells directly leads to immobilisation of the labeled antibodies. The presence of the enzyme substrate (hydrogen peroxide) along with a hydroquinone mediator then allowed an enzymatic reaction to proceed, generating benzoquinone. Reduction of benzoquinone gave rise to positive feedback between the substrate and the SECM microelectrode tip. Control samples such as blank slides or slides not treated with HRP-labeled antibody showed negative feedback effects. Patterns of RT112 cells could be assembled and their expression of the target protein imaged whereas control samples showed minimal activity.

CancerMA: a web-based tool for automatic meta-analysis of public cancer microarray data.

The identification of novel candidate markers is a key challenge in the development of cancer therapies. This can be facilitated by putting accessible and automated approaches analysing the current wealth of 'omic'-scale data in the hands of researchers who are directly addressing biological questions. Data integration techniques and standardized, automated, high-throughput analyses are needed to manage the data available as well as to help narrow down the excessive number of target gene possibilities presented by modern databases and system-level resources. Here we present CancerMA, an online, integrated bioinformatic pipeline for automated identification of novel candidate cancer markers/targets; it operates by means of meta-analysing expression profiles of user-defined sets of biologically significant and related genes across a manually curated database of 80 publicly available cancer microarray datasets covering 13 cancer types. A simple-to-use web interface allows bioinformaticians and non-bioinformaticians alike to initiate new analyses as well as to view and retrieve the meta-analysis results. The functionality of CancerMA is shown by means of two validation datasets.

An in silico study of the differential effect of oxidation on two biologically relevant G-quadruplexes: possible implications in oncogene expression.

G-quadruplex structures, formed from guanine rich sequences, have previously been shown to be involved in various physiological processes including cancer-related gene expression. Furthermore, G-quadruplexes have been found in several oncogene promoter regions, and have been shown to play a role in the regulation of gene expression. The mutagenic properties of oxidative stress on DNA have been widely studied, as has the association with carcinogenesis. Guanine is the most susceptible nucleotide to oxidation, and as such, G-rich sequences that form G-quadruplexes can be viewed as potential "hot-spots" for DNA oxidation. We propose that oxidation may destabilise the G-quadruplex structure, leading to its unfolding into the duplex structure, affecting gene expression. This would imply a possible mechanism by which oxidation may impact on oncogene expression. This work investigates the effect of oxidation on two biologically relevant G-quadruplex structures through 500 ns molecular dynamics simulations on those found in the promoter regions of the c-Kit and c-Myc oncogenes. The results show oxidation having a detrimental effect on stability of the structure, substantially destabilising the c-Kit quadruplex, and with a more attenuated effect on the c-Myc quadruplex. Results are suggestive of a novel route for oxidation-mediated oncogenesis and may have wider implications for genome stability.

Temporal and spatial changes in the microbial bioaerosol communities in green-waste composting.

In this study, the microbial community within compost, emitted into the airstream, downwind and upwind from a composting facility was characterized and compared through phospholipid fatty acid analysis and 16S rRNA gene analysis using denaturing gradient gel electrophoresis and bar-coded pyrosequencing techniques. All methods used suggested that green-waste composting had a significant impact upon bioaerosol community composition. Daily variations of the on-site airborne community showed how specific site parameters such as compost process activity and meteorological conditions affect bioaerosol communities, although more data are required to qualify and quantify the causes for these variations. A notable feature was the dominance of Pseudomonas in downwind samples, suggesting that this genus can disperse downwind in elevated abundances. Thirty-nine phylotypes were homologous to plant or human phylotypes containing pathogens and were found within compost, on-site and downwind microbial communities. Although the significance of this finding in terms of potential health impact was beyond the scope of this study, it clearly illustrated the potential of molecular techniques to improve our understanding of the impact that green-waste composting emissions may have on the human health.