Nuclear translocation and DNA recognition signals colocalized within the bZIP domain of cyclic adenosine 3',5'-monophosphate response element-binding protein CREB.

CREB is a cAMP-responsive nuclear DNA-binding protein that binds to cAMP response elements and stimulates gene transcription upon activation of the cAMP signalling pathway. The protein consists of an amino-terminal transcriptional transactivation domain and a carboxyl-terminal DNA-binding domain (bZIP domain) comprised of a basic region and a leucine zipper involved in DNA recognition and dimerization, respectively. Recently, we discovered a testis-specific transcript of CREB that contains an alternatively spliced exon encoding multiple stop codons. CREB encoded by this transcript is a truncated protein lacking the bZIP domain. We postulated that the antigen detected by CREB antiserum in the cytoplasm of germinal cells is the truncated CREB that must also lack its nuclear translocation signal (NTS). To test this hypothesis we prepared multiple expression plasmids encoding carboxyl-terminal deletions of CREB and transiently expressed them in COS-1 cells. By Western immunoblot analysis as well as immunocytochemistry of transfected cells, we show that CREB proteins truncated to amino acid 286 or shorter are sequestered in the cytoplasm, whereas a CREB of 295 amino acids is translocated into the nucleus. Chimeric CREBs containing a heterologous NTS fused to the first 248 or 261 amino acids of CREB are able to drive the translocation of the protein into the nucleus. Thus, the nine amino acids in the basic region involved in DNA recognition between positions 287 and 295 (RRKKKEYVK) of CREB contain the NTS. Further, mutation of the lysine at position 290 in CREB to an asparagine diminishes nuclear translocation of the protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Trace element chemistry and U-Pb dating of zircons from oceanic gabbros and their relationship with whole rock composition (Lanzo, Italian Alps)

The U-Pb ages and the trace element content of zircon U-Pb along with major and trace element whole rock data on gabbroic dikes from the Lanzo lherzolitic massif, N-Italy, have been determined to constrain crustal accretion in ocean-continent transition zones. Three Fe-Ti gabbros were dated from the central and the southern part of the massif providing middle Jurassic ages of 161 +/- 2, 158 +/- 2 and 163 +/- 1 Ma, which argue for magmatic activity over few millions of years. Zircon crystals are characterized by high but variable Th/U ratios, rare earth element patterns enriched in heavy rare earths, pronounced positive Ce and negative Eu-anomalies consistent with crystallization after substantial plagioclase fractionation. The zircon trace element composition coupled with whole rock chemistry was used to reconstruct the crystallization history of the gabbros. A number of gabbros crystallized in situ, and zircon precipitated from trapped, intercumulus liquid, while other gabbros represent residual liquids that were extracted from a cumulus pile and crystallized along syn-magmatic shear zones. We propose a model in which the emplacement mechanism of gabbroic rocks in ocean-continent transition zones evolves from in situ crystallization to stratified crystallization with efficient extraction of residual liquid along syn-magmatic shear zones. Such an evolution of the crystallization history is probably related to the thermal evolution of the underlying mantle lithosphere.

Influence of the implanted pulse generator as reference electrode in finite element model of monopolar deep brain stimulation.

Electrical deep brain stimulation (DBS) is an efficient method to treat movement disorders. Many models of DBS, based mostly on finite elements, have recently been proposed to better understand the interaction between the electrical stimulation and the brain tissues. In monopolar DBS, clinically widely used, the implanted pulse generator (IPG) is used as reference electrode (RE). In this paper, the influence of the RE model of monopolar DBS is investigated. For that purpose, a finite element model of the full electric loop including the head, the neck and the superior chest is used. Head, neck and superior chest are made of simple structures such as parallelepipeds and cylinders. The tissues surrounding the electrode are accurately modelled from data provided by the diffusion tensor magnetic resonance imaging (DT-MRI). Three different configurations of RE are compared with a commonly used model of reduced size. The electrical impedance seen by the DBS system and the potential distribution are computed for each model. Moreover, axons are modelled to compute the area of tissue activated by stimulation. Results show that these indicators are influenced by the surface and position of the RE. The use of a RE model corresponding to the implanted device rather than the usually simplified model leads to an increase of the system impedance (+48%) and a reduction of the area of activated tissue (-15%).

The cAMP response element binding protein-2 (CREB-2) can interact with the C/EBP-homologous protein (CHOP).

cAMP response element binding protein-2 (CREB-2) is a basic leucine zipper (bZIP) factor that was originally described as a repressor of CRE-dependent transcription but that can also act as a transcriptional activator. Moreover, CREB-2 is able to function in association with the viral Tax protein as an activator of the human T-cell leukemia virus type I (HTLV-I) promoter. Here we show that CREB-2 is able to interact with C/EBP-homologous protein (CHOP), a bZIP transcription factor known to inhibit CAAT/enhancer-dependent transcription. Cotransfection of CHOP with CREB-2 results in decreased activation driven by the cellular CRE motif or the HTLV-I proximal Tax-responsive element, confirming that CREB-2 and CHOP can interact with each other in vivo.

The Arabidopsis PHYTOCHROME KINASE SUBSTRATE2 protein is a phototropin signaling element that regulates leaf flattening and leaf positioning.

In Arabidopsis (Arabidopsis thaliana), the blue light photoreceptor phototropins (phot1 and phot2) fine-tune the photosynthetic status of the plant by controlling several important adaptive processes in response to environmental light variations. These processes include stem and petiole phototropism (leaf positioning), leaf flattening, stomatal opening, and chloroplast movements. The PHYTOCHROME KINASE SUBSTRATE (PKS) protein family comprises four members in Arabidopsis (PKS1-PKS4). PKS1 is a novel phot1 signaling element during phototropism, as it interacts with phot1 and the important signaling element NONPHOTOTROPIC HYPOCOTYL3 (NPH3) and is required for normal phot1-mediated phototropism. In this study, we have analyzed more globally the role of three PKS members (PKS1, PKS2, and PKS4). Systematic analysis of mutants reveals that PKS2 (and to a lesser extent PKS1) act in the same subset of phototropin-controlled responses as NPH3, namely leaf flattening and positioning. PKS1, PKS2, and NPH3 coimmunoprecipitate with both phot1-green fluorescent protein and phot2-green fluorescent protein in leaf extracts. Genetic experiments position PKS2 within phot1 and phot2 pathways controlling leaf positioning and leaf flattening, respectively. NPH3 can act in both phot1 and phot2 pathways, and synergistic interactions observed between pks2 and nph3 mutants suggest complementary roles of PKS2 and NPH3 during phototropin signaling. Finally, several observations further suggest that PKS2 may regulate leaf flattening and positioning by controlling auxin homeostasis. Together with previous findings, our results indicate that the PKS proteins represent an important family of phototropin signaling proteins.

An operon of three transcriptional regulators controls horizontal gene transfer of the integrative and conjugative element ICEclc in Pseudomonas knackmussii B13.

The integrative and conjugative element ICEclc is a mobile genetic element in Pseudomonas knackmussii B13, and an experimental model for a widely distributed group of elements in Proteobacteria. ICEclc is transferred from specialized transfer competent cells, which arise at a frequency of 3-5% in a population at stationary phase. Very little is known about the different factors that control the transfer frequency of this ICE family. Here we report the discovery of a three-gene operon encoded by ICEclc, which exerts global control on transfer initiation. The operon consists of three consecutive regulatory genes, encoding a TetR-type repressor MfsR, a MarR-type regulator and a LysR-type activator TciR. We show that MfsR autoregulates expression of the operon, whereas TciR is a global activator of ICEclc gene expression, but no clear role was yet found for MarR. Deletion of mfsR increases expression of tciR and marR, causing the proportion of transfer competent cells to reach almost 100% and transfer frequencies to approach 1 per donor. mfsR deletion also caused a two orders of magnitude loss in population viability, individual cell growth arrest and loss of ICEclc. This indicates that autoregulation is an important feature maintaining ICE transfer but avoiding fitness loss. Bioinformatic analysis showed that the mfsR-marR-tciR operon is unique for ICEclc and a few highly related ICE, whereas tciR orthologues occur more widely in a large variety of suspected ICE among Proteobacteria.

Trace element monitoring in the ICU: quality and economic impact of a change in sampling practice.

BACKGROUND & AIMS: Trace elements (TE) are involved in the immune and antioxidant defences which are of particular importance during critical illness. Determining plasma TE levels is costly. The present quality control study aimed at assessing the economic impact of a computer reminded blood sampling versus a risk guided on-demand monitoring of plasma concentrations of selenium, copper, and zinc. METHODS: Retrospective analysis of 2 cohorts of patients admitted during 6 months periods in 2006 and 2009 to the ICU of a University hospital. INCLUSION CRITERIA: to receive intravenous micronutrient supplements and/or to have a TE sampling during ICU stay. The TE samplings were triggered by computerized reminder in 2006 versus guided by nutritionists in 2009. RESULTS: During the 2 periods 636 patients met the inclusion criteria out of 2406 consecutive admissions, representing 29.7% and 24.9% respectively of the periods' admissions. The 2009 patients had higher SAPS2 scores (p = 0.02) and lower BMI compared to 2006 (p = 0.007). The number of laboratory determinations was drastically reduced in 2009, particularly during the first week, despite the higher severity of the cohort, resulting in à 55% cost reduction. CONCLUSIONS: The monitoring of TE concentrations guided by a nutritionist resulted in a reduction of the sampling frequency, and targeting on the sickest high risk patients, requiring a nutritional prescription adaptation. This control leads to cost reduction compared to an automated sampling prescription.

A survey of transposable element classification systems--a call for a fundamental update to meet the challenge of their diversity and complexity.

The increase of publicly available sequencing data has allowed for rapid progress in our understanding of genome composition. As new information becomes available we should constantly be updating and reanalyzing existing and newly acquired data. In this report we focus on transposable elements (TEs) which make up a significant portion of nearly all sequenced genomes. Our ability to accurately identify and classify these sequences is critical to understanding their impact on host genomes. At the same time, as we demonstrate in this report, problems with existing classification schemes have led to significant misunderstandings of the evolution of both TE sequences and their host genomes. In a pioneering publication Finnegan (1989) proposed classifying all TE sequences into two classes based on transposition mechanisms and structural features: the retrotransposons (class I) and the DNA transposons (class II). We have retraced how ideas regarding TE classification and annotation in both prokaryotic and eukaryotic scientific communities have changed over time. This has led us to observe that: (1) a number of TEs have convergent structural features and/or transposition mechanisms that have led to misleading conclusions regarding their classification, (2) the evolution of TEs is similar to that of viruses by having several unrelated origins, (3) there might be at least 8 classes and 12 orders of TEs including 10 novel orders. In an effort to address these classification issues we propose: (1) the outline of a universal TE classification, (2) a set of methods and classification rules that could be used by all scientific communities involved in the study of TEs, and (3) a 5-year schedule for the establishment of an International Committee for Taxonomy of Transposable Elements (ICTTE).

Effect of partial-thickness tear on loading capacities of the supraspinatus tendon: a finite element analysis.

Partial-thickness tears of the supraspinatus tendon frequently occur at its insertion on the greater tubercule of the humerus, causing pain and reduced strength and range of motion. The goal of this work was to quantify the loss of loading capacity due to tendon tears at the insertion area. A finite element model of the supraspinatus tendon was developed using in vivo magnetic resonance images data. The tendon was represented by an anisotropic hyperelastic constitutive law identified with experimental measurements. A failure criterion was proposed and calibrated with experimental data. A partial-thickness tear was gradually increased, starting from the deep articular-sided fibres. For different values of tendon tear thickness, the tendon was mechanically loaded up to failure. The numerical model predicted a loss in loading capacity of the tendon as the tear thickness progressed. Tendon failure was more likely when the tendon tear exceeded 20%. The predictions of the model were consistent with experimental studies. Partial-thickness tears below 40% tear are sufficiently stable to persist physiotherapeutic exercises. Above 60% tear surgery should be considered to restore shoulder strength.

Impact of the Femoral Head Position on Moment Arms in Total Hip Arthroplasty: A Parametric Finite Element Study.

BACKGROUND: Although the importance of accurate femoral reconstruction to achieve a good functional outcome is well documented, quantitative data on the effects of a displacement of the femoral center of rotation on moment arms are scarce. The purpose of this study was to calculate moment arms after nonanatomical femoral reconstruction. METHODS: Finite element models of 15 patients including the pelvis, the femur, and the gluteal muscles were developed. Moment arms were calculated within the native anatomy and compared to distinct displacement of the femoral center of rotation (leg lengthening of 10 mm, loss of femoral offset of 20%, anteversion ±10°, and fixed anteversion at 15°). Calculations were performed within the range of motion observed during a normal gait cycle. RESULTS: Although with all evaluated displacements of the femoral center of rotation, the abductor moment arm remained positive, some fibers initially contributing to extension became antagonists (flexors) and vice versa. A loss of 20% of femoral offset led to an average decrease of 15% of abductor moment. Femoral lengthening and changes in femoral anteversion (±10°, fixed at 15°) led to minimal changes in abductor moment arms (maximum change of 5%). Native femoral anteversion correlated with the changes in moment arms induced by the 5 variations of reconstruction. CONCLUSION: Accurate reconstruction of offset is important to maintaining abductor moment arms, while changes of femoral rotation had minimal effects. Patients with larger native femoral anteversion appear to be more susceptible to femoral head displacements.