Structural basis for myopathic defects engendered by alterations in the myosin rod

While mutations in the myosin subfragment 1 motor domain can directly disrupt the generation and transmission of force along myofibrils and lead to myopathy, the mechanism whereby mutations in the myosin rod influences mechanical function is less clear. Here, we used a combination of various imaging techniques and molecular dynamics simulations to test the hypothesis that perturbations in the myosin rod can disturb normal sarcomeric uniformity and, like motor domain lesions, would influence force production and propagation. We show that disrupting the rod can alter its nanomechanical properties and, in vivo, can drive asymmetric myofilament and sarcomere formation. Our imaging results indicate that myosin rod mutations likely disturb production and/or propagation of contractile force. This provides a unifying theory where common pathological cascades accompany both myosin motor and specific rod domain mutations. Finally, we suggest that sarcomeric inhomogeneity, caused by asymmetric thick filaments, could be a useful index of myopathic dysfunction.     

Changes in certain kinetic properties of hepatic microsomal aniline hydroxylase and ethylmorphine demethylase associated with postnatal development and maturation in male rats

1. Changes in certain kinetic properties (V(max.) and apparent K(m)) of hepatic microsomal mixed-function oxidases have been studied as a function of postnatal development and maturation in male rats. 2. Microsomal cytochrome P-450 content changed only slightly between 1 and 12 weeks of age. 3. Aniline hydroxylase activity (V(max.)) increased abruptly between 1 and 2 weeks of age to greater than adult activities and then returned to a plateau value between 4(1/2) and 12 weeks of age. Ethylmorphine demethylase activity remained low and relatively constant between 1 and 3 weeks of age and then increased markedly ( approximately 100%) between 3 and 4(1/2) weeks. 4. The apparent Michaelis constant (K(m)) for aniline hydroxylation increased almost linearly with time between 1 and 6 weeks of age and tended to reach a plateau value thereafter. The apparent K(m) for ethylmorphine demethylation increased between 1 and 3 weeks of age and then decreased abruptly to a constant value between 6 and 12 weeks. 5. The data indicate that developmental changes in the activity of these microsomal oxidases do not correlate temporally with each other or with changes in microsomal cytochrome P-450 content. 6. The most dramatic changes in enzyme activity were associated with early development (1-3 weeks) and weaning (3-4 weeks). 7. Changes in weight of seminal vesicle, a criterion of sexual maturation in male rats, were most prominent between 6 and 8 weeks of age and thus appeared to be separated in time from the prominent changes in enzyme activity.     

New Insights into Muscle Function during Pivot Feeding in Seahorses

Seahorses, pipefish and their syngnathiform relatives are considered unique amongst fishes in using elastic recoil of post-cranial tendons to pivot the head extremely quickly towards small crustacean prey. It is known that pipefish activate the epaxial muscles for a considerable time before striking, at which rotations of the head and the hyoid are temporarily prevented to allow energy storage in the epaxial tendons. Here, we studied the motor control of this system in seahorses using electromyographic recordings of the epaxial muscles and the sternohyoideus-hypaxial muscles with simultaneous high-speed video recordings of prey capture. In addition we present the results from a stimulation experiment including the muscle hypothesised to be responsible for the locking and triggering of pivot feeding in seahorses (m. adductor arcus palatini). Our data confirmed that the epaxial pre-activation pattern observed previously for pipefish also occurs in seahorses. Similar to the epaxials, the sternohyoideus-hypaxial muscle complex shows prolonged anticipatory activity. Although a considerable variation in displacements of the mouth via head rotation could be observed, it could not be demonstrated that seahorses have control over strike distance. In addition, we could not identify the source of the kinematic variability in the activation patterns of the associated muscles. Finally, the stimulation experiment supported the previously hypothesized role of the m. adductor arcus palatini as the trigger in this elastic recoil system. Our results show that pre-stressing of both the head elevators and the hyoid retractors is taking place. As pre-activation of the main muscles involved in pivot feeding has now been demonstrated for both seahorses and pipefish, this is probably a generalized trait of Syngnathidae.     

Plastidial Expression of Type II NAD(P)H Dehydrogenase Increases the Reducing State of Plastoquinones and Hydrogen Photoproduction Rate by the Indirect Pathway in Chlamydomonas reinhardtii

Biological conversion of solar energy into hydrogen is naturally realized by some microalgae species due to a coupling between the photosynthetic electron transport chain and a plastidial hydrogenase. While promising for the production of clean and sustainable hydrogen, this process requires improvement to be economically viable. Two pathways, called direct and indirect photoproduction, lead to sustained hydrogen production in sulfur-deprived Chlamydomonas reinhardtii cultures. The indirect pathway allows an efficient time-based separation of O₂ and H₂ production, thus overcoming the O₂ sensitivity of the hydrogenase, but its activity is low. With the aim of identifying the limiting step of hydrogen production, we succeeded in overexpressing the plastidial type II NAD(P)H dehydrogenase (NDA2). We report that transplastomic strains overexpressing NDA2 show an increased activity of nonphotochemical reduction of plastoquinones (PQs). While hydrogen production by the direct pathway, involving the linear electron flow from photosystem II to photosystem I, was not affected by NDA2 overexpression, the rate of hydrogen production by the indirect pathway was increased in conditions, such as nutrient limitation, where soluble electron donors are not limiting. An increased intracellular starch was observed in response to nutrient deprivation in strains overexpressing NDA2. It is concluded that activity of the indirect pathway is limited by the nonphotochemical reduction of PQs, either by the pool size of soluble electron donors or by the PQ-reducing activity of NDA2 in nutrient-limited conditions. We discuss these data in relation to limitations and biotechnological improvement of hydrogen photoproduction in microalgae.A number of microalgal and cyanobacterial species are able to convert solar energy into hydrogen by photobiological processes and are therefore considered promising organisms for developing clean and sustainable hydrogen production (Benemann, 2000; Ghirardi et al., 2000; Rupprecht et al., 2006). In microalgae, hydrogen photoproduction results from coupling the photosynthetic electron transport chain and a plastidial [FeFe] hydrogenase. Under most conditions, hydrogen photoproduction is a transient phenomenon that lasts from several seconds to a few minutes (Ghirardi et al., 2000; Melis and Happe, 2001). It has been considered a relic of evolution that may now serve, under certain environmental conditions, such as induction of photosynthesis in anoxia (Ghysels et al., 2013), as a safety valve that protects the photosynthetic electron transport chain from photodamage that results from overreduction of electron acceptors (Kessler, 1973; Tolleter et al., 2011). A major limitation to sustained hydrogen photoproduction is due to the oxygen sensitivity of the [FeFe] hydrogenase (Happe et al., 2002; Stripp et al., 2009). Melis et al. (2000) proposed an elegant way to overcome this oxygen sensitivity through a time-based separation of hydrogen and oxygen production phases occurring, for instance, in response to sulfur deficiency in a closed environment. Another limitation is related to the electron supply for the hydrogenase coming from the photosynthetic electron transport chain (Cournac et al., 2002). This limitation is partly due to the fact that other metabolic pathways, such as ferredoxin-NADP⁺ reductase and CO₂ fixation, compete with the hydrogenase for the use of reduced ferredoxin (Gaffron and Rubin, 1942; Hemschemeier et al., 2008). This is also due to upstream regulation of the electron transport chain, recently evidenced from the study of a Chlamydomonas reinhardtii mutant affected in proton gradient regulation-like1 (PGRL1)-mediated cyclic electron flow (CEF) around PSI. The strong enhancement of hydrogen production rates observed in the pgrl1 mutant was interpreted as the release of a control exerted by the transthylakoidal pH gradient on electron supply to the hydrogenase (Tolleter et al., 2011).Two pathways, direct or indirect, can supply electrons to the hydrogenase (Benemann, 2000; Melis and Happe, 2001; Chochois et al., 2009). In the direct pathway, the whole electron transport chain is engaged, with PSII supplying electrons to the plastoquinone (PQ) pool, the cytochrome b₆/f complex, and, in turn, PSI, ferredoxin, and the [FeFe] hydrogenase. Due to the high oxygen sensitivity of the [FeFe] hydrogenase and to the fact that O₂ is produced during photosynthesis at PSII, the direct pathway only operates when PSII activity is lower than mitochondrial respiration, thereby allowing anaerobiosis to be maintained. Such conditions can be obtained by decreasing PSII activity either by means of sulfur deprivation (Melis et al., 2000) or by decreasing light intensity in the photobioreactor (Degrenne et al., 2010). In the indirect pathway, reducing equivalents, stored as starch during the aerobic phase, are subsequently used to fuel hydrogen production. This implies a nonphotochemical reduction of the PQ pool that is at least in part mediated by NDA2, a type II NADH dehydrogenase discovered in C. reinhardtii chloroplasts (Desplats et al., 2009). RNA interference lines expressing lower levels of NDA2 show lower hydrogen production rates, and it was concluded that NDA2 is involved in hydrogen production by the indirect pathway (Jans et al., 2008; Mignolet et al., 2012). The indirect pathway allows for an efficient time-based separation of O₂- and H₂-producing phases because it does not involve PSII activity and does not produce O₂. However, the indirect pathway has a much lower rate than the direct pathway (Cournac et al., 2002; Antal et al., 2009; Chochois et al., 2009). With the aim to identify limiting steps of hydrogen production in microalgae, we attempted to overexpress NDA2 in C. reinhardtii chloroplasts. We report that algal strains displaying a 2-fold increase in NDA2 show an increased nonphotochemical reduction of PQs and an increased rate of hydrogen production by the indirect pathway, the latter being only observed in conditions where stromal reducing equivalents are available in sufficient amounts.     

A Bayesian multi-risks survival (MRS) model in the presence of double censorings

Semi-competing risks data include the time to a nonterminating event and the time to a terminating event, while competing risks data include the time to more than one terminating event. Our work is motivated by a prostate cancer study, which has one nonterminating event and two terminating events with both semi-competing risks and competing risks present as well as two censoring times. In this paper, we propose a new multi-risks survival (MRS) model for this type of data. In addition, the proposed MRS model can accommodate noninformative right-censoring times for nonterminating and terminating events. Properties of the proposed MRS model are examined in detail. Theoretical and empirical results show that the estimates of the cumulative incidence function for a nonterminating event may be biased if the information on a terminating event is ignored. A Markov chain Monte Carlo sampling algorithm is also developed. Our methodology is further assessed using simulations and also an analysis of the real data from a prostate cancer study. As a result, a prostate-specific antigen velocity greater than 2.0 ng/mL per year and higher biopsy Gleason scores are positively associated with a shorter time to death due to prostate cancer.     

A generalized,lumped-parameter model of photosynthesis,evapotranspiration and net primary production in temperate and boreal forest ecosystems

Summary PnET is a simple, lumped-parameter, monthlytime-step model of carbon and water balances of forests built on two principal relationships: 1) maximum photosynthetic rate is a function of foliar nitrogen concentration, and 2) stomatal conductance is a function of realized photosynthetic rate. Monthyly leaf area display and carbon and water balances are predicted by combining these with standard equations describing light attenuation in canopies and photosynthetic response to diminishing radiation intensity, along with effects of soil water stress and vapor pressure deficit (VPD). PnET has been validated against field data from 10 well-studied temperate and boreal forest ecosystems, supporting our central hypothesis that aggregation of climatic data to the monthly scale and biological data such as foliar characteristics to the ecosystem level does not cause a significant loss of information relative to long-term, mean ecosystem responses. Sensitivity analyses reveal a diversity of responses among systems to identical alterations in climatic drivers. This suggests that great care should be used in developing generalizations as to how forests will respond to a changing climate. Also critical is the degree to which the temperature responses of photosynthesis and respiration might acclimate to changes in mean temperatures at decadal time scales. An extreme climate change simulation (+3° C maximum temperature, –25% precipitation with no change in minimum temperature or radiation, direct effects of increased atmospheric CO₂ ignored) suggests that major increases in water stress, and reductions in biomass production (net carbon gain) and water yield would follow such a change.     

The oxidation of glucose by Acinetobacter calcoaceticus: interaction of the quinoprotein glucose dehydrogenase with the electron transport chain

The coupling of the quinoprotein glucose dehydrogenase to the electron transport chain has been investigated in Acinetobacter calcoaceticus. No evidence was obtained to support a previous suggestion that the soluble form of the dehydrogenase and the soluble cytochrome b associated with it are involved in the oxidation of glucose. Analysis of cytochrome content, and of reduction of cytochromes in membranes by substrates, and of sensitivity to cyanide indicated that glucose, succinate and NADH are all oxidized by way of the same b-type cytochrome(s) and cytochrome oxidases (cytochrome o and cytochrome d). Mixed inhibition studies [with KCN and hydroxyquinoline N-oxide (HQNO)] showed that the b-type cytochrome(s) formed a binary complex with the o-type oxidase and that there was thus no communication between the electron transport chains at the cytochrome level. Measurements of the reduction of ubiquinone-9 by glucose and NADH, and inhibitor studies using HQNO, indicated that the ubiquinone mediates electron transport from both the glucose and NADH dehydrogenases. In some conditions the quinone pool facilitated communication between the 'glucose oxidase' and 'NADH oxidase' electron transport chains, but in normal conditions these chains were kinetically distinct.     

Randomized single oral dose phase 1 study of safety,tolerability, and pharmacokinetics of Iminosugar UV-4 Hydrochloride (UV-4B) in healthy subjects

BackgroundUV-4 (N-(9’-methoxynonyl)-1-deoxynojirimycin, also called MON-DNJ) is an iminosugar small-molecule oral drug candidate with in vitro antiviral activity against diverse viruses including dengue, influenza, and filoviruses and demonstrated in vivo efficacy against both dengue and influenza viruses. The antiviral mechanism of action of UV-4 is through inhibition of the host endoplasmic reticulum-resident α-glucosidase 1 and α-glucosidase 2 enzymes. This inhibition prevents proper glycan processing and folding of virus glycoproteins, thereby impacting virus assembly, secretion, and the fitness of nascent virions.Methodology/Principal findingsHere we report a first-in-human, single ascending dose Phase 1a study to evaluate the safety, tolerability, and pharmacokinetics of UV-4 hydrochloride (UV-4B) in healthy subjects (ClinicalTrials.gov Identifier {"type":"clinical-trial","attrs":{"text":"NCT02061358","term_id":"NCT02061358"}}NCT02061358). Sixty-four subjects received single oral doses of UV-4 as the hydrochloride salt equivalent to 3, 10, 30, 90, 180, 360, 720, or 1000 mg of UV-4 (6 subjects per cohort), or placebo (2 subjects per cohort). Single doses of UV-4 hydrochloride were well tolerated with no serious adverse events or dose-dependent increases in adverse events observed. Clinical laboratory results, vital signs, and physical examination data did not reveal any safety signals. Dose-limiting toxicity was not observed; the maximum tolerated dose of UV-4 hydrochloride in humans has not yet been determined (>1000 mg). UV-4 was rapidly absorbed and distributed after dosing with the oral solution formulation used in this study. Median time to reach maximum plasma concentration ranged from 0.5–1 hour and appeared to be independent of dose. Exposure increased approximately in proportion with dose over the 333-fold dose range. UV-4 was quantifiable in pooled urine over the entire collection interval for all doses.Conclusions/SignificanceUV-4 is a host-targeted broad-spectrum antiviral drug candidate. At doses in humans up to 1000 mg there were no serious adverse events reported and no subjects were withdrawn from the study due to treatment-emergent adverse events. These data suggest that therapeutically relevant drug levels of UV-4 can be safely administered to humans and support further clinical development of UV-4 hydrochloride or other candidate antivirals in the iminosugar class.Trial registrationClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT02061358","term_id":"NCT02061358"}}NCT02061358 https://clinicaltrials.gov/ct2/show/{"type":"clinical-trial","attrs":{"text":"NCT02061358","term_id":"NCT02061358"}}NCT02061358.