Synthesis Of Other Proteins Research Articles

Improvement of grain sorghum nutritive properties using modern genetic and biotechnological methods

The paper presents a review of the studies on the use of genetic engineering and genome editing tools for improving nutritional properties of sorghum grain. As a result of experiments performed over the past 5-7 years by several research groups, the created transgenic lines carry genetic constructs for RNA silencing of different kafirin sub-classes (prolamins of sorghum grain). The CRISPR/Cas9 genome editing experiments have yielded mutants with deletions and insertions in the signal sequence of the gene encoding the 22 kDa α-kafirin in sorghum. These lines and mutants were characterized by improved in vitro digestibility of grain proteins, altered ultrastructure of protein bodies and an increased content of lysine. RNA silencing of α-kafirin increased the digestibility of proteins of both raw and cooked flour, while silencing of γ-kafirin led to improved digestibility of proteins of only raw flour. The lines with α-kafirin silencing have kernels with the floury endosperm type that discourages their direct commercial use because of fragility and reduced tolerance to fungal contamination; however, these lines can be used as donors of high digestibility trait when crossed with sorghum lines adapted to local conditions to improve their nutritional value. Kernels of the lines with γ-kafirin silencing may have different endosperm types: floury, vitreous, or a modified type with vitreous endosperm interspersed in the floury endosperm. This fact indicates the possibility of producing agronomically important sorghum lines with high kafirin digestibility and hard endosperm. The increased lysine level in kernels of sorghum lines with the suppressed synthesis of kafirins may be caused by rebalancing of protein synthesis in endosperm of developing kernels due to the synthesis of other proteins, including those with a higher content of essential amino acids. Alongside with improving the digestibility of kafirins, the genetic engineering approach allowed the creation of sorghum lines with a high content of provitamin A in grain and its increased stability during long-term storage. The results of these works show that it is promising to use RNA-interference and genome editing for creating sorghum lines with improved nutritional value of grain.

Total synthesis of TRADD death domain with arginine N-GlcNAcylation by hydrazide-based native chemical ligation

TNFR1-associated death domain protein (TRADD) with arginine N-GlcNAcylation is a novel and structurally unique posttranslational modification (PTM) glycoprotein that blocks the formation of death-inducing signaling complex (DISC), orchestrating host nuclear factor κB (NF-κB) signaling in entero-pathogenic Escherichia coli (EPEC)-infected cells. This particular glycosylated modification plays an extremely vital role for the effective colonization and pathogenesis of pathogens in the gut. Herein we describe the total synthesis of TRADD death domain (residues 195–312) with arginine235 N-GlcNAcylation (Arg-GlcNAc TRADD (195–312)). Two longish peptidyl fragments of the wild-type primary sequence were obtained by robust, microwave-assisted, highly efficient, solid-phase peptide synthesis (SPPS), the N-GlcNAcylated sector was built by total synthesis and attached specifically to resin-bound peptide with an unprotected ornithine residue via silver-promoted on-resin guanidinylation, Arg-GlcNAc TRADD (195–312) was constructed by hydrazide-based native chemical ligation (NCL). The facile synthetic strategy is expected to be generally applicable for the rapid synthesis of other proteins with Arg-GlcNAc modification and to pave the way for the related chemically biological study.

Temperature stress response of heat shock protein 90 (Hsp90) in the clam Paphia undulata

The Heat shock proteins (HSPs) are a group of molecular chaperones that play a crucial role in cell response to various stresses. A full-length cDNA of the heat shock protein 90 (PuHsp90) was cloned and sequenced from the clam Paphia undulata. Phylogenetic analysis revealed that PuHsp90 grouped with the Hsp90 from other metazoan species. Expression of PuHsp90 was highly detected in the gonad, followed by digest gland, gills and heart but was found poorly expressed in mantle, adductor muscle and hemocytes. After a heat shock stress at 32 °C up-regulation of PuHsp90 was detected in the mantle, adductor muscle, gills and hemocytes. Maximal expression levels occurred at 4 h after the heat shock and up-regulation is indicative of protein denaturation and of an increase in energy consumption. In contrast after the heat shock, PuHsp90 was continuously down-regulated in the digestive gland and in the gonad suggesting modifications of the biochemical pathways and energy budgets involved in the synthesis of other protein, such as catalase and of other Hsp proteins. These results reveal that PuHsp90 may play an important role in the clam response to a temperature stress.

Arresting frailty and sarcopenia in cirrhosis: Future prospects.

Arresting frailty and sarcopenia in cirrhosis: Future prospects.

Proteome changes in Garnacha Tintorera red grapes during post-harvest drying

Fruit raisining is accompanied by changes in composition due to the joint action of water loss and modifying enzymes. Cell walls in grape berries form a barrier against diffusion of aromatic and polyphenolic compounds that influence wine quality. In this work, fresh and post-harvest dried grape samples of Vitis vinifera L. Garnacha Tintorera variety were compared in terms of proteome. The main change observed during raisining was over-expression from 14 to 47% of proteins involved in pathogen- and stress-resistance mechanisms, whereas metabolism-related proteins or those involved in the transcription and synthesis of other proteins decreased, respectively, from 43 to 27% and from 29 to 13%. These changes are important in the functional qualities and stability of the sweet wines obtained, since affect interactions with other macromolecules or metabolites.

Less is more: Ebola virus surface glycoprotein expression levels regulate virus production and infectivity.

The Ebola virus (EBOV) surface glycoprotein (GP1,2) mediates host cell attachment and fusion and is the primary target for host neutralizing antibodies. Expression of GP1,2 at high levels disrupts normal cell physiology, and EBOV uses an RNA-editing mechanism to regulate expression of the GP gene. In this study, we demonstrate that high levels of GP1,2 expression impair production and release of EBOV virus-like particles (VLPs) as well as infectivity of GP1,2-pseudotyped viruses. We further show that this effect is mediated through two mechanisms. First, high levels of GP1,2 expression reduce synthesis of other proteins needed for virus assembly. Second, viruses containing high levels of GP1,2 are intrinsically less infectious, possibly due to impaired receptor binding or endosomal processing. Importantly, proteolysis can rescue the infectivity of high-GP1,2-containing viruses. Taken together, our findings indicate that GP1,2 expression levels have a profound effect on factors that contribute to virus fitness and that RNA editing may be an important mechanism employed by EBOV to regulate GP1,2 expression in order to optimize virus production and infectivity. The Ebola virus (EBOV), as well as other members of the Filoviridae family, causes severe hemorrhagic fever that is highly lethal, with up to 90% mortality. The EBOV surface glycoprotein (GP1,2) plays important roles in virus infection and pathogenesis, and its expression is tightly regulated by an RNA-editing mechanism during virus replication. Our study demonstrates that the level of GP1,2 expression profoundly affects virus particle production and release and uncovers a new mechanism by which Ebola virus infectivity is regulated by the level of GP1,2 expression. These findings extend our understanding of EBOV infection and replication in adaptation of host environments, which will aid the development of countermeasures against EBOV infection.

In vitro Studies of the Antibody Response: Antibodies of Different Specificity are Made in Different Populations of Cells.

In vitro Studies of the Antibody Response: Antibodies of Different Specificity are Made in Different Populations of Cells.

Monitoring Collagen Synthesis in Fibroblasts Using Fluorescently Labeled tRNA Pairs

There is a critical need for techniques that directly monitor protein synthesis within cells isolated from normal and diseased tissue. Fibrotic disease, for which there is no drug treatment, is characterized by the overexpression of collagens. Here, we use a bioinformatics approach to identify a pair of glycine and proline isoacceptor tRNAs as being specific for the decoding of collagen mRNAs, leading to development of a FRET-based approach, dicodon monitoring of protein synthesis (DiCoMPS), that directly monitors the synthesis of collagen. DiCoMPS aimed at detecting collagen synthesis will be helpful in identifying novel anti-fibrotic compounds in cells derived from patients with fibrosis of any etiology, and, suitably adapted, should be widely applicable in monitoring the synthesis of other proteins in cells.

Genome-wide analysis of the heat stress response in Zebu (Sahiwal) cattle

Environmental-induced hyperthermia compromises animal production with drastic economic consequences to global animal agriculture and jeopardizes animal welfare. Heat stress is a major stressor that occurs as a result of an imbalance between heat production within the body and its dissipation and it affects animals at cellular, molecular and ecological levels. The molecular mechanism underlying the physiology of heat stress in the cattle remains undefined. The present study sought to evaluate mRNA expression profiles in the cattle blood in response to heat stress. In this study we report the genes that were differentially expressed in response to heat stress using global scale genome expression technology (Microarray). Four Sahiwal heifers were exposed to 42°C with 90% humidity for 4h followed by normothermia. Gene expression changes include activation of heat shock transcription factor 1 (HSF1), increased expression of heat shock proteins (HSP) and decreased expression and synthesis of other proteins, immune system activation via extracellular secretion of HSP. A cDNA microarray analysis found 140 transcripts to be up-regulated and 77 down-regulated in the cattle blood after heat treatment (P<0.05). But still a comprehensive explanation for the direction of fold change and the specific genes involved in response to acute heat stress still remains to be explored. These findings may provide insights into the underlying mechanism of physiology of heat stress in cattle. Understanding the biology and mechanisms of heat stress is critical to developing approaches to ameliorate current production issues for improving animal performance and agriculture economics.

Ghrelin Gene Products and the Regulation of Food Intake and Gut Motility

A breakthrough using "reverse pharmacology" identified and characterized acyl ghrelin from the stomach as the endogenous cognate ligand for the growth hormone (GH) secretagogue receptor (GHS-R) 1a. The unique post-translational modification of O-n-octanoylation at serine 3 is the first in peptide discovery history and is essential for GH-releasing ability. Des-acyl ghrelin, lacking O-n-octanoylation at serine 3, is also produced in the stomach and remains the major molecular form secreted into the circulation. The third ghrelin gene product, obestatin, a novel 23-amino acid peptide identified from rat stomach, was found by comparative genomic analysis. Three ghrelin gene products actively participate in modulating appetite, adipogenesis, gut motility, glucose metabolism, cell proliferation, immune, sleep, memory, anxiety, cognition, and stress. Knockdown or knockout of acyl ghrelin and/or GHS-R1a, and overexpression of des-acyl ghrelin show benefits in the therapy of obesity and metabolic syndrome. By contrast, agonism of acyl ghrelin and/or GHS-R1a could combat human anorexia-cachexia, including anorexia nervosa, chronic heart failure, chronic obstructive pulmonary disease, liver cirrhosis, chronic kidney disease, burn, and postsurgery recovery, as well as restore gut dysmotility, such as diabetic or neurogenic gastroparesis, and postoperative ileus. The ghrelin acyl-modifying enzyme, ghrelin O-Acyltransferase (GOAT), which attaches octanoate to serine-3 of ghrelin, has been identified and characterized also from the stomach. To date, ghrelin is the only protein to be octanylated, and inhibition of GOAT may have effects only on the stomach and is unlikely to affect the synthesis of other proteins. GOAT may provide a critical molecular target in developing novel therapeutics for obesity and type 2 diabetes.

Parasitic castration of Plutella xylostella larvae induced by polydnaviruses and venom of Cotesia vestalis and Diadegma semiclausum

In the present study, we used gamma-ray to irradiate the female parasitoids to make wasp eggs infertile, resulting in pseudoparasitization, which allowed the analysis of maternal secretions such as polydnaviruses (PDVs) and venom in the absence of larval secretions or teratocytes by the growing parasitoids. We then investigated the spermatogenesis and components of testicular proteins of male Plutella xylostella larvae pseudoparasitized by two endoparasitoids (Cotesia vestalis and Diadegma semiclausum). The results showed that pseudoparasitism by the two endoparasitoids at the early third instar host larvae both induced smaller testes in size than those of nonparasitized host larvae. Both of them caused parasitic castration, and the degree of castration is almost as severe as in naturally parasitized hosts. This suggested that PDVs and venom played a major role in the degeneration of host testes. There are significant differences in the degree of castration induced by the two endoparasitoids, with respect to testicular growth, testicular protein concentrations, and histological changes of germ cells. Cotesia vestalis bracovirus always has a significantly stronger effect on host testicular growth and development than D. semiclausum ichnovirus. SDS-PAGE analysis indicated that synthesis of P 65 and P 67 proteins were clearly inhibited in testes of hosts that were pseudoparasitized by C. vestalis while reduction in synthesis of other proteins was not evident.

Globin-methionine complexes formed during labelling studies.

In implementing globin synthetic measurements using our recently described technique of Cellogel separation of 35S-methionine-labelled globin chain we sometimes encountered aberrant results. We discovered the presence of prominent radioactive bands trailing both the alpha and beta globin chains. Such bands were not present when 3H-leucine served as a label. These bands were not due to contaminants in the 35S-methionine. Their formation could be prevented by including in the incubating mixture 1 mM cystine, cysteine or homocysteine. The bands could not be removed by trichloracetic acid or acid acetone precipitation but did disappear almost entirely after prolonged dialysis against a urea-containing buffer. The trailing bands appear to represent the result of the firm, but not covalent, binding of methionine to globin during the protein synthetic process. Since methionine is commonly used as a label for newly synthesized protein, this phenomenon may be important not only in the investigation of globin synthesis, but also in the study of the synthesis of other proteins.

English

&nbsp; Nearly one third of protein in the body consists of collagen. It plays a key role in providing the structural scaffolding for cells, tissues, and organs. With available literature, we show that prolonged exposure of cultures of human connective-tissue cells to ascorbate (vitamin C) induces an eight-fold increase in the synthesis of collagen with no increase in the rate of synthesis of other proteins. Making advantage of increased degree of polarization due to enhanced rate of collagen biosynthesis in the presence of vitamin C, we propose a better imaging modality for diagnosing thermal damage in human tissues. &nbsp; Key words:&nbsp;Collagen, vitamin C, triple helix, birefringence, optical coherence tomography.

Effects of dexamethasone and dibutyryl cAMP on stanniocalcin-1 mRNA expression in rat primary Sertoli and Leydig cells

In this study, STC1 expression levels were determined at various postnatal developmental stages in rat ovaries and testes. The expression pattern of STC1 was found to be sexually dimorphic. In addition we examined the effect of exogenous glucocorticoids, dexamethasone (DEX), testosterone, follicle stimulating hormone (FSH) and dibutyryl cAMP (dbcAMP) on STC1 expression in rat primary Sertoli cell cultures. Our results showed that DEX (via glucocorticoids receptor) stimulated while dbcAMP inhibited STC1 expression in the Sertoli cells. Moreover testosterone and FSH treatment had no significant effect on STC1 expression in the cells. The inhibitory effect of dbcAMP to STC1 expression was consistently demonstrated in rat primary Leydig cell cultures. In human chorinoic gonadotropin (hCG) or dbcAMP treated Leydig cells, a significant reduction of STC1 expression was accompanied with an induction of testosterone secretion. The actions of hCG/dbcAMP were mediated by PKA, as the effect can be rescued by a co-treatment with H89. Using the Sertoli cell model, the modulation of STC1 expression by DEX or dbcAMP was found to be regulated at the transcriptional level as actinomycin D treatment significantly reduced the respective effect. In addition, both DEX and dbcAMP mediated effects were found to be abolished by cycloheximide co-treatment. These indicated that the regulation in the both treatments were indirect and may require prior protein synthesis. We suggested that the de novo synthesis of other protein(s) and mRNA may be involved in the regulation of the steady-state levels of STC1 mRNA in the treatments. Taken together, this study provides the first evidence of the regulation of STC1 expression in the male reproductive system.

Experience of vitamin K2 in Thailand

Vitamin K(2) has dual actions, stimulates osteoblastic functions, for synthesis of osteocalcin, osteonectin and other matrix bone proteins, in addition, new finding, in stem cell culture found osteoblast producing gene expression of collagen type 1, the other action, vitamin K(2) contains mild antiresorpion by inducing the osteoclastic apoptosis. Our study found that postmenopausal and elderly women have high risk of vitamin K(2) deficiency, comparing to the normal value of young, reproductive females. The efficacy of vitamin K(2) will be fulfill benefit after 6 months of administration, prolong use will enhance of bone quality that prevent fracture.

Meiotic Resumption and Spindle Formation of Pig Oocytes

ABSTRACT Fully grown mammalian oocytes arrested at prophase I resume meiosis after gonadotropic stimulation. Oocytes undergo a series of changes including chromosome condensation, nucleolus disassembly, germinal vesicle breakdown (GVBD), and spindle formation. The mouse is the best model for studying the molecular mechanisms underlying the maturation of mammalian oocytes. However, some of the maturational events are different from those in other mammalian species. To study these events, we use pig ovaries, which are available as a byproduct from local slaughterhouses. It has long been known that pig oocytes have a dependence on de novo protein synthesis for GVBD, whereas GVBD in mouse oocytes occurs independently of protein synthesis. The reason seems to be the lack of Cyclin B1 molecules in pig GV-oocytes, although the synthesis of other protein(s) may be required for the GVBD. In mouse oocytes, the spindle is formed through the action of cytoplasmic microtubule organizing centers (MTOCs), and the oocyte...

Light and Metabolic Signals Control the Selective Degradation of Sucrose Synthase in Maize Leaves during Deetiolation

The content and activity of Suc (Suc) synthase (SUS) protein is high in sink organs but low in source organs. In this report, we examined light and metabolic signals regulating SUS protein degradation in maize (Zea mays) leaves during deetiolation. We found that SUS protein accumulated in etiolated leaves of the dark-grown seedlings but was rapidly degraded upon exposure to white, blue, or red light. This occurred concurrent with the accumulation of photosynthetic enzymes, such as Rubisco and Rubisco activase, and enzymes of Suc biosynthesis such as Suc-phosphate synthase. Deetiolation-induced SUS degradation was not inhibited by the proteasome inhibitor MG132. Moreover, neither full-length nor truncated SUS phosphorylated at the serine-170 site was found in the crude 26S proteasome fraction (150,000g postmicrosomal pellet) isolated in the presence of MG132. However, SUS degradation was strongly inhibited by feeding cycloheximide or amino acids to detached leaves, while Suc feeding had no effect. Of the amino acids tested, exogenous glutamate had the greatest effect. Collectively, these results demonstrate that SUS protein degradation during deetiolation: (1) is selective; (2) can be triggered by either blue- or red light-mediated signaling pathways; (3) does not involve the 26S proteasome; and (4) is inhibited by free amino acids. These findings suggest that SUS degradation is important to supply residues for the synthesis of other proteins required for autotrophic metabolism.

The lom gene of bacteriophage lambda is involved in Escherichia coli K12 adhesion to human buccal epithelial cells.

The role of the lom gene of bacteriophage lambda in adhesion of Escherichia coli to human buccal epithelial cells (HBC) was studied testing the adherence of lamda lom+ and lambda lom::TnphoA E. coli lysogens. lambda lom+ prophage increased 50% E. coli adhesion. This effect was not observed with lambda lom::TnphoA. These results suggest that the normal Lom protein participates directly in adhesion or regulates the synthesis of other protein(s), which may be involved in adhesion.

Enzymatic cleavage as a processing step in the maturation of Muc4/sialomucin complex

Cleavage of Muc4/SMC precursor into two subunits is an essential processing step for maturation and occurs within a GD-PH sequence. Recent evidence indicates that cleavage of the precursor of gel-forming mucin MUC2 within the same tetrapeptide sequence occurs by a non-enzymatic, autocatalytic cleavage at low pH, and in cells in the late secretory pathway. Here we provide evidence that the cleavage step of Muc4/SMC processing occurs by a proteolytic mechanism. First, processing of Muc4/SMC precursor to ASGP-2 was inhibited in the presence of the mechanism-based serine protease inhibitor, Pefabloc SC, under conditions that did not block synthesis of other proteins. This inhibition led to an increased level of the precursor. Second, neutralization of the acidic environment of the late secretory pathway with NH4Cl did not inhibit cleavage of Muc4/SMC precursor. These results indicate that the two mucins can be processed by cleavage at the same peptide site by different mechanisms.

Expression of Perchloric Acid Soluble 14KDA Protein in Mouse Brain

Perchloric acid soluble protein (PSP), a member of the group of non-histone chromatin protein, consists of two identical subunits with a molecular mass of 14 kDa. It is highly conserved throughout the animal kingdom from prokaryotes to eukaryotes, reflecting its involvement in basic cellular regulation. It acts as an inhibitor for elongation step of protein synthesis. PSP expression has been reported to be tissue-specific, although it is predominantly present in liver and kidney cells. In this study, we report the expression of PSP in mice brain and compare it with liver. The expression of PSP was found to be much lower in brain than in liver, suggesting a tissue-dependent quantitative variation. The level of PSP mRNA was one-third and that of protein one-tenth in the brain as compared to the liver. Such discrepancy suggests that the brain PSP mRNA is less stable and its small fraction is translated leading to higher synthesis of other proteins in the brain. doi: 10.5214/ans.0972.7531.2005.120203