The effect of cold acclimation on active ion transport in cricket ionoregulatory tissues.

Cold-acclimated insects defend ion and water transport function during cold exposure. We hypothesized that this is achieved via enhanced active transport. The Malpighian tubules and rectum are likely targets for such transport modifications, and recent transcriptomic studies indicate shifts in Na+-K+ ATPase (NKA) and V-ATPase expression in these tissues following cold acclimation. Here we quantify the effect of cold acclimation (one week at 12 °C) on active transport in the ionoregulatory organs of adult Gryllus pennsylvanicus field crickets. We compared primary urine production of warm- and cold-acclimated crickets in excised Malpighian tubules via Ramsay assay at a range of temperatures between 4 and 25 °C. We then compared NKA and V-ATPase activities in Malpighian tubule and rectal homogenates from warm- and cold-acclimated crickets via NADH-linked photometric assays. Malpighian tubules of cold-acclimated crickets excreted fluid at lower rates at all temperatures compared to warm-acclimated crickets. This reduction in Malpighian tubule excretion rates may be attributed to increased NKA activity that we observed for cold-acclimated crickets, but V-ATPase activity was unchanged. Cold acclimation had no effect on rectal NKA activity at either 21 °C or 6 °C, and did not modify rectal V-ATPase activity. Our results suggest that an overall reduction, rather than enhancement of active transport in the Malpighian tubules allows crickets to maintain hemolymph water balance during cold exposure, and increased Malpighian tubule NKA activity may help to defend and/or re-establish ion homeostasis

30 inch Roll-Based Production of High-Quality Graphene Films for Flexible Transparent Electrodes

We report that 30-inch scale multiple roll-to-roll transfer and wet chemical doping considerably enhance the electrical properties of the graphene films grown on roll-type Cu substrates by chemical vapor deposition. The resulting graphene films shows a sheet resistance as low as ~30 Ohm/sq at ~90 % transparency which is superior to commercial transparent electrodes such as indium tin oxides (ITO). The monolayer of graphene shows sheet resistances as low as ~125 Ohm/sq with 97.4% optical transmittance and half-integer quantum Hall effect, indicating the high-quality of these graphene films. As a practical application, we also fabricated a touch screen panel device based on the graphene transparent electrodes, showing extraordinary mechanical and electrical performances

The effects inhibiting the proliferation of cancer cells by far-infrared radiation (FIR) are controlled by the basal expression level of heat shock protein (HSP) 70A

We developed a tissue culture incubator that can continuously irradiate cells with far-infrared radiation (FIR) of wavelengths between 4 and 20 μm with a peak of 7–12 μm, and found that FIR caused different inhibiting effects to five human cancer cell lines, namely A431 (vulva), HSC3 (tongue), Sa3 (gingiva), A549 (lung), and MCF7 (breast). Then, in order to make clear the control system for the effect of FIR, the gene expression concerned to the inhibition effect by FIR were analyzed. In consequence, basal expression level of HSP70A mRNA was higher in A431 and MCF7 cells than in the FIR-sensitive HSC3, Sa3, and A549 cells. Also, the over expression of HSP70 inhibited FIR-induced growth arrest in HSC3 cells, and an HSP70 siRNA inhibited the proliferation of A431 cells by irradiation with FIR. These results indicate that the effect of a body temperature range of FIR suppressing the proliferation of some cancer cells is controlled by the basal expression level of heat shock protein (HSP) 70A. This finding suggested that FIR should be very effective medical treatment for some cancer cells which have a low level of HSP70. Still more, if the level of HSP70 in any cancer of a patient was measured, the effect of medical treatment by FIR can be foreseen for the cancer

Engineering the Controlled Assembly of Filamentous Injectisomes in E. coli K-12 for Protein Translocation into Mammalian Cells.

Bacterial pathogens containing type III protein secretion systems (T3SS) assemble large needle-like protein complexes in the bacterial envelope, called injectisomes, for translocation of protein effectors into host cells. The application of these molecular syringes for the injection of proteins into mammalian cells is hindered by their structural and genomic complexity, requiring multiple polypeptides encoded along with effectors in various transcriptional units (TUs) with intricate regulation. In this work, we have rationally designed the controlled expression of the filamentous injectisomes found in enteropathogenic Escherichia coli (EPEC) in the nonpathogenic strain E. coli K-12. All structural components of EPEC injectisomes, encoded in a genomic island called the locus of enterocyte effacement (LEE), were engineered in five TUs (eLEEs) excluding effectors, promoters and transcriptional regulators. These eLEEs were placed under the control of the IPTG-inducible promoter Ptac and integrated into specific chromosomal sites of E. coli K-12 using a marker-less strategy. The resulting strain, named synthetic injector E. coli (SIEC), assembles filamentous injectisomes similar to those in EPEC. SIEC injectisomes form pores in the host plasma membrane and are able to translocate T3-substrate proteins (e.g., translocated intimin receptor, Tir) into the cytoplasm of HeLa cells reproducing the phenotypes of intimate attachment and polymerization of actin-pedestals elicited by EPEC bacteria. Hence, SIEC strain allows the controlled expression of functional filamentous injectisomes for efficient translocation of proteins with T3S-signals into mammalian cells

Feasibility of hydraulic separation in a novel anaerobic-anoxic upflow reactor for biological nutrient removal

ABSTRACT : This contribution deals with a novel anaerobic-anoxic reactor for biological nutrient removal (BNR) from wastewater, termed AnoxAn. In the AnoxAn reactor, the anaerobic and anoxic zones for phosphate removal and denitrification are integrated in a single continuous upflow sludge blanket reactor, aiming at high compactness and efficiency. Its application is envisaged in those cases where retrofitting of existing wastewater treatment plants for BNR, or the construction of new ones, is limited by the available surface area. The environmental conditions are vertically divided up inside the reactor with the anaerobic zone at the bottom and the anoxic zone above. The capability of the AnoxAn configuration to establish two hydraulically separated zones inside the single reactor was assessed by means of hydraulic characterization experiments and model simulations. Residence time distribution (RTD) experiments in clean water were performed in a bench-scale (48.4 L) AnoxAn prototype. The required hydraulic separation between the anaerobic and anoxic zones, as well as adequate mixing in the individual zones, was obtained through selected mixing devices. The observed behaviour was described by a hydraulic model consisting of continuous stirred tank reactors and plug-flow reactors. The impact of the denitrification process in the anoxic zone on the hydraulic separation was subsequently evaluated through model simulations. The desired hydraulic behaviour proved feasible, involving little mixing between the anaerobic and anoxic zones (mixing flowrate 40.2% of influent flowrate) and negligible nitrate concentration in the anaerobic zone (less than 0.1 mgN L-1) when denitrification was considered

Wafer-Scale, Sub-5 nm Junction Formation by Monolayer Doping and Conventional Spike Annealing

We report the formation of sub-5 nm ultrashallow junctions in 4 inch Si wafers enabled by the molecular monolayer doping of phosphorous and boron atoms and the use of conventional spike annealing. The junctions are characterized by secondary ion mass spectrometry and non-contact sheet resistance measurements. It is found that the majority (~70%) of the incorporated dopants are electrically active, therefore, enabling a low sheet resistance for a given dopant areal dose. The wafer-scale uniformity is investigated and found to be limited by the temperature homogeneity of the spike anneal tool used in the experiments. Notably, minimal junction leakage currents (<1 uA/cm2) are observed which highlights the quality of the junctions formed by this process. The results clearly demonstrate the versatility and potency of the monolayer doping approach for enabling controlled, molecular-scale ultrashallow junction formation without introducing defects in the semiconductor.Comment: 21 pages, 5 figure

The genomes of two key bumblebee species with primitive eusocial organization

Background: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats. Results: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits. Conclusions: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation

Longitudinal Replication Studies of GWAS Risk SNPs Influencing Body Mass Index over the Course of Childhood and Adulthood

Genome-wide association studies (GWAS) have identified multiple common variants associated with body mass index (BMI). In this study, we tested 23 genotyped GWAS-significant SNPs (p-value<5*10-8) for longitudinal associations with BMI during childhood (3–17 years) and adulthood (18–45 years) for 658 subjects. We also proposed a heuristic forward search for the best joint effect model to explain the longitudinal BMI variation. After using false discovery rate (FDR) to adjust for multiple tests, childhood and adulthood BMI were found to be significantly associated with six SNPs each (q-value<0.05), with one SNP associated with both BMI measurements: KCTD15 rs29941 (q-value<7.6*10-4). These 12 SNPs are located at or near genes either expressed in the brain (BDNF, KCTD15, TMEM18, MTCH2, and FTO) or implicated in cell apoptosis and proliferation (FAIM2, MAP2K5, and TFAP2B). The longitudinal effects of FAIM2 rs7138803 on childhood BMI and MAP2K5 rs2241423 on adulthood BMI decreased as age increased (q-value<0.05). The FTO candidate SNPs, rs6499640 at the 5 ′-end and rs1121980 and rs8050136 downstream, were associated with childhood and adulthood BMI, respectively, and the risk effects of rs6499640 and rs1121980 increased as birth weight decreased. The best joint effect model for childhood and adulthood BMI contained 14 and 15 SNPs each, with 11 in common, and the percentage of explained variance increased from 0.17% and 9.0*10−6% to 2.22% and 2.71%, respectively. In summary, this study evidenced the presence of long-term major effects of genes on obesity development, implicated in pathways related to neural development and cell metabolism, and different sets of genes associated with childhood and adulthood BMI, respectively. The gene effects can vary with age and be modified by prenatal development. The best joint effect model indicated that multiple variants with effects that are weak or absent alone can nevertheless jointly exert a large longitudinal effect on BMI

SPAG5 as a prognostic biomarker and chemotherapy sensitivity predictor in breast cancer: a retrospective, integrated genomic, transcriptomic, and protein analysis

Background: Although the use of proliferation markers/profiles has been recommended when choosing the appropriate systemic-treatment for breast cancer (BC), the best molecular-marker/test that should be used needs to be identified. Methods: To identify factors that drive proliferation and its associated features in BC an artificial neural network (ANN) based integrative data-mining methodology was applied to three cohorts [(Nottingham-discovery (ND), Uppsala and METABRIC (Molecular Taxonomy of Breast Cancer International Consortium)]. The most prominent genes in the resulting interactome-map were then identified. Given that SPAG5 was associated with many features of proliferation, featured prominently in the interactome-map and has a fundamental role in mitotic-progression,, we hypothesized that it could be a better indicator of proliferation activity. (BC). Subsequently to test if it could provide a more accurate guide for the delivery of systemic therapies in BC, we investigated the clinico-pathological utility of SPAG5: gene copy number aberrations (CNAs); mRNA and protein expression, in over 10,000 BCs. Integrated analysis of SPAG5-gene CNAs, transcript and protein expression was conducted in the ND cohort (n=171) and validated in the METABRIC cohort (n=1980). In addition, the associations of SPAG5 CNAs, transcript and/or protein with BC specific survival (BCSS), disease free survival (DFS) and/or distant relapse free survival (DRFS) were analysed in multiple cohorts including Uppsala (n=249), METABRIC, three-untreated lymph node (LN) negative cohorts (n=684), a combined multicentre clinical data set (n=5439), Nottingham historical early-stage-primary BC (Nottingham-HES-BC; n=1650), Nottingham oestrogen receptor (ER) negative BC (n=697), Nottingham anthracycline-Neoadjuvant-chemotherapy (Nottingham-AC-Neo-ACT; n=200), and MD Anderson Cancer Centre Taxane/anthracycline (MDACC-T/AC-Neo-ACT; n=508) cohorts. The association of SPAG5 transcript and protein expression with pathological response rate (pCR) were also tested in [MDACC-T/AC-Neo-ACT (n=508) and the phase II trial NCT00455533; n=253)] and [Nottingham-AC-Neo-ACT (n=200)] cohorts; respectively. Findings: SPAG5 gene gain/amplification at the Ch17q11·2 locus was found in 10.4% of BC (206/1980 (; METABRIC) and was reported in 19·4% of PAM50-HER2 (46/237) and 17·8% of PAM50-LumB (87/488). SPAG5-CNA gain/amplification and high SPAG5-transcript and SPAG5-protein were associated with increased risk of death from BC [Uppsala; (HR (CI 95%): 1·50 (1·18-1·92); p=0·00010, METABRIC; (HR (CI 95%): 1·68 (1·40-2·01) p<0·0001), and Nottingham-HSE-BC; (HR (CI 95%): 1·68 (1·32-2·12), p<0·0001); respectively]. Multivariable Cox regression models, including other validated-prognostic factors, (Uppsala: age, size, LN-stage, genomic grade index (GGI), ER, TP53 mutation and MKi67; METABRIC: age, size, LN-stage, histologic-grade, ER, progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), hormone-therapy, chemotherapy, interaction term of SPAG5 and both chemotherapyy and hormonotherapy; Desmedt-untreated LN- cohort: ER, Nottingham prognostic index (NPI), 76-gene prognostic signature (Veridex) and Adjuvant-Online (AOL); Nottingham-HES-BC: menopausal status, size, LN- stage, histologic-grade, ER, PR, HER2, ki67, hormone-therapy, chemotherapy, interaction term of SPAG5 and both chemotherapy[y and hormonotherapy), showed that high SPAG5-transcript and high SPAG5-protein were associated with shorter BCSS [Uppsala: (HR (CI 95%): 1·62 (1·03-2·53) p=0·036); METABRIC: (HR (CI 95%): 1·27 (1·02-1·58) p=0·034); Desmedt-untreated LN- cohort: (HR (CI 95%): 2·34 (1·24-4·42) p=0·0090), and Nottingham-HES-BC (HR (CI 95%): 1·73 (1·23-2·46) p=0·0020); respectively]. In ER-negative-BC with high SPAG5-protein, administration of anthracycline-adjuvant-chemotherapy had reduced the risk of death by 60% compared to chemotherapy-naive (HR (95% CI): 0·37 (0·20-0·60); p=0·0010). A multivariable Cox regression analysis, which included other validated prognostic factors for chemotherapy (e.g., menopausal status, size, lymph node stage, histologic grade, ER, PR, HER2, Bcl2, chemotherapy, interaction term of SPAG5 and both chemotherapy[y), revealed that SPAG5-transcript+ was independently associated with decreased risk of DRFS after receiving Taxane/anthracycline-Neo-ACT [MDACC-T/AC-Neo-ACT: (HR (CI 95%): 0·68 (0·48-0·97); p=0·0070)]. In multivariable logistic regression analysis, both SPAG5-transcript+ and SPAG5-protein+ and were independent predictors for higher pCR after combination-cytotoxic chemotherapy [MDACC-T/AC-Neo-ACT: (OR (95% CI) 1·71 (1·07-2·74); p=0·024), and Nottingham-AC-Neo-AC: (OR (95% CI): 8·75 (2·42-31); p=0·0010); respectively]. Interpretation: SPAG5 is a novel amplified gene on Ch17q11.2 in PAM50-LumB and PAM-HER2 BC, and its transcript and protein products are independent prognostic and predictive biomarkers, with potential clinical utility as a biomarker for combination cytotoxic chemotherapy sensitivity, especially in ER- BC

Cell Free Expression of hif1α and p21 in Maternal Peripheral Blood as a Marker for Preeclampsia and Fetal Growth Restriction

Preeclampsia, a severe unpredictable complication of pregnancy, occurs in 6% of pregnancies, usually in the second or third trimester. The specific etiology of preeclampsia remains unclear, although the pathophysiological hallmark of this condition appears to be an inadequate blood supply to the placenta. As a result of the impaired placental blood flow, intrauterine growth restriction (IUGR) and consequential fetal oxidative stress may occur. Consistent with this view, pregnancies complicated by preeclampsia and IUGR are characterized by up-regulation of key transcriptional regulators of the hypoxic response including, hif1α and as well as p53 and its target genes. Recently, the presence of circulating cell-free fetal RNA has been documented in maternal plasma. We speculated that pregnancies complicated by preeclampsia and IUGR, will be associated with an abnormal expression of p53 and/or hif1α related genes in the maternal plasma. Maternal plasma from 113 singleton pregnancies (72 normal and 41 complicated pregnancies) and 19 twins (9 normal and 10 complicated pregnancies) were collected and cell free RNA was extracted. The expression of 18 genes was measured by one step real-time RT-PCR and was analyzed for prevalence of positive/negative expression levels. Results indicate that, among the genes examined, cell free plasma expressions of p21 and hif1α were more prevalent in pregnancies complicated by hypoxia and/or IUGR (p<0.001). To conclude, we present in this manuscript data to support the association between two possible surrogate markers of hypoxia and common complications of pregnancy. More work is needed in order to implement these findings in clinical practice