The functional roles of disulfide bonds in the β-subunit of (Na,K)ATPase as studied by site-directed mutagenesis

AbstractThe β-subunit of Torpedo californica (Na,K)ATPase contains seven cysteine residues; one (Cys46) is in the single transmembrane segment and the other six (Cys127, Cys150, Cys160, Cys176, Cys215 and Cys278) are in the extracellular domain and form three highly conserved disulfide bonds. Aβ-subunit mutant with replacement of Cys46 by Ser could assemble with the α-subunit, and the resulting αβ-complex was catalytically active. Mutants in which either the N-terminal side or both Cys residues of the Cys127-Cys150 bond were replaced by Ser could also tightly assemble with the α-subunit, but the resulting αβ-complex was catalytically inactive. On the other hand, disruption of either the Cys160-Cys176 or Cys215-Cys278 bond by substituting the N-terminal side only or both Cys residues with Ser led to a β-subunit that could not assemble with the α-subunit. We conclude that the structure of the β-subunit around the Cys160-Cys176 and Cys215-Cys278 loops is indispensable for assembly with the α-subunit, whereas the Cys127-Cys150 loop is not essential for assembly but is required for enzyme activity

An enhancer unit of L-type pyruvate kinase gene is responsible for transcriptional stimulation by dietary fructose as well as glucose in transgenic mice

AbstractWe produced three lines of transgenic mice containing the 5' flanking region of the L-type pyruvate kinase gene from nucleotides −189 to +37, which includes an enhancer unit and TATA box as functional elements, linked to the chloramphenicol acetyltransferase gene. Since transgene expression was stimulated by both dietary fructose and glucose in a tissue-dependent manner, we suggest that this unit is responsive to both stimuli

A mindfulness-based stress management program and treatment with omega-3 fatty acids to maintain a healthy mental state in hospital nurses (Happy Nurse Project): study protocol for a randomized controlled trial

Background: It is reported that nursing is one of the most vulnerable jobs for developing depression. While they may not be clinically diagnosed as depressed, nurses often suffer from depression and anxiety symptoms, which can lead to a low level of patient care. However, there is no rigorous evidence base for determining an effective prevention strategy for these symptoms in nurses. After reviewing previous literature, we chose a strategy of treatment with omega-3 fatty acids and a mindfulness-based stress management program for this purpose. We aim to explore the effectiveness of these intervention options for junior nurses working in hospital wards in Japan. Methods/Design: A factorial-design multi-center randomized trial is currently being conducted. A total of 120 nurses without a managerial position, who work for general hospitals and gave informed consent, have been randomly allocated to a stress management program or psychoeducation using a leaflet, and to omega-3 fatty acids or identical placebo pills. The stress management program has been developed according to mindfulness cognitive therapy and consists of four 30-minute individual sessions conducted using a detailed manual. These sessions are conducted by nurses with a managerial position. Participants allocated to the omega-3 fatty acid groups are provided with 1, 200 mg/day of eicosapentaenoic acid and 600 mg/day of docosahexaenoic acid for 90 days. Discussion: An effective preventive intervention may not only lead to the maintenance of a healthy mental state in nurses, but also to better quality of care for inpatients. This paper outlines the background and methods of a randomized trial that evaluates the possible additive value of omega-3 fatty acids and a mindfulness-based stress management program for reducing depression in nurses

Intrinsic Shape of Star-Forming BzK Galaxies at z~2 in GOODS-N

We study structure of star-forming galaxies at z~2 in GOODS-N field selected as sBzK galaxies down to K_{AB} <24.0 mag. Among 1029 sBzK galaxies, 551 galaxies (54%) show a single component in ACS/F850LP image obtained with the Hubble Space Telescope; the rest show multiple components. We fit the single-component sBzK galaxies with the single S\'ersic profile using the ACS/F850LP image and find that a majority of them (64%) show S\'ersic index of n=0.5-2.5, indicating that they have a disk-like structure. The resulting effective radii typically range from 1.0 to 3.0 kpc in the rest-frame UV wavelength. After correcting the effective radii to those in the rest-frame optical wavelength, we find that the single-component sBzK galaxies locate in the region where the local and z~1 disk galaxies distribute in the stellar mass-size diagram, suggesting comparable surface stellar mass density between the sBzK and z~0-1 disk galaxies. All these properties suggest that the single-component sBzK galaxies are progenitors of the present-day disk galaxies. However, by studying their intrinsic shape through comparison between the observed distribution of apparent axial ratios and the distribution for triaxial models with axes (A>B>C), we find that the mean B/A ratio is 0.61^{+0.05}_{-0.08} and disk thickness C/A is 0.28^{+0.03}_{-0.04}. This indicates that the single-component sBzK galaxies at z~2 have a bar-like or oval shape rather than a round disk shape. The shape seems to resemble to a bar/oval structure that form through bar instability; if it is the case, the intrinsic shape may give us a clue to understand dynamical evolution of baryonic matter in a dark matter halo.Comment: 14 pages, 13 figure, 2 tables, Accepted for publication in Ap

Polygenic architecture informs potential vulnerability to drug-induced liver injury

Drug-Induced-Liver-Injury (DILI) is a leading cause of termination in drug development programs and removal of drugs from the market, and this is partially due to the inability to identify patients who are at risk1. Here, we developed a polygenic risk score (PRS) for DILI by aggregating effects of numerous genome-wide loci identified from previous large-scale genome-wide association studies (GWAS)2. The PRS predicted the susceptibility to DILI in patients treated with fasiglifam, amoxicillin-clavulanate or flucloxacillin, and in primary hepatocytes and stem cell-derived organoids from multiple donors treated with over 10 different drugs. Pathway analysis highlighted processes previously implicated in DILI, including unfolded protein responses and oxidative stress. In silico screening identified compounds that elicit transcriptomic signatures present in hepatocytes from individuals with elevated PRS, supporting mechanistic links and suggesting a novel screen for safety of new drug candidates. This genetic-, cellular-, organoid- and human-scale evidence underscored the polygenic architecture underlying DILI vulnerability at the level of hepatocytes, thus facilitating future mechanistic studies. Moreover, the proposed “polygenicity-in-a-dish” strategy might potentially inform designs of safer, more efficient, and robust clinical trials

Differential tumor surveillance by natural killer (NK) and NKT cells

Natural tumor surveillance capabilities of the host were investigated in six different mouse tumor models where endogenous interleukin (IL)-12. does or does not dictate the efficiency of the innate immune response. Gene-targeted and lymphocyte subset-depleted mice were used to establish the relative importance of natural killer (NK) and NK1.1(+) T (NKT) cells in protection from tumor initiation and metastasis. In the models examined, CD3(-) NK cells were responsible for tumor rejection and protection from metastasis in models where control of major histocompatibility complex class I-deficient tumors was independent of IL-12, A protective role for NKT cells was only observed when tumor rejection required endogenous IL-12 activity. In particular, T cell receptor J alpha 281 gene-targeted mice confirmed a critical function for NKT cells in protection from spontaneous tumors initiated by the chemical carcinogen, methylcholanthrene. This is the first description of an antitumor function for NKT cells in the absence of exogenously administered potent stimulators such as IL-12 or alpha-galactosylceramide

Abnormal spermatogenesis and male infertility in testicular zinc finger protein Zfp318-knockout mice

Zfp318, a mouse gene with a Cys2/His2 zinc finger motif, is mainly expressed in germ cells in the testis. It encodes two alternative transcripts, which regulate androgen receptor-mediated transcriptional activation or repression by overexpression of them. However, the role of Zfp318 is still obscure in vivo, especially in spermatogenesis. To elucidate the role of Zfp318 during gamete production, we established a knockout mouse line. Zfp318-null male mice exhibited infertility, whereas Zfp318-null female mice displayed normal fertility. ZFP318 was expressed during multiple stages of spermatogenesis, from spermatocytes to round spermatids. The nuclei of secondary spermatocytes showed high levels of expression. Histological analysis and quantitative analysis of DNA content showed decreased numbers of both spermatids in the seminiferous tubules and mature spermatozoa in the epididymides of Zfp318-null mice. These results suggest that Zfp318 is expressed as a functional protein in testicular germ cells and plays an important role in meiosis during spermatogenesis