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Epithelial to mesenchymal transition (EMT) is a dynamic process that drives cancer cell plasticity and is thought to play a major role in metastasis. Here we show, using MDA-MB-231 cells as a model, that the plasticity of at least some metastatic breast cancer cells is dependent on the transcriptional co-regulator CBFβ. We demonstrate that CBFβ is essential to maintain the mesenchymal phenotype of triple-negative breast cancer cells and that CBFβ-depleted cells undergo a mesenchymal to epithelial transition (MET) and re-organise into acini-like structures, reminiscent of those formed by epithelial breast cells. We subsequently show, using an inducible CBFβ system, that the MET can be reversed, thus demonstrating the plasticity of CBFβ-mediated EMT. Moreover, the MET can be reversed by expression of the EMT transcription factor Slug whose expression is dependent on CBFβ. Finally, we demonstrate that loss of CBFβ inhibits the ability of metastatic breast cancer cells to invade bone cell cultures and suppresses their ability to form bone metastases in vivo. Together our findings demonstrate that CBFβ can determine the plasticity of the metastatic cancer cell phenotype, suggesting that its regulation in different micro-environments may play a key role in the establishment of metastatic tumours.

Small size ultra-precision Computer Numerical Control (CNC) machines require high dynamic performance. Flexible frame phenomena can limit the machine dynamic performance, particularly in small size machines. A novel accelerometer based feedback concept for improving machine dynamic performance was developed and realised, a virtual metrology frame. It expends the limited techniques for improving dynamic performance of a small size machine by measuring the flexible frame displacement, and feeding it into the controller. The concept was implemented in a simplified linear motion system, and showed a 12dB reduction in the magnitude of the first resonance in the plant frequency response function. This allowed improving the servo bandwidth by 58% based on a PID controller. A new technique for real-time dynamic displacement measurements using accelerometer was developed. It shows a low sensor noise σ<30 nm; thus, accelerometers are used as a displacement sensor in a control system.

Despite a paucity of supporting literature, acetazolamide is commonly used in critically ill children with metabolic alkalosis (elevated plasma bicarbonate pHco–3 and pH). The objective of this study was to assess the change in 18 hours after initiation of acetazolamide therapy.

The Wilms' tumor 1 protein WT1 is a transcriptional regulator that is involved in cell growth and differentiation. The transcriptional corepressor BASP1 interacts with WT1 and converts WT1 from a transcriptional activator to a repressor. Here, we demonstrate that the N-terminal myristoylation of BASP1 is required in order to elicit transcriptional repression at WT1 target genes. We show that myristoylated BASP1 binds to nuclear PIP2, which leads to the recruitment of PIP2 to the promoter regions of WT1-dependent target genes. BASP1's myristoylation and association with PIP2 are required for the interaction of BASP1 with HDAC1, which mediates the recruitment of HDAC1 to the promoter and elicits transcriptional repression. Our findings uncover a role for myristoylation in transcription, as well as a critical function for PIP2 in gene-specific transcriptional repression through the recruitment of histone deacetylase.

To generate a preliminary bedside predictor of rapid time-to-death after withdrawal of support in children to help identify potential candidates for organ donation after circulatory death.

Nanometre-scale figuring technique at atmospheric pressure for large optical surfaces is a high profile research topic which attracts numerous competing state-of-the-art technologies. In this context, a dry chemical process, called Reactive Atom Plasma (RAP), was developed as a prospectively ideal alternative to CNC polishing or Ion Beam Figuring. The RAP process combines high material removal rates, nanometre level repeatability and absence of subsurface damage. A RAP figuring facility with metre-scale processing capability, Helios 1200, was then established in the Precision Engineering Centre at Cranfield University. The work presented in this paper is carried out using Helios 1200 and demonstrates the rapid figuring capability of the RAP process. First experimental tests of figure correction are performed on fused silica substrates over 100 mm diameter areas. A 500 nm deep spherical hollow shape is etched onto the central region of 200x200 mm polished surfaces. The test is carried out twice for reproducibility purposes. After two iterative steps, a residual figure error of ~16 nm rms is achieved. Subsequently, the process is scaled up to 140 mm diameter areas and two tests are carried out. First, the developed algorithm for 500 nm deep spherical hollow test is confirmed. Residual deviation over processed area is ~18 nm rms after three iterations. Finally, a surface characterised by random topography (79 nm rms initial figure error) is smoothed down to ~ 16 nm rms within three iteration steps. All results presented in this paper are achieved by means of an in-house developed tool-path algorithm. This can be described as a staggered meander-type tool motion path specifically designed to reduce heat transfer and consequently temperature gradient on the surface. Contiguously, classical de-convolution methods are adapted to non-linear etching rates for the derivation of the surface scanning speed maps. The figuring procedure is carried out iteratively. It is noteworthy that iteration steps never exceed ~7 minutes mean processing time.

Throughout the world there has been concern that persons with disabilities are being perceived as "other" than us and that differences imparted by disability result in more dissimilarity than actually exists. A summary of this concept of "otherness" and disability in the United States, Eastern Europe, and India is presented. [ABSTRACT FROM AUTHOR]

Drainage of cerebrospinal fluid (CSF) is routinely used in the treatment of severe traumatic brain injury (TBI), either continuously or intermittently in response to increases in intracranial pressure (ICP). There has been little study of the effect of CSF drainage method on the biochemistry, pathophysiology or outcome of TBI in adults or children. Having previously reported that a variety of markers of injury or repair increase in CSF after severe TBI, we chose to evaluate directly the effect of CSF drainage method on the biochemistry and volume of CSF drained as well as ICP. We hypothesized that concentrations of these markers would be similar in CSF drained continuously vs intermittently. We compared CSF levels of markers of neuronal injury (neuron specific enolase, [NSE]), glial injury (s100B), inflammation (interleukin-6 [IL-6]), and regeneration (vascular endothelial growth factor [VEGF]) (measured by ELISA) in 80 CSF samples from 19 severely injured children whose CSF was drained continuously (n = 13) versus intermittently (n = 6) as part of standard care in two institutions. Compared to continuous CSF drainage, intermittent drainage of CSF was associated with twofold greater CSF concentrations of NSE, s100B, IL-6 and VEGF (p < 0.05) and with about half the volume of CSF removal than continuous drainage (p = 0.002). The resulting elimination (concentration x volume) of these biochemicals, however, was not influenced by drainage method. Patients treated with continuous drainage had lower mean ICPs than those with intermittent drainage (13.6 ± 0.69 vs. 21.8 ± 0.95 mm Hg, p < 0.0001). We conclude that the method of CSF drainage greatly affects concentrations of CSF markers after TBI and may influence ICP. The influence of method on CSF marker concentration must be kept in mind when interpreting

The ETS domain family of transcription factors is comprised of several important proteins that are involved in controlling key cellular events such as proliferation, differentiation, and development. One such protein, Elk-1, regulates the activity of the c-fospromoter in response to extracellular stimuli. Elk-1 is representative of a subgroup of ETS domain proteins that utilize a bipartite recognition mechanism that is mediated by both protein-DNA and protein-protein interactions. In this study, we have overexpressed, purified, and characterized the ETS DNA-binding domain of Elk-1 (Elk-93). Elk-93 was expressed in Escherichia colias a fusion protein with glutathione S-transferase and purified to homogeneity from both the soluble and insoluble fractions using a two-column protocol. A combination of CD, NMR, and fluorescence spectroscopy demonstrates that Elk-93 represents an independently folded domain of mixed α/β structure in which the three conserved tryptophans appear to contribute to the hydrophobic core of the protein. Moreover, DNA binding studies demonstrate that Elk-93 binds DNA with both high affinity (Kd≈ 0.85 × 10−10M) and specificity. Circular permutation analysis indicates that DNA binding by Elk-93 does not induce significant bending of the DNA. Our results are discussed with respect to predictive models for the structure of the ETS DNA-binding domain.

Enzyme-catalysed DNA methylation provides an opportunity for the modulation of protein-DNA recognition in biological systems. Recently we have demonstrated that the smaller of the two subunits of the heterodimeric, cytosine-specific DNA methyltransferase, M. AquI, is largely responsible for sequence-specific DNA recognition. Here we present evidence from a series of NMR, fluorescence and circular dichroism spectroscopy experiments that the DNA binding subunit of M. AquI has the characteristics of a molten globule in the absence of the catalytic machinery. In this metastable state this subunit retains its ability to bind DNA in a sequence-specific manner. We believe this finding offers an insight into the structural flexibility which underpins the mechansim of action of these enzymes, and may provide a possible biological role for molten globules in protein function.

Enzyme‐catalysed DNA methylation provides an opportunity for the modulation of protein‐DNA recognition in biological systems. Recently we have demonstrated that the smaller of the two subunits of the heterodimeric, cytosine‐specific DNA methyltransferase, M.AquI, is largely responsible for sequence‐specific DNA recognition. Here we present evidence from a series of NMR, fluorescence and circular dichroism spectroscopy experiments that the DNA binding subunit of M.AquI has the characteristics of a molten globule in the absence of the catalytic machinery. In this metastable state this subunit retains its ability to bind DNA in a sequence‐specific manner. We believe this finding offers an insight into the structural flexibility which underpins the mechansim of action of these enzymes, and may provide a possible biological role for molten globules in protein function.

The serum response factor (SRF) and myocyte enhancer factor 2A (MEF2A) represent two human members of the MADS-box transcription factor family. Each protein has a distinct biological function which is reflected by the distinct specificities of the proteins for coregulatory protein partners and DNA-binding sites. In this study, we have investigated the mechanism of DNA binding utilized by these two related transcription factors. Although SRF and MEF2A belong to the same family and contain related DNA-binding domains, their DNA-binding mechanisms differ in several key aspects. In contrast to the dramatic DNA bending induced by SRF, MEF2A induces minimal DNA distortion. A combination of loss- and gain-of-function mutagenesis identified a single amino acid residue located at the N terminus of the recognition helices as the critical mediator of this differential DNA bending. This residue is also involved in determining DNA-binding specificity, thus indicating a link between DNA bending and DNA-binding specificity determination. Furthermore, different basic residues within the putative recognition α-helices are critical for DNA binding, and the role of the C-terminal extensions to the MADS box in dimerization between SRF and MEF2A also differs. These important differences in the molecular interactions of SRF and MEF2A are likely to contribute to their differing roles in the regulation of specific gene transcription.

In previous studies, lower levels of serum total protein, serum albumin and plasma ascorbic acid were found in patients with chronic peptic ulcer than in comparable persons without clinical evidence of gastrointestinal disorder.1 Since modification of the chemical structure of blood plasma influences the clinical course of ulcer through its action on motor and secretory functions of the stomach as well as by its effect on healing of the actual ulcer, it seemed of value to determine whether these changes in the blood were an integral part of the disease complex or reflected a deficiency in the diet.This study presents an analysis of the dietary records of 16 of the group whose blood studies have been reported. These patients, while considered representative of the group as a whole, were selected primarily because of their willingness to cooperate. Their average age was 45, the youngest 21 and the oldest

Radial and Linear Fresnel Lenses are finding application as light concentrators for Concentrated Photovoltaic and Concentrated Solar Thermal power applications. The efficiency of these diffractive lenses directly affects the yield of such systems. Peaks and valleys of the optical facets of the Fresnel lens must be sharp in order to prevent diffusion and transmission loss due to rounding. For diamond turned mould masters, optical facet tip sharpness is affected by machining accuracy, tool-path and tool wear/mileage. Strategies to optimise optical facet tip sharpness are presented which enable production of large lenses with minimal degradation of optical quality. Radial Fresnel produced with diameters over 500mm and Linear Fresnel over 1m long are discussed with data on structure fidelity and tool wear.

The ETS DNA-binding domain is conserved amongst many eukaryotic transcription factors. ETS-domains bind differentially to specific ONA sites containing a central GGA trinucleotide motif. The nucleotides flank- Ing this motif define the binding specificity of Individual proteins. In this study we have investigated binding specificity of the ETS-domains from two members of the ternary complex factor (TCF) subfamily, Elk-1 and SAP-1. The ETS DNA-binding domains of Elk-1 (Elk-93) and SAP-1 (SAP-92) select similar sites from random pools of double stranded oligonucleotides based on the consensus sequence ACCGGAAGTR. However, SAP-92 shows a more relaxed binding site selectivity and binds efficiently to a greater spectrum of sites than does Elk-93. This more relaxed DNA binding site selectivity Is most pronounced in nucleotides located on the 3′ side of the GGA motif. This differential DNA-blnding specificity is also exhibited by longer TCF derivatives and, indeed by the full-length proteins. Our results suggest that the range of potential In vivo target sites for SAP-1 is likely to be greater than for Elk-1. We discuss our results in relation to other similar studies carried out with more divergent ETS-domains.

Transcriptional induction of the c-fos protc-oncogene in response to serum growth factors is mediated in part by a ternary complex that forms on the serum response element (SRE) within Its promoter. This complex consists of Elk-1, serum response factor (SRF) and the SRE. Elk-1 Is phosphorylated by MAP kinase, which correlates with the induction of c-fos transcription. In this study we have investigated the protein-induced DNA bending which occurs during the formation and post-translational modification of the ternary complex that forms at the c-fos SRE. Circular permutation analysis demonstrates that the minimal DNA-blnding domain of SRF, which contains the MADS box, is sufficient to Induce flexibility into the centre of its binding site within the SRE. Phasing analysis indicates that at least part of this flexibility results in the production of a directional bend towards the minor groove. The Isolated ETS domains from Elk-1 and SAP-1 induce neither DNA bending nor increased DNA flexibility. Formation of ternary complexes by binding of Elk-1 to the binary SRF:SRE complex results In a change in the flexibility of the SRE. Phosphorylatlon of Elk-1 by MAP kinase (p42/ERK2) induces urther minor changes In this DNA flexiility. However, phasinganalysis reveals that the recruitment of Elk-1 to form the ternary complex affects the SRF-induced directional DNA bend in the SRE. The potetial roles of DNA bending at the c-fos SR are discussed.