Mapping metabolic fluxes in plant cells to understand carbon-nitrogen interactions and nitrogen storage and cycling

Plants provide commodities like food, fiber, fuel and chemicals. Understanding plant metabolism will help find genetic engineering targets that enhance production of these commodities. Interactions between the macronutrients - carbon (C) and nitrogen (N) determine growth and developmental functions in plants (Nunes-Nesi, Fernie, and Stitt 2010; Sakakibara, Takei, and Hirose 2006) and are regulated by complex mechanisms that need systems-level analyses. Metabolic fluxes, the rates of C flow in metabolic pathways, provide a system-wide view of metabolism and are quantified by steady state metabolic flux analysis (MFA) wherein isotopic tracers (13C, 15N) are fed to the cells and the resulting labeling patterns of biomass components are used to fit the fluxes. In this study we i) statistically designed isotope labeling experiments (ILEs) in silico to enhance accuracy of flux estimates through the pentose phosphate pathway (PPP) ii) conducted MFA on heterotrophic cell suspensions of Arabidopsis thaliana (Arabidopsis), a model plant, to investigate regulatory role of light in cell metabolism and iii) conducted MFA on cell suspensions of poplar (Populus tremula × Populus alba; clone N 717-B4), a potential biofuel crop, to understand C-N interactions. In silico label design studies determined that accuracy of flux estimates in the PPP improves by ILEs with 1,2-13C glucose and measuring labeling patterns of sugars, especially ribose. Metabolic fluxes, estimated by the designed ILEs on Arabidopsis cells, under continuous light or dark, showed negligible changes between treatments indicating that light does not regulate central carbon metabolism in heterotrophic Arabidopsis cells. The designed ILEs improved confidences of non-oxidative PPP flux estimates by 40-80% from previous studies (Masakapalli et al. 2009a). ILEs on poplar cell suspensions, grown in batch cultures, displayed unexpected back-mixing between unlabeled seed biomass and newly synthesized labeled biomass. Novel metabolic network models were developed that successfully account for observed back-mixing. ILEs on poplar cells, subjected to different C-N supply treatments to understand C-N interactions showed significant differences in phenylalanine labeling which may implicate role of flavonoid biosynthesis pathway in C-N interactions. Design of ILEs and subsequent improvement in flux estimates and the improvements in modeling metabolic networks are the novel contributions of this work

Mechanism (S) of Metal-Induced Apoptosis in Saccharomyces Cerevisiae

Heavy metals, such as copper and cadmium have been linked to a number of cellular dysfunctions in single and multicellular organisms that are associated with apoptosis. The yeast, Saccharomyces cerevisiae, provides a valuable model for elucidating apoptosis mechanisms, and this study extends that capability to Cu and Cd-induced apoptosis. We demonstrate that S. cerevisiae undergoes a glucose-dependent, programmed cell death in response to low cadmium concentrations, which is initiated within the first hour of Cd exposure. The response was associated with induction of the yeast caspase, Yca1p, and was abolished in YCA1∆ mutant. Other apoptotic markers, including sub-G1 DNA fragmentation and hyper-polarization of mitochondrial membranes, were also evident among Cd-exposed cells. We also show that low levels of copper can induce a similar apoptotic response in yeast within the first hour of exposure. Such cellular responses were verified by analyzing mitochondrial perturbation, generation of superoxide ions, activation of the yeast caspase1, and the eventual fragmentation of nuclear DNA (through TUNEL). In analyzing the response of yeast to the different metals, we also demonstrated that the metal-induced PCD is instigated through the sequential activity of at least two caspase-like proteins (i.e., Yca1 and Atg4), both of which appear to be in involved in the process of inducing mitochondrial stress. The additional caspase-like activity is shown to be derived from an enzyme involved in the latter stages of autophagy (Atg4), and provides an intriguing association of apoptosis with autophagy. Here we also demonstrate that metals such as copper and cadmium causes oxidative damage to mitochondrial proteins. Such oxidative attack is targeted and we show that oxidation of certain crucial proteins is required for apoptosis upon metal exposure. By showing that such targeted protein oxidation is dependent on YCA1 and ATG, we also confirm the finding that in yeast that have been exposed to a heavy metal, YCA1 and ATG are essential for damaging mitochondria and to initiate apoptosis. These novel findings highlight several new perspectives about the mechanism of metal-dependent apoptosis, while opening up future analyses to the power of the yeast model system

Synthesis and biological evaluation of novel pyrazole derivatives as urease inhibitors

Studies on enzyme inhibition remain an important area of pharmaceutical research since these studies have led to the discoveries of drugs useful in a variety of physiological conditions. The enzyme inhibitors can interact with enzymes and block their activity towards natural substrates. Urease inhibitors have recently attracted much attention as potential new anti-ulcer drugs. A series of novel substituted pyrazoles 8(a-j) has been synthesized by diazotization of fluoro chloro aniline (1) and the reaction of the corresponding diazonium salt solution (2) with ethyl cyanoacetate (3) to give the intermediate, ethyl 2-((3-chloro-4-fluorophenyl) diazenyl-2- cyanoacetate (4). The intermediate is then cyclised with chloroacetonitrile (5) using triethyl amine as the base to give the final compound, ethyl 4-amino-1-(3-chloro-4-flurophenyl)-5- cyano-1H-pyrazole-3-carboxylate (6). Nucleophilic substitution group is removed from the final compound and 8(a-j) derivatives have been synthesized. All the synthesized compounds were characterized by physical data (M.P. & TLC) and spectral Data (IR & 1H NMR). The synthesized compounds were evaluated for urease-inhibition activity. Molecular docking studies were carried out for these compounds with the enzyme urease. From the observations it has been noticed that some of the compounds possesses remarkable urease-inhibitory effect

Analysis of flow of polar and non polar incompressible ferrofluids

In this paper, flow between two parallel plates is analyzed for both polar and non polar ferrofluids. Velocity is obtained without pressure gradient for polar fluid and with pressure gradient for non polar fluid. The solution of the spin velocity is found in terms of applied magnetic field and magnetic flux density for polar fluid. Shear stress is calculated for both polar and non polar ferrofluid

SPOTR: Spatio-temporal Pose Transformers for Human Motion Prediction

3D human motion prediction is a research area of high significance and a challenge in computer vision. It is useful for the design of many applications including robotics and autonomous driving. Traditionally, autogregressive models have been used to predict human motion. However, these models have high computation needs and error accumulation that make it difficult to use them for realtime applications. In this paper, we present a non-autogressive model for human motion prediction. We focus on learning spatio-temporal representations non-autoregressively for generation of plausible future motions. We propose a novel architecture that leverages the recently proposed Transformers. Human motion involves complex spatio-temporal dynamics with joints affecting the position and rotation of each other even though they are not connected directly. The proposed model extracts these dynamics using both convolutions and the self-attention mechanism. Using specialized spatial and temporal self-attention to augment the features extracted through convolution allows our model to generate spatio-temporally coherent predictions in parallel independent of the activity. Our contributions are threefold: (i) we frame human motion prediction as a sequence-to-sequence problem and propose a non-autoregressive Transformer to forecast a sequence of poses in parallel; (ii) our method is activity agnostic; (iii) we show that despite its simplicity, our approach is able to make accurate predictions, achieving better or comparable results compared to the state-of-the-art on two public datasets, with far fewer parameters and much faster inference

HOMOTOPY ANALYSIS METHOD TO SOLVE BOUSSINESQ EQUATIONS

In this paper, Homotopy analysis method is applied to the nonlinear coupled differential equations of classical Boussinesq system. We have applied Homotopy analysis method (HAM) for the application problems in [1, 2, 3, 4]. We have also plotted Domb-Sykes plot for the region of convergence. We have applied Pade for the HAM series to identify the singularity and reflect it in the graph. The HAM is a analytical technique which is used to solve non-linear problems to generate a convergent series. HAM gives complete freedom to choose the initial approximation of the solution, it is the auxiliary parameter h which gives us a convenient way to guarantee the convergence of homotopy series solution. It seems that more artificial degrees of freedom implies larger possibility to gain better approximations by HAM

An Analysis of Internal Gravity Wave Tunnelling in a Stratified Region Along with Rotation

The internal gravity wave tunnelling in presence of earths rotation is studied for different density barriers. An exponential approximation used reveals the existence of evanescence in the barrier region which signifies the trapping of wave energy in the tunnelling region. The Transmission coefficients are computed for different density barriers and the comparative study shows that across the locally mixed region the transmission is enhanced. The asymptotic analysis of the transmission co-efficient using the rotational parameter reveals the convergence and the graphs shows that the transmission decreases continuously and leads to the non-rotating case. The results are compared with the non-rotations case and we observe that the evanescence caused by the rotation makes the waves travel more along the horizontal direction than in the vertical direction

Case report: Pregnancy After Vitrification of Biopsied Human Blastocysts Previously Frozen by the Slow Method

Pre-implantation genetic screening (PGS) with trophectoderm biopsy is an extremely powerful technique for the determination of embryos with a high implantation potential. Patients with cryopreserved embryos seeking PGS have had limited access to this treatment due to the need to thaw, biopsy, and refreeze these embryos. This is especially true for patients with embryos cryopreserved by the slow freeze technique due to the low survival rate after thawing. In this case report, we describe the application of refreezing with the vitrification technique to embryos thawed with the slow technique and biopsied for PGS. The patient had a total of 8 blastocysts thawed, biopsied, and refrozen with vitrification. Next Generation Sequencing (NGS) of these embryos revealed that 4 blastocysts were euploid. The patients achieved a pregnancy on the the first frozen embryo transfer procedure that terminated after 11 days. The second frozen embryo transfer procedure resulted in the live birth of a 3800g boy, demonstrating that thaw, biopsy, and refreeze are applicable to human reproduction even in cases where embryos have been cryopreserved by the slow techniqu

Igf2/H19 Imprinting Control Region (ICR): An Insulator or a Position-Dependent Silencer?

The imprinting control region (ICR) located far upstream of the H19 gene, in conjunction with enhancers, modulates the transcription of Igf2 and H19 genes in an allele-specific manner. On paternal inheritance, the methylated ICR silences the H19 gene and indirectly facilitates transcription from the distant Igf2 promoter, whereas on the maternal chromosome the unmethylated ICR, together with enhancers, activates transcription of the H19 gene and thereby contributes to the repression of Igf2. This repression of maternal Igf2 has recently been postulated to be due to a chromatin boundary or insulator function of the unmethylated ICR. Central to the insulator model is the site-specific binding of a ubiquitous nuclear factor CTCF which exhibits remarkable flexibility in functioning as transcriptional activator or silencer. We suggest that the ICR positioned close to the enhancers in an episomal context might function as a transcriptional silencer by virtue of interaction of CTCF with its modifiers such as SIN3A and histone deacetylases. Furthermore, a localised folded chromatin structure resulting from juxtaposition of two disparate regulatory sequences (enhancer ICR) could be the mechanistic basis of ICR-mediated position-dependent (ICR-promoter) transcriptional repression in transgenic Drosophila