BORIS/CTCFL-mediated transcriptional regulation of the hTERT telomerase gene in testicular and ovarian tumor cells.

Telomerase activity, not detectable in somatic cells but frequently activated during carcinogenesis, confers immortality to tumors. Mechanisms governing expression of the catalytic subunit hTERT, the limiting factor for telomerase activity, still remain unclear. We previously proposed a model in which the binding of the transcription factor CTCF to the two first exons of hTERT results in transcriptional inhibition in normal cells. This inhibition is abrogated, however, by methylation of CTCF binding sites in 85% of tumors. Here, we showed that hTERT was unmethylated in testicular and ovarian tumors and in derivative cell lines. We demonstrated that CTCF and its paralogue, BORIS/CTCFL, were both present in the nucleus of the same cancer cells and bound to the first exon of hTERT in vivo. Moreover, exogenous BORIS expression in normal BORIS-negative cells was sufficient to activate hTERT transcription with an increasing number of cell passages. Thus, expression of BORIS was sufficient to allow hTERT transcription in normal cells and to counteract the inhibitory effect of CTCF in testicular and ovarian tumor cells. These results define an important contribution of BORIS to immortalization during tumorigenesis.

Preparation and properties of starch-based biopolymers modified with difunctional isocyanates

The present work reports on the preparation of thermoplastic starch (TPS) modified in situ with a diisocyanate derivative. Evidence of the condensation reaction between the hydroxyl groups of starch and glycerol with the isocyanate function (NCO) was confirmed by FTIR analysis. The evolution of the properties of the ensuing TPS, in term of mechanical properties, microstructure, and water sensitivity, was investigated using tensile mechanical, dynamic mechanical thermal analysis (DMTA), X-ray diffraction (XRD), and water uptake. The results showed that the addition of isocyanate did not affect the crystallinity of the TPS and slightly reduced the water uptake of the material. The evolution of the mechanical properties with ageing became less pronounced by the addition of the isocyanate as their amount exceeded 4 to 6wt%.

2q34-qter duplication and 4q34.2-qter deletion in a patient with developmental delay.

The 2q3 duplication and 4q3 deletion syndromes are two conditions with variable phenotypes including Pierre-Robin sequence (PRS), limb anomalies, congenital heart defects (CHD), developmental delays and intellectual disabilities. We describe a patient born to a mother with a balanced t(2; 4) translocation who combines both a 2q34-qter duplication and a 4q34.2-qter deletion through inheritance of the derivative chromosome 4 (der(4)). He showed developmental delay, growth retardation, hearing problems, minor facial and non-facial anomalies, such as bilateral fifth finger shortness and clinodactyly, but no PRS or CHD. The comparison of his features with those of 46 and 65 published cases of 2q3 duplication and 4q3 deletion, respectively, allows us to further restrict the size of the proposed critical intervals for PRS and CHD on chromosome 4.

Nuclear relaxation contribution to static and dynamic (infinite frequency approximation) nonlinear optical properties by means of electrical property expansions: Application to HF, CH4, CF4, and SF6

Electrical property derivative expressions are presented for the nuclear relaxation contribution to static and dynamic (infinite frequency approximation) nonlinear optical properties. For CF4 and SF6, as opposed to HF and CH4, a term that is quadratic in the vibrational anharmonicity (and not previously evaluated for any molecule) makes an important contribution to the static second vibrational hyperpolarizability of CF4 and SF6. A comparison between calculated and experimental values for the difference between the (anisotropic) Kerr effect and electric field induced second-harmonic generation shows that, at the Hartree-Fock level, the nuclear relaxation/infinite frequency approximation gives the correct trend (in the series CH4, CF4, SF6) but is of the order of 50% too small

Relations among several nuclear and electronic density functional reactivity indexes

A set of connections among several nuclear and electronic indexes of reactivity in the framework of the conceptual Density Functional Theory by using an expansion ofthe energy functional in terms of the total number of electrons and the normal coordinates within a canonical ensemble was derived. The relations obtained provided explicit links between important quantities related to the chemical reactivity of a system. This paper particularly demonstrates that the derivative of the electronic energy with respect to the external potential of a system in its equilibrium geometry was equal to the negative of the nuclear repulsion derivative with respect to the external potential

Binding of low concentration of peptide to H-2Kd produced in insect cells requires mouse beta 2-microglobulin co-expression.

A recombinant baculovirus expressing the murine class I MHC heavy chain H-2Kd cDNA under the transcriptional control of Autografa californica nuclear polyhedrosis virus (AcNPV) polyhedrin promoter has been isolated and used to infect Sf9 lepidopteran cells either alone or in association with a previously isolated virus expressing mouse beta 2-microglobulina (beta 2-ma). When infected with the heavy chain-encoding virus alone, H-2Kd was produced in a beta 2-m-free conformation detected on the surface of infected cells by conformation-independent antibodies. When Sf9 cells were co-infected with both viruses, approximately 10% of the heavy chain pool was engaged in the formation of native heterodimeric MHC class I molecules, which were glycosylated and transported to the cell surface as demonstrated by radio-binding experiments and flow cytometry. The assembly of the recombinant class I molecule was dependent on peptide, since heterodimer formation was brought about by H-2Kd-specific peptide ligands both in vivo, upon incubation with dually infected cells, and in vitro, in cell-free detergent extracts. In addition, a change in heavy chain conformation was brought about upon incubation with high concentrations (100 microM) of an H-2Kd-restricted octapeptide epitope from Plasmodium berghei. Furthermore, using low concentrations (3 nM) of a photoaffinity label derivative of this peptide, we show direct binding to cells co-expressing class I heavy chain and mouse beta 2-m but not to cells expressing free heavy chain only.

Targeted Gene Deletion and In Vivo Analysis of Putative Virulence Gene Function in the Pathogenic Dermatophyte Arthroderma benhamiae.

Dermatophytes cause the majority of superficial mycoses in humans and animals. However, little is known about the pathogenicity of this specialized group of filamentous fungi, for which molecular research has been limited thus far. During experimental infection of guinea pigs by the human pathogenic dermatophyte Arthroderma benhamiae, we recently detected the activation of the fungal gene encoding malate synthase AcuE, a key enzyme of the glyoxylate cycle. By the establishment of the first genetic system for A. benhamiae, specific ΔacuE mutants were constructed in a wild-type strain and, in addition, in a derivative in which we inactivated the nonhomologous end-joining pathway by deletion of the A. benhamiae KU70 gene. The absence of AbenKU70 resulted in an increased frequency of the targeted insertion of linear DNA by homologous recombination, without notably altering the monitored in vitro growth abilities of the fungus or its virulence in a guinea pig infection model. Phenotypic analyses of ΔacuE mutants and complemented strains depicted that malate synthase is required for the growth of A. benhamiae on lipids, major constituents of the skin. However, mutant analysis did not reveal a pathogenic role of the A. benhamiae enzyme in guinea pig dermatophytosis or during epidermal invasion of the fungus in an in vitro model of reconstituted human epidermis. The presented efficient system for targeted genetic manipulation in A. benhamiae, paired with the analyzed infection models, will advance the functional characterization of putative virulence determinants in medically important dermatophytes.

Comparative gene finding in chicken indicates that we are closing in on the set of multi-exonic widely-expressed human genes

The recent availability of the chicken genome sequence poses the question of whether there are human protein-coding genes conserved in chicken that are currently not included in the human gene catalog. Here, we show, using comparative gene finding followed by experimental verification of exon pairs by RT–PCR, that the addition to the multi-exonic subset of this catalog could be as little as 0.2%, suggesting that we may be closing in on the human gene set. Our protocol, however, has two shortcomings: (i) the bioinformatic screening of the predicted genes, applied to filter out false positives, cannot handle intronless genes; and (ii) the experimental verification could fail to identify expression at a specific developmental time. This highlights the importance of developing methods that could provide a reliable estimate of the number of these two types of genes.

Nematode selenoproteome: the use of the selenocysteine insertion system to decode one codon in an animal genome?

Selenocysteine (Sec) is co-translationally inserted into selenoproteins in response to codon UGA with the help of the selenocysteine insertion sequence (SECIS) element. The number of selenoproteins in animals varies, with humans having 25 and mice having 24 selenoproteins. To date, however, only one selenoprotein, thioredoxin reductase, has been detected in Caenorhabditis elegans, and this enzyme contains only one Sec. Here, we characterize the selenoproteomes of C.elegans and Caenorhabditis briggsae with three independent algorithms, one searching for pairs of homologous nematode SECIS elements, another searching for Cys- or Sec-containing homologs of potential nematode selenoprotein genes and the third identifying Sec-containing homologs of annotated nematode proteins. These methods suggest that thioredoxin reductase is the only Sec-containing protein in the C.elegans and C.briggsae genomes. In contrast, we identified additional selenoproteins in other nematodes. Assuming that Sec insertion mechanisms are conserved between nematodes and other eukaryotes, the data suggest that nematode selenoproteomes were reduced during evolution, and that in an extreme reduction case Sec insertion systems probably decode only a single UGA codon in C.elegans and C.briggsae genomes. In addition, all detected genes had a rare form of SECIS element containing a guanosine in place of a conserved adenosine present in most other SECIS structures, suggesting that in organisms with small selenoproteomes SECIS elements may change rapidly.

Feeding and the Equilibrium Feeder Animal Price-Weight Schedule, June 2005

The feeder animal price is a derivative in the sense that its value depends upon the price of animals for the consumption market. It also depends upon the biological growth technology and feed costs. Daily maintenance costs are of particular interest to the husbander because they can be avoided through accelerated feeding. In this paper, the optimal feeding path under equilibrium feeder animal prices is established. This analysis is used to gain a better understanding of feeding decisions, regulation in feedstuff markets, and the consequences of genetic innovations. It is shown that days on feed can increase or decrease with a genetic innovation or other improvement in feed conversion efficiency. The structure of comparative prices for feeder animals at different weights, the early slaughter decision, and equilibrium in feeder animal markets are also developed. Feeder animal prices can increase over a weight interval if biological feed efficiency parameters are low over the interval.

Photoaffinity labeling of the T cell receptor on cloned cytotoxic T lymphocytes by covalent photoreactive ligand.

The interaction of the T cell antigen receptor with a photoreactive antigenic peptide derivative bound covalently to the H-2Kd (Kd) molecule was studied by photoaffinity labeling on cloned, CD8 positive cytotoxic T lymphocytes. The Kd-restricted Plasmodium berghei circumsporozoite peptide 253-260 (YIPS-AEKI) was conjugated with iodo-4-azidosalicylic acid at the N terminus and with 4-azidobenzoic acid at the T cell receptor residue Lys-259. Cell-associated or soluble Kd molecules were photoaffinity-labeled with the peptide derivative by selective photoactivation of the N-terminal photoreactive group. Incubation of cell-associated or soluble covalent Kd-peptide derivative complexes (ligands) with cytotoxic T lymphocytes that recognized this peptide derivative and activation of the orthogonal photoreactive group resulted in specific photoaffinity labeling of the T cell receptor. The labeling was inhibitable by an anti-Kd antibody and was absent on Kd-restricted cytotoxic T lymphocytes of different specificity. The binding of the soluble ligand reached a maximum after 2-4 min at 37 degrees C, after 30 min at 18 degrees C, and after 3 h at 4 degrees C. In contrast, binding of the cell-associated ligand reached a transient maxima after 50 and 110 min at 37 and 18 degrees C, respectively. The degree of binding at 37 degrees C was approximately 30% lower than that at 18 degrees C. No binding took place at 4 degrees C. Inhibition studies with antibodies and drugs indicated that the binding of the cell-associated, but not the soluble ligand, was highly dependent on T cell-target cell conjugate formation, whereas the binding of the soluble ligand was greatly dependent on CD8.

The influence of learning on evolution: a mathematical framework.

The Baldwin effect can be observed if phenotypic learning influences the evolutionary fitness of individuals, which can in turn accelerate or decelerate evolutionary change. Evidence for both learning-induced acceleration and deceleration can be found in the literature. Although the results for both outcomes were supported by specific mathematical or simulation models, no general predictions have been achieved so far. Here we propose a general framework to predict whether evolution benefits from learning or not. It is formulated in terms of the gain function, which quantifies the proportional change of fitness due to learning depending on the genotype value. With an inductive proof we show that a positive gain-function derivative implies that learning accelerates evolution, and a negative one implies deceleration under the condition that the population is distributed on a monotonic part of the fitness landscape. We show that the gain-function framework explains the results of several specific simulation models. We also use the gain-function framework to shed some light on the results of a recent biological experiment with fruit flies.

Importance of genotypic and phenotypic tolerance in the treatment of experimental endocarditis due to Streptococcus gordonii.

Genotypic and phenotypic tolerance was studied in penicillin treatment of experimental endocarditis due to nontolerant and tolerant Streptococcus gordonii and to their backcross transformants. The organisms were matched for in vitro and in vivo growth rates. Rats with aortic endocarditis were treated for 3 or 5 days, starting 12, 24, or 48 h after inoculation. When started at 12 h, during fast intravegetation growth, 3 days of treatment cured 80% of the nontolerant parent compared with <30% of the tolerant derivative (P < .005). When started at 24 or 48 h and if intravegetation growth had reached a plateau, 3 days of treatment failed against both bacteria. However, a significant difference between the 2 organisms was restored when treatment was extended to 5 days. Thus, genotypic tolerance conferred a survival advantage in both fast- and slow-growing bacteria, demonstrating that the in vitro-defined tolerant phenotype also carried the risk of treatment failure in vivo.

Development of Implantable Flow-Through Left Ventricular Pressure Telemetric Transmitter for small Rodent Animals

Objective: Although 24-hour arterial blood pressure can be monitored in a free-moving animal using pressure telemetric transmitter mostly from Data Science International (DSI), accurate monitoring of 24-hour mouse left ventricular pressure (LVP) is not available because of its insufficient frequency response to a high frequency signal such as the maximum derivative of mouse LVP (LVdP/dtmax and LVdP/dtmin). The aim of the study was to develop a tiny implantable flow-through LVP telemetric transmitter for small rodent animals, which can be potentially adapted for human 24 hour BP and LVP accurate monitoring. Design and Method: The mouse LVP telemetric transmitter (Diameter: _12 mm, _0.4 g) was assembled by a pressure sensor, a passive RF telemetry chip, and to a 1.2F Polyurethane (PU) catheter tip. The device was developed in two configurations and compared with existing DSI system: (a) prototype-I: a new flow-through pressure sensor with wire link and (b) prototype-II: prototype-I plus a telemetry chip and its receiver. All the devices were applied in C57BL/6J mice. Data are mean_SEM. Results: A high frequency response (>100 Hz) PU heparin saline-filled catheter was inserted into mouse left ventricle via right carotid artery and implanted, LV systolic pressure (LVSP), LVdP/dtmax, and LVdP/dtmin were recorded on day2, 3, 4, 5, and 7 in conscious mice. The hemodynamic values were consistent and comparable (139_4 mmHg, 16634_319, - 12283_184 mmHg/s, n¼5) to one recorded by a validated Pebax03 catheter (138_2mmHg, 16045_443 and -12112_357 mmHg/s, n¼9). Similar LV hemodynamic values were obtained with Prototype-I. The same LVP waveforms were synchronically recorded by Notocord wire and Senimed wireless software through prototype-II in anesthetized mice. Conclusion: An implantable flow-through LVP transmitter (prototype-I) is generated for LVP accurate assessment in conscious mice. The prototype-II needs a further improvement on data transmission bandwidth and signal coupling distance to its receiver for accurate monitoring of LVP in a freemoving mouse.

Long-term follow-up of European APL 2000 trial, evaluating the role of cytarabine combined with ATRA and Daunorubicin in the treatment of nonelderly APL patients.

All-trans retinoic acid (ATRA) combined to anthracycline-based chemotherapy is the reference treatment of acute promyelocytic leukemia (APL). Whereas, in high-risk patients, cytarabine (AraC) is often considered useful in combination with anthracycline to prevent relapse, its usefulness in standard-risk APL is uncertain. In APL 2000 trial, patients with standard-risk APL [i.e., with baseline white blood cell (WBC) count <10,000/mm(3) ] were randomized between treatment with ATRA with Daunorubicin (DNR) and AraC (AraC group) and ATRA with DNR but without AraC (no AraC group). All patients subsequently received combined maintenance treatment. The trial had been prematurely terminated due to significantly more relapses in the no AraC group (J Clin Oncol, (24) 2006, 5703-10), but follow-up was still relatively short. With long-term follow-up (median 103 months), the 7-year cumulative incidence of relapses was 28.6% in the no AraC group, compared to 12.9% in the AraC group (P = 0.0065). In standard-risk APL, at least when the anthracycline used is DNR, avoiding AraC may lead to an increased risk of relapse suggesting that the need for AraC is regimen-dependent.