Neurospora crassa transcriptomics reveals oxidative stress and plasma membrane homeostasis biology genes as key targets in response to chitosan

PubMed Central

Lopez-Moya, Federico; Kowbel, David; Nueda, Ma José; Palma-Guerrero, Javier; Glass, N. Louise; Lopez-Llorca, Luis Vicente

2016-01-01

Chitosan is a natural polymer with antimicrobial activity. Chitosan causes plasma membrane permeabilization and induction of intracellular reactive oxygen species (ROS) in Neurospora crassa. We have determined the transcriptional profile of N. crassa to chitosan and identified the main gene targets involved in the cellular response to this compound. Global network analyses showed membrane, transport and oxidoreductase activity as key nodes affected by chitosan. Activation of oxidative metabolism indicates the importance of ROS and cell energy together with plasma membrane homeostasis in N. crassa response to chitosan. Deletion strain analysis of chitosan susceptibility pointed, NCU03639 encoding a class 3 lipase, involved in plasma membrane repair by lipid replacement and NCU04537 a MFS monosaccharide transporter related with assimilation of simple sugars, as main gene targets of chitosan. NCU10521, a glutathione S-transferase-4 involved in the generation of reducing power for scavenging intracellular ROS is also a determinant chitosan gene target. Ca2+ increased tolerance to chitosan in N. crassa. Growth of NCU10610 (fig 1 domain) and SYT1 (a synaptotagmin) deletion strains was significantly increased by Ca2+ in presence of chitosan. Both genes play a determinant role in N. crassa membrane homeostasis. Our results are of paramount importance for developing chitosan as antifungal. PMID:26694141

Yarrowia lipolytica possesses two plasma membrane alkali metal cation/H+ antiporters with different functions in cell physiology.

PubMed

Papouskova, Klara; Sychrova, Hana

2006-04-03

The family of Nha antiporters mediating the efflux of alkali metal cations in exchange for protons across the plasma membrane is conserved in all yeast species. Yarrowia lipolytica is a dimorphic yeast, phylogenetically very distant from the model yeast Saccharomyces cerevisiae. A search in its sequenced genome revealed two genes (designated as YlNHA1 and YlNHA2) with homology to the S. cerevisiae NHA1 gene, which encodes a plasma membrane alkali metal cation/H+ antiporter. Upon heterologous expression of both YlNHA genes in S. cerevisiae, we showed that Y. lipolytica antiporters differ not only in length and sequence, but also in their affinity for individual substrates. While the YlNha1 protein mainly increased cell tolerance to potassium, YlNha2p displayed a remarkable transport capacity for sodium. Thus, Y. lipolytica is the first example of a yeast species with two plasma membrane alkali metal cation/H+ antiporters differing in their putative functions in cell physiology; cell detoxification vs. the maintenance of stable intracellular pH, potassium content and cell volume.

The Complete Set of Genes Encoding Major Intrinsic Proteins in Arabidopsis Provides a Framework for a New Nomenclature for Major Intrinsic Proteins in Plants1

PubMed Central

Johanson, Urban; Karlsson, Maria; Johansson, Ingela; Gustavsson, Sofia; Sjövall, Sara; Fraysse, Laure; Weig, Alfons R.; Kjellbom, Per

2001-01-01

Major intrinsic proteins (MIPs) facilitate the passive transport of small polar molecules across membranes. MIPs constitute a very old family of proteins and different forms have been found in all kinds of living organisms, including bacteria, fungi, animals, and plants. In the genomic sequence of Arabidopsis, we have identified 35 different MIP-encoding genes. Based on sequence similarity, these 35 proteins are divided into four different subfamilies: plasma membrane intrinsic proteins, tonoplast intrinsic proteins, NOD26-like intrinsic proteins also called NOD26-like MIPs, and the recently discovered small basic intrinsic proteins. In Arabidopsis, there are 13 plasma membrane intrinsic proteins, 10 tonoplast intrinsic proteins, nine NOD26-like intrinsic proteins, and three small basic intrinsic proteins. The gene structure in general is conserved within each subfamily, although there is a tendency to lose introns. Based on phylogenetic comparisons of maize (Zea mays) and Arabidopsis MIPs (AtMIPs), it is argued that the general intron patterns in the subfamilies were formed before the split of monocotyledons and dicotyledons. Although the gene structure is unique for each subfamily, there is a common pattern in how transmembrane helices are encoded on the exons in three of the subfamilies. The nomenclature for plant MIPs varies widely between different species but also between subfamilies in the same species. Based on the phylogeny of all AtMIPs, a new and more consistent nomenclature is proposed. The complete set of AtMIPs, together with the new nomenclature, will facilitate the isolation, classification, and labeling of plant MIPs from other species. PMID:11500536

Maternal Betaine Supplementation during Gestation Enhances Expression of mtDNA-Encoded Genes through D-Loop DNA Hypomethylation in the Skeletal Muscle of Newborn Piglets.

PubMed

Jia, Yimin; Song, Haogang; Gao, Guichao; Cai, Demin; Yang, Xiaojing; Zhao, Ruqian

2015-11-25

Betaine has been widely used in animal and human nutrition to promote muscle growth and performance, yet it remains unknown whether maternal betaine supplementation during gestation affects the metabolic characteristics of neonatal skeletal muscles. In the present study, feeding sows with betaine-supplemented diets throughout gestation significantly upregulated the expression of mtDNA-encoded OXPHOS genes (p < 0.05), including COX1, COX2, and ND5, in the muscle of newborn piglets, which was associated with enhanced mitochondrial COX enzyme activity (p < 0.05). Concurrently, maternal betaine supplementation increased the plasma betaine concentration and muscle expression of methyl transfer enzymes (p < 0.05), BHMT and GNMT, in offspring piglets. Nevertheless, Dnmt3a was downregulated at the level of both mRNA and protein, which was associated with a hypomethylated mtDNA D-loop region (p < 0.05). These results suggest that maternal betaine supplementation during gestation enhances expression of mtDNA-encoded genes through D-loop DNA hypomethylation in the skeletal muscle of newborn piglets.

A heteromeric potassium channel involved in the modulation of the plasma membrane potential is essential for the survival of African trypanosomes.

PubMed

Steinmann, Michael E; González-Salgado, Amaia; Bütikofer, Peter; Mäser, Pascal; Sigel, Erwin

2015-08-01

Discovery of novel drug targets may lead to improved treatment of trypanosomiasis. We characterize here 2 gene products of Trypanosoma brucei that are essential for the growth of bloodstream form (BSF) parasites, as shown by RNA interference (RNAi)-mediated down-regulation of the individual mRNAs. The primary sequences of the 2 proteins--protein encoded by gene Tb927.1.4450 (TbK1) and protein encoded by gene Tb927.9.4820 (TbK2)--indicate that both belong to the family of putative, Ca(2+)-activated potassium channels. The proteins were expressed in Xenopus laevis oocytes and their functions investigated by use of electrophysiological techniques. Only combined expression of TbK1 and TbK2 results in the formation of sizeable currents, indicating that these proteins probably assemble into a heteromeric ion channel. The current mediated by this channel shows little time and voltage dependence and displays a permeability ratio of K(+)/Na(+) of >20. The known potassium channel blocker barium inhibits this channel with a half-maximal inhibitory concentration (IC50) of 98 ± 15 μM. The membrane potential of trypanosomes was measured with a fluorescent dye. Individual RNAi-mediated down-regulation of TbK1 or TbK2 eliminates a potassium conductance in the plasma membrane of BSF. Thus, this heteromeric potassium channel is involved in the modulation of the plasma membrane potential and represents a novel drug target in T. brucei. © FASEB.

High interpatient variability of raltegravir CSF concentrations in HIV-positive patients: a pharmacogenetic analysis.

PubMed

Calcagno, Andrea; Cusato, Jessica; Simiele, Marco; Motta, Ilaria; Audagnotto, Sabrina; Bracchi, Margherita; D'Avolio, Antonio; Di Perri, Giovanni; Bonora, Stefano

2014-01-01

To analyse the determinants of raltegravir CSF penetration, including the pharmacogenetics of drug transporters located at the blood-brain barrier or blood-CSF barrier. Plasma and CSF raltegravir concentrations were determined by a validated HPLC coupled with mass spectrometry method in adults on raltegravir-based combination antiretroviral therapy undergoing a lumbar puncture. Single nucleotide polymorphisms in the genes encoding drugs transporters (ABCB1 3435, SLCO1A2, ABCC2 and SLC22A6) and the gene encoding hepatocyte nuclear factor 4 α (HNF4α) were determined by real-time PCR. In 41 patients (73.2% male, 95.1% Caucasians), the median raltegravir plasma and CSF concentrations were 165 ng/mL (83-552) and 31 ng/mL (21-56), respectively. CSF-to-plasma ratios (CPRs) ranged from 0.005 to 1.33 (median 0.20, IQR 0.04-0.36). Raltegravir trough CSF concentrations (n = 35) correlated with raltegravir plasma levels (ρ = 0.395, P = 0.019); CPRs were higher in patients with blood-brain barrier damage (0.47 versus 0.18, P = 0.02). HNF4α 613 CG genotype carriers had lower trough CSF concentrations (20 versus 37 ng/mL, P = 0.03) and CPRs (0.12 versus 0.27, P = 0.02). Following multivariate linear regression analysis, the CSF-to-serum albumin ratio was the only independent predictor of raltegravir penetration into the CSF. Raltegravir penetration into the CSF shows a large interpatient variability, although CSF concentrations were above the wild-type IC50 in all patients (and above IC95 in 28.6%). In this cohort, blood-brain barrier permeability is the only independent predictor of raltegravir CPR. The impact of single nucleotide polymorphisms in selected genes on raltegravir penetration warrants further studies.

Transport and metabolism of glycerophosphodiesters produced through phospholipid deacylation.

PubMed

Patton-Vogt, Jana

2007-03-01

Phospholipid deacylation results in the formation of glycerophosphodiesters and free fatty acids. In Saccharomyces cerevisiae, four gene products with phospholipase B (deacylating) activity have been characterized (PLB1, PLB2, PLB3, NTE1), and those activities account for most, if not all, of the glycerophosphodiester production observed to date. The glycerophosphodiesters themselves are hydrolyzed into glycerol-3-phosphate and the corresponding alcohol by glycerophosphodiester phosphodiesterases. Although only one glycerophosphodiester phosphodiesterase-encoding gene (GDE1) has been characterized in S. cerevisiae, others certainly exist. Both internal and external glycerophosphodiesters (primarily glycerophosphocholine and glycerophosphoinositol) are formed as a result of phospholipid turnover in S. cerevisiae. A permease encoded by the GIT1 gene imports extracellular glycerophosphodiesters across the plasma membrane, where their hydrolytic products can provide crucial nutrients such as inositol, choline, and phosphate to the cell. The importance of this metabolic pathway in various aspects of S. cerevisiae cell physiology is being explored.

Trypanosoma cruzi H+-ATPase 1 (TcHA1) and 2 (TcHA2) genes complement yeast mutants defective in H+ pumps and encode plasma membrane P-type H+-ATPases with different enzymatic properties.

PubMed

Luo, Shuhong; Scott, David A; Docampo, Roberto

2002-11-15

Previous studies in Trypanosoma cruzi have shown that intracellular pH homeostasis requires ATP and is affected by H(+)-ATPase inhibitors, indicating a major role for ATP-driven proton pumps in intracellular pH control. In the present study, we report the cloning and sequencing of a pair of genes linked in tandem (TcHA1 and TcHA2) in T. cruzi which encode proteins with homology to fungal and plant P-type proton-pumping ATPases. The genes are expressed at the mRNA level in different developmental stages of T. cruzi: TcHA1 is expressed maximally in epimastigotes, whereas TcHA2 is expressed predominantly in trypomastigotes. The proteins predicted from the nucleotide sequence of the genes have 875 and 917 amino acids and molecular masses of 96.3 and 101.2 kDa, respectively. Full-length TcHA1 and an N-terminal truncated version of TcHA2 complemented a Saccharomyces cerevisiae strain deficient in P-type H(+)-ATPase activity, the proteins localized to the yeast plasma membrane, and ATP-driven proton pumping could be detected in proteoliposomes reconstituted from plasma membrane purified from transfected yeast. The reconstituted proton transport activity was reduced by inhibitors of P-type H(+)-ATPases. C-terminal truncation did not affect complementation of mutant yeast, suggesting the lack of C-terminal autoinhibitory domains in these proteins. ATPase activity in plasma membrane from TcHA1- and (N-terminal truncated) TcHA2-transfected yeast was inhibited to different extents by vanadate, whereas the latter yeast strain was more resistant to extremes of pH, suggesting that the native proteins may serve different functions at different stages in the T. cruzi life cycle.

Gene encoding prolactin in cinnamon clownfish Amphiprion melanopus and its expression upon acclimation to low salinities

PubMed Central

2013-01-01

Background Prolactin (PRL) is a key hormone for osmoregulation in fish. Levels of PRL in the pituitary gland and plasma ion composition of clownfish seem to change to regulate their hydromineral balance during adaptation to waters of different salinities. In order to understand osmoregulatory mechanism and its association with growth performance and PRL in fish, the gene encoding PRL and its expression level in cinnamon clownfish Amphiprion melanopus upon acclimation to low salinity was analyzed. Results The PRL gene of A. melanopus encoded a protein of 212 amino acid residues comprised of a putative signal peptide of 24 amino acids and a mature protein of 188 amino acids. Analysis of growth performance under different salinities of 34, 25, 15, and 10 ppt indicated that cinnamon clownfish could survive under salinities as low as 10 ppt. A higher rate of growth was observed at the lower salinities as compared to that of 34 ppt. Upon shifting the salinity of the surrounding water from 34 ppt to 15 ppt, the level of the PRL transcripts gradually increased to reach the peak level until 24 h of acclimation at 15 ppt, but decreased back as adaptation continued to 144 h. In contrast, levels of plasma Na+, Cl-, and osmolality decreased at the initial stage (4–8 h) of acclimation at 15 pt but increased back as adaptation continued till 144 h. Conclusion Cinnamon clownfish could survive under salinities as low as 10 ppt. Upon shifting the salinity of the surrounding water from 34 ppt to 15 ppt, the level of the PRL transcripts gradually increased during the initial stage of acclimation but decreased back to the normal level as adaptation continued. An opposite pattern of changes - decrease at the beginning followed by an increase - in the levels of plasma Na+, Cl-, and osmolality was found upon acclimation to low salinity. The results suggest an involvement of PRL in the processes of osmoregulation and homeostasis in A. melanopus. PMID:23276106

Gene encoding prolactin in cinnamon clownfish Amphiprion melanopus and its expression upon acclimation to low salinities.

PubMed

Noh, Gyeong Eon; Rho, Sum; Chang, Yong Jin; Min, Byung Hwa; Kim, Jong-Myoung

2013-01-01

Prolactin (PRL) is a key hormone for osmoregulation in fish. Levels of PRL in the pituitary gland and plasma ion composition of clownfish seem to change to regulate their hydromineral balance during adaptation to waters of different salinities. In order to understand osmoregulatory mechanism and its association with growth performance and PRL in fish, the gene encoding PRL and its expression level in cinnamon clownfish Amphiprion melanopus upon acclimation to low salinity was analyzed. The PRL gene of A. melanopus encoded a protein of 212 amino acid residues comprised of a putative signal peptide of 24 amino acids and a mature protein of 188 amino acids. Analysis of growth performance under different salinities of 34, 25, 15, and 10 ppt indicated that cinnamon clownfish could survive under salinities as low as 10 ppt. A higher rate of growth was observed at the lower salinities as compared to that of 34 ppt. Upon shifting the salinity of the surrounding water from 34 ppt to 15 ppt, the level of the PRL transcripts gradually increased to reach the peak level until 24 h of acclimation at 15 ppt, but decreased back as adaptation continued to 144 h. In contrast, levels of plasma Na+, Cl-, and osmolality decreased at the initial stage (4-8 h) of acclimation at 15 pt but increased back as adaptation continued till 144 h. Cinnamon clownfish could survive under salinities as low as 10 ppt. Upon shifting the salinity of the surrounding water from 34 ppt to 15 ppt, the level of the PRL transcripts gradually increased during the initial stage of acclimation but decreased back to the normal level as adaptation continued. An opposite pattern of changes - decrease at the beginning followed by an increase - in the levels of plasma Na+, Cl-, and osmolality was found upon acclimation to low salinity. The results suggest an involvement of PRL in the processes of osmoregulation and homeostasis in A. melanopus.

NaCl regulation of plasma membrane H(+)-ATPase gene expression in a glycophyte and a halophyte.

PubMed

Niu, X; Narasimhan, M L; Salzman, R A; Bressan, R A; Hasegawa, P M

1993-11-01

NaCl regulation of plasma membrane H(+)-ATPase gene expression in the glycophyte tobacco (Nicotiana tabacum L. var Wisconsin 38) and the halophyte Atriplex nummularia L. was evaluated by comparison of organ-specific mRNA abundance using homologous cDNA probes encoding the ATPases of the respective plants. Accumulation of mRNA was induced by NaCl in fully expanded leaves and in roots but not in expanding leaves or stems. The NaCl responsiveness of the halophyte to accumulate plasma membrane H(+)-ATPase mRNA in roots was substantially greater than that of the glycophyte. Salt-induced transcript accumulation in A. nummularia roots was localized by in situ hybridization predominantly to the elongation zone, but mRNA levels also increased in the zone of differentiation. Increased message accumulation in A. nummularia roots could be detected within 8 h after NaCl (400 mM) treatment, and maximal levels were severalfold greater than in roots of untreated control plants. NaCl-induced plasma membrane H(+)-ATPase gene expression in expanded leaves and roots presumably indicates that these organs require increased H(+)-electrochemical potential gradients for the maintenance of plant ion homeostasis for salt adaptation. The greater capacity of the halophyte to induce plasma membrane H(+)-ATPase gene expression in response to NaCl may be a salt-tolerance determinant.

NaCl regulation of plasma membrane H(+)-ATPase gene expression in a glycophyte and a halophyte.

PubMed Central

Niu, X; Narasimhan, M L; Salzman, R A; Bressan, R A; Hasegawa, P M

1993-01-01

NaCl regulation of plasma membrane H(+)-ATPase gene expression in the glycophyte tobacco (Nicotiana tabacum L. var Wisconsin 38) and the halophyte Atriplex nummularia L. was evaluated by comparison of organ-specific mRNA abundance using homologous cDNA probes encoding the ATPases of the respective plants. Accumulation of mRNA was induced by NaCl in fully expanded leaves and in roots but not in expanding leaves or stems. The NaCl responsiveness of the halophyte to accumulate plasma membrane H(+)-ATPase mRNA in roots was substantially greater than that of the glycophyte. Salt-induced transcript accumulation in A. nummularia roots was localized by in situ hybridization predominantly to the elongation zone, but mRNA levels also increased in the zone of differentiation. Increased message accumulation in A. nummularia roots could be detected within 8 h after NaCl (400 mM) treatment, and maximal levels were severalfold greater than in roots of untreated control plants. NaCl-induced plasma membrane H(+)-ATPase gene expression in expanded leaves and roots presumably indicates that these organs require increased H(+)-electrochemical potential gradients for the maintenance of plant ion homeostasis for salt adaptation. The greater capacity of the halophyte to induce plasma membrane H(+)-ATPase gene expression in response to NaCl may be a salt-tolerance determinant. PMID:8022933

Anion channels: master switches of stress responses.

PubMed

Roelfsema, M Rob G; Hedrich, Rainer; Geiger, Dietmar

2012-04-01

During stress, plant cells activate anion channels and trigger the release of anions across the plasma membrane. Recently, two new gene families have been identified that encode major groups of anion channels. The SLAC/SLAH channels are characterized by slow voltage-dependent activation (S-type), whereas ALMT genes encode rapid-activating channels (R-type). Both S- and R-type channels are stimulated in guard cells by the stress hormone ABA, which leads to stomatal closure. Besides their role in ABA-dependent stomatal movement, anion channels are also activated by biotic stress factors such as microbe-associated molecular patterns (MAMPs). Given that anion channels occur throughout the plant kingdom, they are likely to serve a general function as master switches of stress responses. Copyright © 2012 Elsevier Ltd. All rights reserved.

Neurospora crassa transcriptomics reveals oxidative stress and plasma membrane homeostasis biology genes as key targets in response to chitosan

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lopez-Moya, Federico; Kowbel, David; Nueda, Ma Jose

Chitosan is a natural polymer with antimicrobial activity. Chitosan causes plasma membrane permeabilization and induction of intracellular reactive oxygen species (ROS) in Neurospora crassa. In this paper, we have determined the transcriptional profile of N. crassa to chitosan and identified the main gene targets involved in the cellular response to this compound. Global network analyses showed membrane, transport and oxidoreductase activity as key nodes affected by chitosan. Activation of oxidative metabolism indicates the importance of ROS and cell energy together with plasma membrane homeostasis in N. crassa response to chitosan. Deletion strain analysis of chitosan susceptibility pointed NCU03639 encoding amore » class 3 lipase, involved in plasma membrane repair by lipid replacement, and NCU04537 a MFS monosaccharide transporter related to assimilation of simple sugars, as main gene targets of chitosan. NCU10521, a glutathione S-transferase-4 involved in the generation of reducing power for scavenging intracellular ROS is also a determinant chitosan gene target. Ca 2+ increased tolerance to chitosan in N. crassa. Growth of NCU10610 (fig 1 domain) and SYT1 (a synaptotagmin) deletion strains was significantly increased by Ca 2+ in the presence of chitosan. Both genes play a determinant role in N. crassa membrane homeostasis. Finally, our results are of paramount importance for developing chitosan as an antifungal.« less

Neurospora crassa transcriptomics reveals oxidative stress and plasma membrane homeostasis biology genes as key targets in response to chitosan

DOE PAGES

Lopez-Moya, Federico; Kowbel, David; Nueda, Ma Jose; ...

2015-12-01

Chitosan is a natural polymer with antimicrobial activity. Chitosan causes plasma membrane permeabilization and induction of intracellular reactive oxygen species (ROS) in Neurospora crassa. In this paper, we have determined the transcriptional profile of N. crassa to chitosan and identified the main gene targets involved in the cellular response to this compound. Global network analyses showed membrane, transport and oxidoreductase activity as key nodes affected by chitosan. Activation of oxidative metabolism indicates the importance of ROS and cell energy together with plasma membrane homeostasis in N. crassa response to chitosan. Deletion strain analysis of chitosan susceptibility pointed NCU03639 encoding amore » class 3 lipase, involved in plasma membrane repair by lipid replacement, and NCU04537 a MFS monosaccharide transporter related to assimilation of simple sugars, as main gene targets of chitosan. NCU10521, a glutathione S-transferase-4 involved in the generation of reducing power for scavenging intracellular ROS is also a determinant chitosan gene target. Ca 2+ increased tolerance to chitosan in N. crassa. Growth of NCU10610 (fig 1 domain) and SYT1 (a synaptotagmin) deletion strains was significantly increased by Ca 2+ in the presence of chitosan. Both genes play a determinant role in N. crassa membrane homeostasis. Finally, our results are of paramount importance for developing chitosan as an antifungal.« less

Plasma-membrane H(+)-ATPase gene expression is regulated by NaCl in cells of the halophyte Atriplex nummularia L.

PubMed

Niu, X; Zhu, J K; Narasimhan, M L; Bressan, R A; Hasegawa, P M

1993-01-01

An Atriplex nummularia L. cDNA probe encoding the partial sequence of an isoform of the plasma-membrane H(+)-ATPase was isolated, and used to characterize the NaCl regulation of mRNA accumulation in cultured cells of this halophyte. The peptide (477 amino acids) translated from the open reading frame has the highest sequence homology to the Nicotiana plumbaginifolia plasma-membrane H(+)-ATPase isoform pma4 (greater than 80% identity) and detected a transcript of approximately 3.7 kb on Northern blots of both total and poly(A)+ RNA. The mRNA levels were comparable in unadapted cells, adapted cells (cells adapted to and growing in 342 mM NaCl) and deadapted cells (cells previously adapted to 342 mM NaCl that are now growing without salt). Increased mRNA abundance was detected in deadapted cells within 24 h after exposure to NaCl but not in unadapted cells with similar salt treatments. The NaCl up-regulation of message abundance in deadapted cells was subject to developmental control. Analogous to those reported for glycophytes, the plasma-membrane H(+)-ATPase are encoded by a multigene family in the halophyte.

Whole genome sequencing in the search for genes associated with the control of SIV infection in the Mauritian macaque model.

PubMed

de Manuel, Marc; Shiina, Takashi; Suzuki, Shingo; Dereuddre-Bosquet, Nathalie; Garchon, Henri-Jean; Tanaka, Masayuki; Congy-Jolivet, Nicolas; Aarnink, Alice; Le Grand, Roger; Marques-Bonet, Tomas; Blancher, Antoine

2018-05-08

In the Mauritian macaque experimentally inoculated with SIV, gene polymorphisms potentially associated with the plasma virus load at a set point, approximately 100 days post inoculation, were investigated. Among the 42 animals inoculated with 50 AID 50 of the same strain of SIV, none of which received any preventive or curative treatment, nine individuals were selected: three with a plasma virus load (PVL) among the lowest, three with intermediate PVL values and three among the highest PVL values. The complete genomes of these nine animals were then analyzed. Initially, attention was focused on variants with a potential functional impact on protein encoding genes (non-synonymous SNPs (NS-SNPs) and splicing variants). Thus, 424 NS-SNPs possibly associated with PVL were detected. The 424 candidates SNPs were genotyped in these 42 SIV experimentally infected animals (including the nine animals subjected to whole genome sequencing). The genes containing variants most probably associated with PVL at a set time point are analyzed herein.

Analysis of tomato plasma membrane H(+)-ATPase gene family suggests a mycorrhiza-mediated regulatory mechanism conserved in diverse plant species.

PubMed

Liu, Junli; Liu, Jianjian; Chen, Aiqun; Ji, Minjie; Chen, Jiadong; Yang, Xiaofeng; Gu, Mian; Qu, Hongye; Xu, Guohua

2016-10-01

In plants, the plasma membrane H(+)-ATPase (HA) is considered to play a crucial role in regulating plant growth and respoding to environment stresses. Multiple paralogous genes encoding different isozymes of HA have been identified and characterized in several model plants, while limited information of the HA gene family is available to date for tomato. Here, we describe the molecular and expression features of eight HA-encoding genes (SlHA1-8) from tomato. All these genes are interrupted by multiple introns with conserved positions. SlHA1, 2, and 4 were widely expressed in all tissues, while SlHA5, 6, and 7 were almost only expressed in flowers. SlHA8, the transcripts of which were barely detectable under normal or nutrient-/salt-stress growth conditions, was strongly activated in arbuscular mycorrhizal (AM) fungal-colonized roots. Extreme lack of SlHA8 expression in M161, a mutant defective to AM fungal colonization, provided genetic evidence towards the dependence of its expression on AM symbiosis. A 1521-bp SlHA8 promoter could direct the GUS reporter expression specifically in colonized cells of transgenic tobacco, soybean, and rice mycorrhizal roots. Promoter deletion assay revealed a 223-bp promoter fragment of SlHA8 containing a variant of AM-specific cis-element MYCS (vMYCS) sufficient to confer the AM-induced activity. Targeted deletion of this motif in the corresponding promoter region causes complete abolishment of GUS staining in mycorrhizal roots. Together, these results lend cogent evidence towards the evolutionary conservation of a potential regulatory mechanism mediating the activation of AM-responsive HA genes in diverse mycorrhizal plant species.

Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants

PubMed Central

Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

2016-01-01

The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na+/H+) antiporter that transports Na+ into the vacuole and exports H+ into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na+/H+ antiporter that exports Na+ to the extracellular space and imports H+ into the plant cell. Plants rely on these enzymes either to keep Na+ out of the cell or to sequester Na+ into vacuoles to avoid the toxic level of Na+ in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. PMID:26985021

The severity of hereditary porphyria is modulated by the porphyrin exporter and Lan antigen ABCB6

PubMed Central

Fukuda, Yu; Cheong, Pak Leng; Lynch, John; Brighton, Cheryl; Frase, Sharon; Kargas, Vasileios; Rampersaud, Evadnie; Wang, Yao; Sankaran, Vijay G.; Yu, Bing; Ney, Paul A.; Weiss, Mitchell J.; Vogel, Peter; Bond, Peter J.; Ford, Robert C.; Trent, Ronald J.; Schuetz, John D.

2016-01-01

Hereditary porphyrias are caused by mutations in genes that encode haem biosynthetic enzymes with resultant buildup of cytotoxic metabolic porphyrin intermediates. A long-standing open question is why the same causal porphyria mutations exhibit widely variable penetrance and expressivity in different individuals. Here we show that severely affected porphyria patients harbour variant alleles in the ABCB6 gene, also known as Lan, which encodes an ATP-binding cassette (ABC) transporter. Plasma membrane ABCB6 exports a variety of disease-related porphyrins. Functional studies show that most of these ABCB6 variants are expressed poorly and/or have impaired function. Accordingly, homozygous disruption of the Abcb6 gene in mice exacerbates porphyria phenotypes in the Fechm1Pas mouse model, as evidenced by increased porphyrin accumulation, and marked liver injury. Collectively, these studies support ABCB6 role as a genetic modifier of porphyria and suggest that porphyrin-inducing drugs may produce excessive toxicities in individuals with the rare Lan(−) blood type. PMID:27507172

No association of defined variability in leptin, leptin receptor, adiponectin, proopiomelanocortin and ghrelin gene with food preferences in the Czech population.

PubMed

Bienertova-Vasku, Julie; Bienert, Petr; Tomandl, Josef; Forejt, Martin; Vavrina, Martin; Kudelkova, Jana; Vasku, Anna

2008-02-01

Previously, it has been reported that mutations in the genes encoding for adipokines may be associated with impaired food intake and may serve as potential obesity biomarkers. The aim of this study was to investigate the possible associations of defined variability in leptin, leptin receptor, adiponectin, proopiomelanocortin and ghrelin genes with food preferences in the obese and non-obese Czech population and evaluate their potential as the obesity susceptibility genes. Using PCR followed by restriction analysis, we studied 185 volunteers. Basic anthropometrical characteristics associated to obesity were measured and the food intake was monitored using a 7-day record method. In the group of obese individuals, a subset of 34 morbidly obese patients was studied for plasma leptin and soluble leptin receptor levels. None of the examined polymorphisms was associated to anthropometrical or demographic characteristics of the study subjects. The Gln223Arg polymorphism within the leptin receptor gene was significantly associated with lower plasma leptin levels (the RR genotype being more frequent in patients with lower plasma leptin levels; P = 0.001). No associations of the examined polymorphisms with food preferences was observed. Based on our results, the examined polymorphisms in the adipokine genes do not seem to be the major risk factor for obesity development in the Czech population nor significantly affect food preferences.

Gene-diet interaction of a common FADS1 variant with marine polyunsaturated fatty acids for fatty acid composition in plasma and erythrocytes among men.

PubMed

Takkunen, Markus J; de Mello, Vanessa D; Schwab, Ursula S; Kuusisto, Johanna; Vaittinen, Maija; Ågren, Jyrki J; Laakso, Markku; Pihlajamäki, Jussi; Uusitupa, Matti I J

2016-02-01

Limited information exists on how the relationship between dietary intake of fat and fatty acids in erythrocytes and plasma is modulated by polymorphisms in the FADS gene cluster. We examined gene-diet interaction of total marine PUFA intake with a known gene encoding Δ-5 desaturase enzyme (FADS1) variant (rs174550) for fatty acids in erythrocyte membranes and plasma phospholipids (PL), cholesteryl esters (CE), and triglycerides (TG). In this cross-sectional study, fatty acid compositions were measured using GC, and total intake of polyunsaturated fat from fish and fish oil was estimated using a food frequency questionnaire in a subsample (n = 962) of the Metabolic Syndrome in Men Study. We found nominally significant gene-diet interactions for eicosapentaenoic acid (EPA, 20:5n-3) in erythrocytes (pinteraction = 0.032) and for EPA in plasma PL (pinteraction = 0.062), CE (pinteraction = 0.035), and TG (pinteraction = 0.035), as well as for docosapentaenoic acid (22:5n-3) in PL (pinteraction = 0.007). After excluding omega-3 supplement users, we found a significant gene-diet interaction for EPA in erythrocytes (pinteraction < 0.003). In a separate cohort of the Kuopio Obesity Surgery Study, the same locus was strongly associated with hepatic mRNA expression of FADS1 (p = 1.5 × 10(-10) ). FADS1 variants may modulate the relationship between marine fatty acid intake and circulating levels of long-chain omega-3 fatty acids. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A gene-environment interaction analysis of plasma selenium with prevalent and incident diabetes: The Hortega study.

PubMed

Galan-Chilet, Inmaculada; Grau-Perez, Maria; De Marco, Griselda; Guallar, Eliseo; Martin-Escudero, Juan Carlos; Dominguez-Lucas, Alejandro; Gonzalez-Manzano, Isabel; Lopez-Izquierdo, Raul; Briongos-Figuero, Laisa Socorro; Redon, Josep; Chaves, Felipe Javier; Tellez-Plaza, Maria

2017-08-01

Selenium and single-nucleotide-polymorphisms in selenoprotein genes have been associated to diabetes. However, the interaction of selenium with genetic variation in diabetes and oxidative stress-related genes has not been evaluated as a potential determinant of diabetes risk. We evaluated the cross-sectional and prospective associations of plasma selenium concentrations with type 2 diabetes, and the interaction of selenium concentrations with genetic variation in candidate polymorphisms, in a representative sample of 1452 men and women aged 18-85 years from Spain. The geometric mean of plasma selenium levels in the study sample was 84.2µg/L. 120 participants had diabetes at baseline. Among diabetes-free participants who were not lost during the follow-up (N=1234), 75 developed diabetes over time. The multivariable adjusted odds ratios (95% confidence interval) for diabetes prevalence comparing the second and third to the first tertiles of plasma selenium levels were 1.80 (1.03, 3.14) and 1.97 (1.14, 3.41), respectively. The corresponding hazard ratios (95% CI) for diabetes incidence were 1.76 (0.96, 3.22) and 1.80 (0.98, 3.31), respectively. In addition, we observed significant interactions between selenium and polymorphisms in PPARGC1A, and in genes encoding mitochondrial proteins, such as BCS1L and SDHA, and suggestive interactions of selenium with other genes related to selenoproteins and redox metabolism. Plasma selenium was positively associated with prevalent and incident diabetes. While the statistical interactions of selenium with polymorphisms involved in regulation of redox and insulin signaling pathways provide biological plausibility to the positive associations of selenium with diabetes, further research is needed to elucidate the causal pathways underlying these associations. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Deletion of acetate transporter gene ADY2 improved tolerance of Saccharomyces cerevisiae against multiple stresses and enhanced ethanol production in the presence of acetic acid.

PubMed

Zhang, Mingming; Zhang, Keyu; Mehmood, Muhammad Aamer; Zhao, Zongbao Kent; Bai, Fengwu; Zhao, Xinqing

2017-12-01

The aim of this work was to study the effects of deleting acetate transporter gene ADY2 on growth and fermentation of Saccharomyces cerevisiae in the presence of inhibitors. Comparative transcriptome analysis revealed that three genes encoding plasma membrane carboxylic acid transporters, especially ADY2, were significantly downregulated under the zinc sulfate addition condition in the presence of acetic acid stress, and the deletion of ADY2 improved growth of S. cerevisiae under acetic acid, ethanol and hydrogen peroxide stresses. Consistently, a concomitant increase in ethanol production by 14.7% in the presence of 3.6g/L acetic acid was observed in the ADY2 deletion mutant of S. cerevisiae BY4741. Decreased intracellular acetic acid, ROS accumulation, and plasma membrane permeability were observed in the ADY2 deletion mutant. These findings would be useful for developing robust yeast strains for efficient ethanol production. Copyright © 2017 Elsevier Ltd. All rights reserved.

Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants.

PubMed

Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

2016-05-01

The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na(+)/H(+)) antiporter that transports Na(+) into the vacuole and exports H(+) into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na(+)/H(+) antiporter that exports Na(+) to the extracellular space and imports H(+) into the plant cell. Plants rely on these enzymes either to keep Na(+) out of the cell or to sequester Na(+) into vacuoles to avoid the toxic level of Na(+) in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Two Membrane-Associated Tyrosine Phosphatase Homologs Potentiate C. elegans AKT-1/PKB Signaling

PubMed Central

Hu, Patrick J; Xu, Jinling; Ruvkun, Gary

2006-01-01

Akt/protein kinase B (PKB) functions in conserved signaling cascades that regulate growth and metabolism. In humans, Akt/PKB is dysregulated in diabetes and cancer; in Caenorhabditis elegans, Akt/PKB functions in an insulin-like signaling pathway to regulate larval development. To identify molecules that modulate C. elegans Akt/PKB signaling, we performed a genetic screen for enhancers of the akt-1 mutant phenotype (eak). We report the analysis of three eak genes. eak-6 and eak-5/sdf-9 encode protein tyrosine phosphatase homologs; eak-4 encodes a novel protein with an N-myristoylation signal. All three genes are expressed primarily in the two endocrine XXX cells, and their predicted gene products localize to the plasma membrane. Genetic evidence indicates that these proteins function in parallel to AKT-1 to inhibit the FoxO transcription factor DAF-16. These results define two membrane-associated protein tyrosine phosphatase homologs that may potentiate C. elegans Akt/PKB signaling by cell autonomous and cell nonautonomous mechanisms. Similar molecules may modulate Akt/PKB signaling in human endocrine tissues. PMID:16839187

Salt stress-induced proline transporters and salt stress-repressed broad specificity amino acid permeases identified by suppression of a yeast amino acid permease-targeting mutant.

PubMed Central

Rentsch, D; Hirner, B; Schmelzer, E; Frommer, W B

1996-01-01

A yeast mutant lacking SHR3, a protein specifically required for correct targeting of plasma membrane amino acid permeases, was used to study the targeting of plant transporters and as a tool to isolate new SHR3-independent amino acid transporters. For this purpose, an shr3 mutant was transformed with an Arabidopsis cDNA library. Thirty-four clones were capable of growth under selective conditions, but none showed homology with SHR3. However, genes encoding eight different amino acid transporters belonging to three different transporter families were isolated. Five of these are members of the general amino acid permease (AAP) gene family, one is a member of the NTR family, encoding an oligopeptide transporter, and two belong to a new class of transporter genes. A functional analysis of the latter two genes revealed that they encode specific proline transporters (ProT) that are distantly related to the AAP gene family. ProT1 was found to be expressed in all organs, but highest levels were found in roots, stems, and flowers. Expression in flowers was highest in the floral stalk phloem that enters the carpels and was downregulated after fertilization, indicating a specific role in supplying the ovules with proline. ProT2 transcripts were found ubiquitously throughout the plant, but expression was strongly induced under water or salt stress, implying that ProT2 plays an important role in nitrogen distribution during water stress, unlike members of the AAP gene family whose expression was repressed under the same conditions. These results corroborate the general finding that under water stress, amino acid export is impaired whereas proline export is increased. PMID:8776904

Arabidopsis ACA7, encoding a putative auto-regulated Ca(2+)-ATPase, is required for normal pollen development.

PubMed

Lucca, Noel; León, Gabriel

2012-04-01

Microgametogenesis is a complex process that involves numerous well-coordinated cell activities, ending with the production of pollen grains. Pollen development has been studied at the cytological level in Arabidopsis and other plant species, where its temporal time course has been defined. However, the molecular mechanism underlying this process is still unclear, since a relative small number of genes and/or processes have been identified as essential for pollen development. We have designed a methodology to select candidate genes for functional analysis, based on transcriptomic data obtained from different stages of pollen development. From our analyses, we selected At2g22950 as a candidate gene; this gene encodes a protein belonging to the auto-regulated Ca(2+)-ATPase family, ACA7. Microarray data indicate that ACA7 is expressed exclusively in developing pollen grains, with the highest level of mRNA at the time of the second pollen mitosis. Our RT-PCR experiments showed that ACA7 mRNA is detected exclusively in developing flowers. Confocal microscopy experiments showed a plasma membrane localization for the recombinant GFP:ACA7 protein. We identified two different insertional mutant lines, aca7-1 and aca7-2; plants from both mutant lines displayed a normal vegetative development but showed large amounts of dead pollen grains in mature flowers assayed by Alexander's staining. Histological analysis indicated that abnormalities are detected after the first pollen mitosis and we found a strong correlation between ACA7 mRNA accumulation and the severity of the phenotype. Our results indicate that ACA7 is a plasma membrane protein that has an important role during pollen development, possibly through regulation of Ca(2+) homeostasis. © Springer-Verlag 2011

Genetic control of the alternative pathway of complement in humans and age-related macular degeneration

PubMed Central

Hecker, Laura A.; Edwards, Albert O.; Ryu, Euijung; Tosakulwong, Nirubol; Baratz, Keith H.; Brown, William L.; Issa, Peter Charbel; Scholl, Hendrik P.; Pollok-Kopp, Beatrix; Schmid-Kubista, Katharina E.; Bailey, Kent R.; Oppermann, Martin

2010-01-01

Activation of the alternative pathway of complement is implicated in common neurodegenerative diseases including age-related macular degeneration (AMD). We explored the impact of common variation in genes encoding proteins of the alternative pathway on complement activation in human blood and in AMD. Genetic variation across the genes encoding complement factor H (CFH), factor B (CFB) and component 3 (C3) was determined. The influence of common haplotypes defining transcriptional and translational units on complement activation in blood was determined in a quantitative genomic association study. Individual haplotypes in CFH and CFB were associated with distinct and novel effects on plasma levels of precursors, regulators and activation products of the alternative pathway of complement in human blood. Further, genetic variation in CFH thought to influence cell surface regulation of complement did not alter plasma complement levels in human blood. Plasma markers of chronic activation (split-products Ba and C3d) and an activating enzyme (factor D) were elevated in AMD subjects. Most of the elevation in AMD was accounted for by the genetic variation controlling complement activation in human blood. Activation of the alternative pathway of complement in blood is under genetic control and increases with age. The genetic variation associated with increased activation of complement in human blood also increased the risk of AMD. Our data are consistent with a disease model in which genetic variation in the complement system increases the risk of AMD by a combination of systemic complement activation and abnormal regulation of complement activation in local tissues. PMID:19825847

Bioinformatics analysis and characteristics of VP23 encoded by the newly identified UL18 gene of duck enteritis virus

NASA Astrophysics Data System (ADS)

Chen, Xiwen; Cheng, Anchun; Wang, Mingshu; Xiang, Jun

2011-10-01

In this study, the predicted information about structures and functions of VP23 encoded by the newly identified DEV UL18 gene through bioinformatics softwares and tools. The DEV UL18 was predicted to encode a polypeptide with 322 amino acids, termed VP23, with a putative molecular mass of 35.250 kDa and a predicted isoelectric point (PI) of 8.37, no signal peptide and transmembrane domain in the polypeptide. The prediction of subcellular localization showed that the DEV-VP23 located at endoplasmic reticulum with 33.3%, mitochondrial with 22.2%, extracellular, including cell wall with 11.1%, vesicles of secretory system with 11.1%, Golgi with 11.1%, and plasma membrane with 11.1%. The acid sequence of analysis showed that the potential antigenic epitopes are situated in 45-47, 53-60, 102-105, 173-180, 185-189, 260-265, 267-271, and 292-299 amino acids. All the consequences inevitably provide some insights for further research about the DEV-VP23 and also provide a fundament for further study on the the new type clinical diagnosis of DEV and can be used for the development of new DEV vaccine.

A Member of the Sugar Transporter Family, Stl1p Is the Glycerol/H+ Symporter in Saccharomyces cerevisiae

PubMed Central

Ferreira, Célia; van Voorst, Frank; Martins, António; Neves, Luisa; Oliveira, Rui; Kielland-Brandt, Morten C.; Lucas, Cândida; Brandt, Anders

2005-01-01

Glycerol and other polyols are used as osmoprotectants by many organisms. Several yeasts and other fungi can take up glycerol by proton symport. To identify genes involved in active glycerol uptake in Saccharomyces cerevisiae we screened a deletion mutant collection comprising 321 genes encoding proteins with 6 or more predicted transmembrane domains for impaired growth on glycerol medium. Deletion of STL1, which encodes a member of the sugar transporter family, eliminates active glycerol transport. Stl1p is present in the plasma membrane in S. cerevisiae during conditions where glycerol symport is functional. Both the Stl1 protein and the active glycerol transport are subject to glucose-induced inactivation, following identical patterns. Furthermore, the Stl1 protein and the glycerol symporter activity are strongly but transiently induced when cells are subjected to osmotic shock. STL1 was heterologously expressed in Schizosaccharomyces pombe, a yeast that does not contain its own active glycerol transport system. In S. pombe, STL1 conferred the ability to take up glycerol against a concentration gradient in a proton motive force-dependent manner. We conclude that the glycerol proton symporter in S. cerevisiae is encoded by STL1. PMID:15703210

Improved α-Amylase Production by Dephosphorylation Mutation of CreD, an Arrestin-Like Protein Required for Glucose-Induced Endocytosis of Maltose Permease and Carbon Catabolite Derepression in Aspergillus oryzae

PubMed Central

Tanaka, Mizuki; Hiramoto, Tetsuya; Tada, Hinako; Shintani, Takahiro

2017-01-01

ABSTRACT Aspergillus oryzae produces copious amount of amylolytic enzymes, and MalP, a major maltose permease, is required for the expression of amylase-encoding genes. The expression of these genes is strongly repressed by carbon catabolite repression (CCR) in the presence of glucose. MalP is transported from the plasma membrane to the vacuole by endocytosis, which requires the homolog of E6-AP carboxyl terminus ubiquitin ligase HulA, an ortholog of yeast Rsp5. In yeast, arrestin-like proteins mediate endocytosis as adaptors of Rsp5 and transporters. In the present study, we examined the involvement of CreD, an arrestin-like protein, in glucose-induced MalP endocytosis and CCR of amylase-encoding genes. Deletion of creD inhibited the glucose-induced endocytosis of MalP, and CreD showed physical interaction with HulA. Phosphorylation of CreD was detected by Western blotting, and two serine residues were determined as the putative phosphorylation sites. However, the phosphorylation state of the serine residues did not regulate MalP endocytosis and its interaction with HulA. Although α-amylase production was significantly repressed by creD deletion, both phosphorylation and dephosphorylation mimics of CreD had a negligible effect on α-amylase activity. Interestingly, dephosphorylation of CreD was required for CCR relief of amylase genes that was triggered by disruption of the deubiquitinating enzyme-encoding gene creB. The α-amylase activity of the creB mutant was 1.6-fold higher than that of the wild type, and the dephosphorylation mimic of CreD further improved the α-amylase activity by 2.6-fold. These results indicate that a combination of the dephosphorylation mutation of CreD and creB disruption increased the production of amylolytic enzymes in A. oryzae. IMPORTANCE In eukaryotes, glucose induces carbon catabolite repression (CCR) and proteolytic degradation of plasma membrane transporters via endocytosis. Glucose-induced endocytosis of transporters is mediated by their ubiquitination, and arrestin-like proteins act as adaptors of transporters and ubiquitin ligases. In this study, we showed that CreD, an arrestin-like protein, is involved in glucose-induced endocytosis of maltose permease and carbon catabolite derepression of amylase gene expression in Aspergillus oryzae. Dephosphorylation of CreD was required for CCR relief triggered by the disruption of creB, which encodes a deubiquitinating enzyme; a combination of the phosphorylation-defective mutation of CreD and creB disruption dramatically improved α-amylase production. This study shows the dual function of an arrestin-like protein and provides a novel approach for improving the production of amylolytic enzymes in A. oryzae. PMID:28455339

Improved α-Amylase Production by Dephosphorylation Mutation of CreD, an Arrestin-Like Protein Required for Glucose-Induced Endocytosis of Maltose Permease and Carbon Catabolite Derepression in Aspergillus oryzae.

PubMed

Tanaka, Mizuki; Hiramoto, Tetsuya; Tada, Hinako; Shintani, Takahiro; Gomi, Katsuya

2017-07-01

Aspergillus oryzae produces copious amount of amylolytic enzymes, and MalP, a major maltose permease, is required for the expression of amylase-encoding genes. The expression of these genes is strongly repressed by carbon catabolite repression (CCR) in the presence of glucose. MalP is transported from the plasma membrane to the vacuole by endocytosis, which requires the homolog of E6-AP carboxyl terminus ubiquitin ligase HulA, an ortholog of yeast Rsp5. In yeast, arrestin-like proteins mediate endocytosis as adaptors of Rsp5 and transporters. In the present study, we examined the involvement of CreD, an arrestin-like protein, in glucose-induced MalP endocytosis and CCR of amylase-encoding genes. Deletion of creD inhibited the glucose-induced endocytosis of MalP, and CreD showed physical interaction with HulA. Phosphorylation of CreD was detected by Western blotting, and two serine residues were determined as the putative phosphorylation sites. However, the phosphorylation state of the serine residues did not regulate MalP endocytosis and its interaction with HulA. Although α-amylase production was significantly repressed by creD deletion, both phosphorylation and dephosphorylation mimics of CreD had a negligible effect on α-amylase activity. Interestingly, dephosphorylation of CreD was required for CCR relief of amylase genes that was triggered by disruption of the deubiquitinating enzyme-encoding gene creB The α-amylase activity of the creB mutant was 1.6-fold higher than that of the wild type, and the dephosphorylation mimic of CreD further improved the α-amylase activity by 2.6-fold. These results indicate that a combination of the dephosphorylation mutation of CreD and creB disruption increased the production of amylolytic enzymes in A. oryzae IMPORTANCE In eukaryotes, glucose induces carbon catabolite repression (CCR) and proteolytic degradation of plasma membrane transporters via endocytosis. Glucose-induced endocytosis of transporters is mediated by their ubiquitination, and arrestin-like proteins act as adaptors of transporters and ubiquitin ligases. In this study, we showed that CreD, an arrestin-like protein, is involved in glucose-induced endocytosis of maltose permease and carbon catabolite derepression of amylase gene expression in Aspergillus oryzae Dephosphorylation of CreD was required for CCR relief triggered by the disruption of creB , which encodes a deubiquitinating enzyme; a combination of the phosphorylation-defective mutation of CreD and creB disruption dramatically improved α-amylase production. This study shows the dual function of an arrestin-like protein and provides a novel approach for improving the production of amylolytic enzymes in A. oryzae . Copyright © 2017 American Society for Microbiology.

Clarin-1, encoded by the Usher Syndrome III causative gene, forms a membranous microdomain: possible role of clarin-1 in organizing the actin cytoskeleton.

PubMed

Tian, Guilian; Zhou, Yun; Hajkova, Dagmar; Miyagi, Masaru; Dinculescu, Astra; Hauswirth, William W; Palczewski, Krzysztof; Geng, Ruishuang; Alagramam, Kumar N; Isosomppi, Juha; Sankila, Eeva-Marja; Flannery, John G; Imanishi, Yoshikazu

2009-07-10

Clarin-1 is the protein product encoded by the gene mutated in Usher syndrome III. Although the molecular function of clarin-1 is unknown, its primary structure predicts four transmembrane domains similar to a large family of membrane proteins that include tetraspanins. Here we investigated the role of clarin-1 by using heterologous expression and in vivo model systems. When expressed in HEK293 cells, clarin-1 localized to the plasma membrane and concentrated in low density compartments distinct from lipid rafts. Clarin-1 reorganized actin filament structures and induced lamellipodia. This actin-reorganizing function was absent in the modified protein encoded by the most prevalent North American Usher syndrome III mutation, the N48K form of clarin-1 deficient in N-linked glycosylation. Proteomics analyses revealed a number of clarin-1-interacting proteins involved in cell-cell adhesion, focal adhesions, cell migration, tight junctions, and regulation of the actin cytoskeleton. Consistent with the hypothesized role of clarin-1 in actin organization, F-actin-enriched stereocilia of auditory hair cells evidenced structural disorganization in Clrn1(-/-) mice. These observations suggest a possible role for clarin-1 in the regulation and homeostasis of actin filaments, and link clarin-1 to the interactive network of Usher syndrome gene products.

Temporal Dynamics of Microbial Rhodopsin Fluorescence Reports Absolute Membrane Voltage

PubMed Central

Hou, Jennifer H.; Venkatachalam, Veena; Cohen, Adam E.

2014-01-01

Plasma membrane voltage is a fundamentally important property of a living cell; its value is tightly coupled to membrane transport, the dynamics of transmembrane proteins, and to intercellular communication. Accurate measurement of the membrane voltage could elucidate subtle changes in cellular physiology, but existing genetically encoded fluorescent voltage reporters are better at reporting relative changes than absolute numbers. We developed an Archaerhodopsin-based fluorescent voltage sensor whose time-domain response to a stepwise change in illumination encodes the absolute membrane voltage. We validated this sensor in human embryonic kidney cells. Measurements were robust to variation in imaging parameters and in gene expression levels, and reported voltage with an absolute accuracy of 10 mV. With further improvements in membrane trafficking and signal amplitude, time-domain encoding of absolute voltage could be applied to investigate many important and previously intractable bioelectric phenomena. PMID:24507604

Expression of Pkd2l2 in testis is implicated in spermatogenesis.

PubMed

Chen, Ye; Zhang, Zheng; Lv, Xiao-Yan; Wang, Yi-Dong; Hu, Zhong-Guo; Sun, Huan; Tan, Rui-Zhi; Liu, Yu-Hang; Bian, Guo-Hui; Xiao, Yan; Li, Qin-Wei; Yang, Qiu-Tan; Ai, Jian-Zhong; Feng, Lu; Yang, Yang; Wei, Yu-Quan; Zhou, Qin

2008-08-01

Pkd2l2 is a novel member of the polycystic kidney disease (PKD) gene family in mammals. Prominently expressed in testis, this gene is still poorly understood. In this study, reverse transcription polymerase chain reaction (RT-PCR) results showed a time-dependent expression pattern of Pkd2l2 in postnatal mouse testis. Immunohistochemical analysis revealed that Pkd2l2 encoded a protein, polycystin-L2, which was predominantly detectable in the plasma membrane of spermatocytes and round spermatids, as well as in the head and tail of elongating spermatids within seminiferous tubules in mouse testis tissue sections of postnatal day 14 and adult mice. A green fluorescent fusion protein of Pkd2l2 resided in the plasma membrane of HEK 293 and MDCK cells, suggesting that it functions as a plasma membrane protein. Overexpression of Pkd2l2 increased the intracellular calcium concentration of MDCK cells, as detected by flow cytometry. Collectively, these data indicated that Pkd2l2 may be involved in the mid-late stage of spermatogenesis through modulation of the intracellular calcium concentration.

Palmitoylated SCP1 is targeted to the plasma membrane and negatively regulates angiogenesis

PubMed Central

Liao, Peng; Wang, Weichao; Li, Yu; Wang, Rui; Jin, Jiali; Pang, Weijuan; Chen, Yunfei; Shen, Mingyue; Wang, Xinbo; Jiang, Dongyang; Pang, Jinjiang; Liu, Mingyao; Lin, Xia; Feng, Xin-Hua; Wang, Ping; Ge, Xin

2017-01-01

SCP1 as a nuclear transcriptional regulator acts globally to silence neuronal genes and to affect the dephosphorylation of RNA Pol ll. However, we report the first finding and description of SCP1 as a plasma membrane-localized protein in various cancer cells using EGFP- or other epitope-fused SCP1. Membrane-located SCP1 dephosphorylates AKT at serine 473, leading to the abolishment of serine 473 phosphorylation that results in suppressed angiogenesis and a decreased risk of tumorigenesis. Consistently, we observed increased AKT phosphorylation and angiogenesis followed by enhanced tumorigenesis in Ctdsp1 (which encodes SCP1) gene - knockout mice. Importantly, we discovered that the membrane localization of SCP1 is crucial for impeding angiogenesis and tumor growth, and this localization depends on palmitoylation of a conserved cysteine motif within its NH2 terminus. Thus, our study discovers a novel mechanism underlying SCP1 shuttling between the plasma membrane and nucleus, which constitutes a unique pathway in transducing AKT signaling that is closely linked to angiogenesis and tumorigenesis. DOI: http://dx.doi.org/10.7554/eLife.22058.001 PMID:28440748

Lack of Association between Polymorphisms of Hepatic Lipase with Lipid Profile in Young Jordanian Adults

PubMed Central

Khabour, Omar F; Alomari, Mahmoud A; Alzoubi, Karem H; Gharaibeh, Mohammad Y; Alhashimi, Farah H

2014-01-01

The human hepatic lipase (LIPC) gene encodes hepatic lipase, an enzyme involved in lipoprotein metabolism and regulation. Therefore, variants in LIPC gene may influence plasma lipoprotein levels. In this study, the association of LIPC C-514T and G-250A polymorphisms with plasma lipid profiles in 348 young Jordanians was investigated. Genotyping of C-514T and G-250A was performed by polymerase chain reaction and subsequent digestion with DraI and NiaIII restriction enzymes, respectively, while Roche analyzer was used to determine plasma total cholesterol, triglycerides, low-and high-density lipoprotein. The G-250 and C-514 alleles were most abundant in Jordanians with 79 and 80% frequencies, respectively. Additionally, no difference was found in the lipid–lipoprotein profile between the different genotype groups of C-514T or G-250A polymorphisms, even when males and females were examined separately (P > 0.05). In young Jordanian adults, the examined LIPC polymorphisms seem to play a limited role in determining the lipid profile. PMID:25278769

Specialized sugar sensing in diverse fungi.

PubMed

Brown, Victoria; Sabina, Jeffrey; Johnston, Mark

2009-03-10

S. cerevisiae senses glucose and galactose differently. Glucose is detected through sensors that reside in the cellular plasma membrane. When activated, the sensors initiate a signal-transduction cascade that ultimately inactivates the Rgt1 transcriptional repressor by causing degradation of its corepressors Mth1 and Std1. This results in the expression of many HXT genes encoding glucose transporters. The ensuing flood of glucose into the cell activates Mig1, a transcriptional repressor that mediates "glucose repression" of many genes, including the GAL genes; hence, glucose sensing hinders galactose utilization. Galactose is sensed in the cytoplasm via Gal3. Upon binding galactose (and ATP), Gal3 sequesters the Gal80 protein, thereby emancipating the Gal4 transcriptional activator of the GAL genes. Gal4 also activates expression of MTH1, encoding a corepressor critical for Rgt1 function. Thus, galactose inhibits glucose assimilation by encouraging repression of HXT genes. C. albicans senses glucose similarly to S. cerevisiae but does not sense galactose through Gal3-Gal80-Gal4. Its genome harbors no GAL80 ortholog, and the severely truncated CaGal4 does not regulate CaGAL genes. We present evidence that C. albicans senses galactose with its Hgt4 glucose sensor, a capability that is enabled by transcriptional "rewiring" of its sugar-sensing signal-transduction pathways. We suggest that galactose sensing through Hgt4 is ancestral in fungi.

Host-derived viral transporter protein for nitrogen uptake in infected marine phytoplankton

PubMed Central

Chambouvet, Aurélie; Milner, David S.; Attah, Victoria; Terrado, Ramón; Lovejoy, Connie; Moreau, Hervé; Derelle, Évelyne; Richards, Thomas A.

2017-01-01

Phytoplankton community structure is shaped by both bottom–up factors, such as nutrient availability, and top–down processes, such as predation. Here we show that marine viruses can blur these distinctions, being able to amend how host cells acquire nutrients from their environment while also predating and lysing their algal hosts. Viral genomes often encode genes derived from their host. These genes may allow the virus to manipulate host metabolism to improve viral fitness. We identify in the genome of a phytoplankton virus, which infects the small green alga Ostreococcus tauri, a host-derived ammonium transporter. This gene is transcribed during infection and when expressed in yeast mutants the viral protein is located to the plasma membrane and rescues growth when cultured with ammonium as the sole nitrogen source. We also show that viral infection alters the nature of nitrogen compound uptake of host cells, by both increasing substrate affinity and allowing the host to access diverse nitrogen sources. This is important because the availability of nitrogen often limits phytoplankton growth. Collectively, these data show that a virus can acquire genes encoding nutrient transporters from a host genome and that expression of the viral gene can alter the nutrient uptake behavior of host cells. These results have implications for understanding how viruses manipulate the physiology and ecology of phytoplankton, influence marine nutrient cycles, and act as vectors for horizontal gene transfer. PMID:28827361

Uncovering the functional constraints underlying the genomic organization of the odorant-binding protein genes.

PubMed

Librado, Pablo; Rozas, Julio

2013-01-01

Animal olfactory systems have a critical role for the survival and reproduction of individuals. In insects, the odorant-binding proteins (OBPs) are encoded by a moderately sized gene family, and mediate the first steps of the olfactory processing. Most OBPs are organized in clusters of a few paralogs, which are conserved over time. Currently, the biological mechanism explaining the close physical proximity among OBPs is not yet established. Here, we conducted a comprehensive study aiming to gain insights into the mechanisms underlying the OBP genomic organization. We found that the OBP clusters are embedded within large conserved arrangements. These organizations also include other non-OBP genes, which often encode proteins integral to plasma membrane. Moreover, the conservation degree of such large clusters is related to the following: 1) the promoter architecture of the confined genes, 2) a characteristic transcriptional environment, and 3) the chromatin conformation of the chromosomal region. Our results suggest that chromatin domains may restrict the location of OBP genes to regions having the appropriate transcriptional environment, leading to the OBP cluster structure. However, the appropriate transcriptional environment for OBP and the other neighbor genes is not dominated by reduced levels of expression noise. Indeed, the stochastic fluctuations in the OBP transcript abundance may have a critical role in the combinatorial nature of the olfactory coding process.

Systematic Analysis and Comparison of Nucleotide-Binding Site Disease Resistance Genes in a Diploid Cotton Gossypium raimondii

PubMed Central

Wei, Hengling; Li, Wei; Sun, Xiwei; Zhu, Shuijin; Zhu, Jun

2013-01-01

Plant disease resistance genes are a key component of defending plants from a range of pathogens. The majority of these resistance genes belong to the super-family that harbors a Nucleotide-binding site (NBS). A number of studies have focused on NBS-encoding genes in disease resistant breeding programs for diverse plants. However, little information has been reported with an emphasis on systematic analysis and comparison of NBS-encoding genes in cotton. To fill this gap of knowledge, in this study, we identified and investigated the NBS-encoding resistance genes in cotton using the whole genome sequence information of Gossypium raimondii. Totally, 355 NBS-encoding resistance genes were identified. Analyses of the conserved motifs and structural diversity showed that the most two distinct features for these genes are the high proportion of non-regular NBS genes and the high diversity of N-termini domains. Analyses of the physical locations and duplications of NBS-encoding genes showed that gene duplication of disease resistance genes could play an important role in cotton by leading to an increase in the functional diversity of the cotton NBS-encoding genes. Analyses of phylogenetic comparisons indicated that, in cotton, the NBS-encoding genes with TIR domain not only have their own evolution pattern different from those of genes without TIR domain, but also have their own species-specific pattern that differs from those of TIR genes in other plants. Analyses of the correlation between disease resistance QTL and NBS-encoding resistance genes showed that there could be more than half of the disease resistance QTL associated to the NBS-encoding genes in cotton, which agrees with previous studies establishing that more than half of plant resistance genes are NBS-encoding genes. PMID:23936305

Genome-wide comparative analysis of NBS-encoding genes between Brassica species and Arabidopsis thaliana.

PubMed

Yu, Jingyin; Tehrim, Sadia; Zhang, Fengqi; Tong, Chaobo; Huang, Junyan; Cheng, Xiaohui; Dong, Caihua; Zhou, Yanqiu; Qin, Rui; Hua, Wei; Liu, Shengyi

2014-01-03

Plant disease resistance (R) genes with the nucleotide binding site (NBS) play an important role in offering resistance to pathogens. The availability of complete genome sequences of Brassica oleracea and Brassica rapa provides an important opportunity for researchers to identify and characterize NBS-encoding R genes in Brassica species and to compare with analogues in Arabidopsis thaliana based on a comparative genomics approach. However, little is known about the evolutionary fate of NBS-encoding genes in the Brassica lineage after split from A. thaliana. Here we present genome-wide analysis of NBS-encoding genes in B. oleracea, B. rapa and A. thaliana. Through the employment of HMM search and manual curation, we identified 157, 206 and 167 NBS-encoding genes in B. oleracea, B. rapa and A. thaliana genomes, respectively. Phylogenetic analysis among 3 species classified NBS-encoding genes into 6 subgroups. Tandem duplication and whole genome triplication (WGT) analyses revealed that after WGT of the Brassica ancestor, NBS-encoding homologous gene pairs on triplicated regions in Brassica ancestor were deleted or lost quickly, but NBS-encoding genes in Brassica species experienced species-specific gene amplification by tandem duplication after divergence of B. rapa and B. oleracea. Expression profiling of NBS-encoding orthologous gene pairs indicated the differential expression pattern of retained orthologous gene copies in B. oleracea and B. rapa. Furthermore, evolutionary analysis of CNL type NBS-encoding orthologous gene pairs among 3 species suggested that orthologous genes in B. rapa species have undergone stronger negative selection than those in B .oleracea species. But for TNL type, there are no significant differences in the orthologous gene pairs between the two species. This study is first identification and characterization of NBS-encoding genes in B. rapa and B. oleracea based on whole genome sequences. Through tandem duplication and whole genome triplication analysis in B. oleracea, B. rapa and A. thaliana genomes, our study provides insight into the evolutionary history of NBS-encoding genes after divergence of A. thaliana and the Brassica lineage. These results together with expression pattern analysis of NBS-encoding orthologous genes provide useful resource for functional characterization of these genes and genetic improvement of relevant crops.

Association between maternal micronutrient status, oxidative stress, and common genetic variants in antioxidant enzymes at 15 weeks׳ gestation in nulliparous women who subsequently develop preeclampsia.

PubMed

Mistry, Hiten D; Gill, Carolyn A; Kurlak, Lesia O; Seed, Paul T; Hesketh, John E; Méplan, Catherine; Schomburg, Lutz; Chappell, Lucy C; Morgan, Linda; Poston, Lucilla

2015-01-01

Preeclampsia is a pregnancy-specific condition affecting 2-7% of women and a leading cause of perinatal and maternal morbidity and mortality. Deficiencies of specific micronutrient antioxidant activities associated with copper, selenium, zinc, and manganese have previously been linked to preeclampsia at the time of disease. Our aims were to investigate whether maternal plasma micronutrient concentrations and related antioxidant enzyme activities are altered before preeclampsia onset and to examine the dependence on genetic variations in these antioxidant enzymes. Predisease plasma samples (15±1 weeks׳ gestation) were obtained from women enrolled in the international Screening for Pregnancy Endpoints (SCOPE) study who subsequently developed preeclampsia (n=244) and from age- and BMI-matched normotensive controls (n=472). Micronutrient concentrations were measured by inductively coupled plasma mass spectrometry; associated antioxidant enzyme activities, selenoprotein-P, ceruloplasmin concentration and activity, antioxidant capacity, and markers of oxidative stress were measured by colorimetric assays. Sixty-four tag-single-nucleotide polymorphisms (SNPs) within genes encoding the antioxidant enzymes and selenoprotein-P were genotyped using allele-specific competitive PCR. Plasma copper and ceruloplasmin concentrations were modestly but significantly elevated in women who subsequently developed preeclampsia (both P<0.001) compared to controls (median (IQR), copper, 1957.4 (1787, 2177.5) vs 1850.0 (1663.5, 2051.5) µg/L; ceruloplasmin, 2.5 (1.4, 3.2) vs 2.2 (1.2, 3.0) µg/ml). There were no differences in other micronutrients or enzymes between groups. No relationship was observed between genotype for SNPs and antioxidant enzyme activity. This analysis of a prospective cohort study reports maternal micronutrient concentrations in combination with associated antioxidant enzymes and SNPs in their encoding genes in women at 15 weeks׳ gestation that subsequently developed preeclampsia. The modest elevation in copper may contribute to oxidative stress, later in pregnancy, in those women that go on to develop preeclampsia. The lack of evidence to support the hypothesis that functional SNPs influence antioxidant enzyme activity in pregnant women argues against a role for these genes in the etiology of preeclampsia. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Cardiometabolic risk factors are influenced by Stearoyl-CoA Desaturase (SCD) -1 gene polymorphisms and n-3 polyunsaturated fatty acid supplementation.

PubMed

Rudkowska, Iwona; Julien, Pierre; Couture, Patrick; Lemieux, Simone; Tchernof, André; Barbier, Olivier; Vohl, Marie-Claude

2014-05-01

To determine if single nucleotide polymorphisms (SNPs) in stearoyl-CoA desaturase (SCD)-1 gene that encodes a key enzyme for fatty acid metabolism are associated with the response of cardiometabolic risk factors to n-3 PUFA supplementation. Two hundred and ten subjects completed a 2-week run-in period followed by 6-week supplementation with 5 g of fish oil (1.9-2.2 g eicosapentaenoic acid and 1.1 g docosahexaenoic acid). Risk factors were measured pre and post n-3 supplementation. Fatty acid composition of plasma phospholipids was analyzed by GC and the desaturase indices SCD16 (16:1n-7/16:0) and SCD18 (18:1n-9/18:0) were calculated. Genotyping of eight SNPs of the SCD1 gene was performed. N-3 PUFA supplementation decreased plasma triglycerides, as well as SCD16 and SCD18 indices, but increased fasting plasma glucose concentrations. SNPs in SCD1-modified cardiometabolic risk factors pre and post n-3 PUFA supplementation: triglyceride (rs508384, p = 0.0086), IL6 (rs3071, p = 0.0485), C-reactive protein (rs3829160, p = 0.0489), and SCD18 indices (rs2234970, p = 0.0337). A significant interaction effect between the SNP and n-3 PUFA supplementation was also observed for fasting plasma glucose levels (rs508384, p = 0.0262). These results suggest that cardiometabolic risk factors are modulated by genetic variations in the SCD1 gene alone or in combination with n-3 PUFA supplementation. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Further evidence for plasma progranulin as a biomarker in bipolar disorder.

PubMed

Kittel-Schneider, Sarah; Weigl, Johannes; Volkert, Julia; Geßner, Alexandra; Schmidt, Brigitte; Hempel, Susanne; Kiel, Tilman; Olmes, David G; Bartl, Jasmin; Weber, Heike; Kopf, Juliane; Reif, Andreas

2014-03-01

A recent study suggested that progranulin (encoded by the fronto-temporal dementia risk gene GRN) plasma levels are decreased in bipolar disorder (BD). Replication of this finding is however lacking. Progranulin plasma levels of bipolar patients (n=104) and healthy controls (n=80) were measured by enzyme-linked immunosorbent assay (ELISA). Participants were also genotyped for three single nucleotide polymorphisms (SNPs) in the GRN gene (rs2879096, rs4792938 and rs5848), and the effect of genetic variation on progranulin levels was examined. Plasma progranulin levels were decreased in BD (ANCOVA, p=0.001). Furthermore, age was significantly and positively correlated with plasma progranulin (Pearson׳s correlation, r=0.269, p<0.001). Also, lithium treatment but no other medication had a significant effect on progranulin plasma levels (ANCOVA, p=0.007). Specifically in BD, the GRN SNP rs5848 was associated with progranulin plasma levels (Kruskal-Wallis test, p<0.005). Subgroup analysis regarding bipolar I vs. bipolar II subtype and polarity of the episode at sampling (manic vs. depressed vs. mixed vs. rapid cycling vs. euthymic) could only be performed with limited validity due to the relatively small sample size. The suitability of peripheral progranulin as a biomarker for BD is limited due to the overlap between patients and controls. The findings strengthen the evidence for progranulin being involved in pathomechanisms of bipolar disorder, and suggest a genetic determinant of progranulin concentrations that is relevant specifically in bipolar patients. Copyright © 2014 Elsevier B.V. All rights reserved.

The root application of a purified leonardite humic acid modifies the transcriptional regulation of the main physiological root responses to Fe deficiency in Fe-sufficient cucumber plants.

PubMed

Aguirre, Elena; Elena, Aguirre; Leménager, Diane; Diane, Leménager; Bacaicoa, Eva; Eva, Bacaicoa; Fuentes, Marta; Marta, Fuentes; Baigorri, Roberto; Roberto, Baigorri; Zamarreño, Angel Ma; García-Mina, José Ma

2009-03-01

The aim of this study is to investigate the effect of a well-characterized purified humic acid (non-measurable concentrations of the main plant hormones were detected) on the transcriptional regulation of the principal molecular agents involved in iron assimilation. To this end, non-deficient cucumber plants were treated with different concentrations of a purified humic acid (PHA) (2, 5, 100 and 250 mg of organic carbonL(-1)) and harvested 4, 24, 48, 76 and 92 h from the onset of the treatment. At harvest times, the mRNA transcript accumulation of CsFRO1 encoding for Fe(III) chelate-reductase (EC 1.16.1.7); CsHa1 and CsHa2 encoding for plasma membrane H+-ATPase (EC 3.6.3.6); and CsIRT1 encoding for Fe(II) high-affinity transporter, was quantified by real-time RT-PCR. Meanwhile, the respective enzyme activity of the Fe(III) chelate-reductase and plasma membrane H+-ATPase was also investigated. The results obtained indicated that PHA root treatments affected the regulation of the expression of the studied genes, but this effect was transient and differed (up-regulation or down-regulation) depending on the genes studied. Thus, principally the higher doses of PHA caused a transient increase in the expression of the CsHa2 isoform for 24 and 48 h whereas the CsHa1 isoform was unaffected or down-regulated. These effects were accompanied by an increase in the plasma membrane H+-ATPase activity for 4, 48 and 96 h. Likewise, PHA root treatments (principally the higher doses) up-regulated CsFRO1 and CsIRT1 expression for 48 and 72 h; whereas these genes were down-regulated by PHA for 96 h. These effects were associated with an increase in the Fe(III) chelate-reductase activity for 72 h. These effects were not associated with a significant decrease in the Fe root or leaf concentrations, although an eventual effect on the Fe root assimilation pattern cannot be ruled out. These results stress the close relationships between the effects of humic substances on plant development and iron nutrition. However, further studies are needed in order to elucidate if these effects at molecular level are caused by mechanisms involving hormone-like actions and/or nutritional factors.

Expression of the Aspergillus terreus itaconic acid biosynthesis cluster in Aspergillus niger

PubMed Central

2014-01-01

Background Aspergillus terreus is a natural producer of itaconic acid and is currently used to produce itaconic acid on an industrial scale. The metabolic process for itaconic acid biosynthesis is very similar to the production of citric acid in Aspergillus niger. However, a key enzyme in A. niger, cis-aconitate decarboxylase, is missing. The introduction of the A. terreus cadA gene in A. niger exploits the high level of citric acid production (over 200 g per liter) and theoretically can lead to production levels of over 135 g per liter of itaconic acid in A. niger. Given the potential for higher production levels in A. niger, production of itaconic acid in this host was investigated. Results Expression of Aspergillus terreus cis-aconitate decarboxylase in Aspergillus niger resulted in the production of a low concentration (0.05 g/L) of itaconic acid. Overexpression of codon-optimized genes for cis-aconitate decarboxylase, a mitochondrial transporter and a plasma membrane transporter in an oxaloacetate hydrolase and glucose oxidase deficient A. niger strain led to highly increased yields and itaconic acid production titers. At these higher production titers, the effect of the mitochondrial and plasma membrane transporters was much more pronounced, with levels being 5–8 times higher than previously described. Conclusions Itaconic acid can be produced in A. niger by the introduction of the A. terreus cis-aconitate decarboxylase encoding cadA gene. This results in a low itaconic acid production level, which can be increased by codon-optimization of the cadA gene for A. niger. A second crucial requirement for efficient production of itaconic acid is the expression of the A. terreus mttA gene, encoding a putative mitochondrial transporter. Expression of this transporter results in a twenty-fold increase in the secretion of itaconic acid. Expression of the A. terreus itaconic acid cluster consisting of the cadA gene, the mttA gene and the mfsA gene results in A. niger strains that produce over twenty five-fold higher levels of itaconic acid and show a twenty-fold increase in yield compared to a strain expressing only CadA. PMID:24438100

Expression of the Aspergillus terreus itaconic acid biosynthesis cluster in Aspergillus niger.

PubMed

van der Straat, Laura; Vernooij, Marloes; Lammers, Marieke; van den Berg, Willy; Schonewille, Tom; Cordewener, Jan; van der Meer, Ingrid; Koops, Andries; de Graaff, Leo H

2014-01-17

Aspergillus terreus is a natural producer of itaconic acid and is currently used to produce itaconic acid on an industrial scale. The metabolic process for itaconic acid biosynthesis is very similar to the production of citric acid in Aspergillus niger. However, a key enzyme in A. niger, cis-aconitate decarboxylase, is missing. The introduction of the A. terreus cadA gene in A. niger exploits the high level of citric acid production (over 200 g per liter) and theoretically can lead to production levels of over 135 g per liter of itaconic acid in A. niger. Given the potential for higher production levels in A. niger, production of itaconic acid in this host was investigated. Expression of Aspergillus terreus cis-aconitate decarboxylase in Aspergillus niger resulted in the production of a low concentration (0.05 g/L) of itaconic acid. Overexpression of codon-optimized genes for cis-aconitate decarboxylase, a mitochondrial transporter and a plasma membrane transporter in an oxaloacetate hydrolase and glucose oxidase deficient A. niger strain led to highly increased yields and itaconic acid production titers. At these higher production titers, the effect of the mitochondrial and plasma membrane transporters was much more pronounced, with levels being 5-8 times higher than previously described. Itaconic acid can be produced in A. niger by the introduction of the A. terreus cis-aconitate decarboxylase encoding cadA gene. This results in a low itaconic acid production level, which can be increased by codon-optimization of the cadA gene for A. niger. A second crucial requirement for efficient production of itaconic acid is the expression of the A. terreus mttA gene, encoding a putative mitochondrial transporter. Expression of this transporter results in a twenty-fold increase in the secretion of itaconic acid. Expression of the A. terreus itaconic acid cluster consisting of the cadA gene, the mttA gene and the mfsA gene results in A. niger strains that produce over twenty five-fold higher levels of itaconic acid and show a twenty-fold increase in yield compared to a strain expressing only CadA.

Topological and organizational properties of the products of house-keeping and tissue-specific genes in protein-protein interaction networks.

PubMed

Lin, Wen-Hsien; Liu, Wei-Chung; Hwang, Ming-Jing

2009-03-11

Human cells of various tissue types differ greatly in morphology despite having the same set of genetic information. Some genes are expressed in all cell types to perform house-keeping functions, while some are selectively expressed to perform tissue-specific functions. In this study, we wished to elucidate how proteins encoded by human house-keeping genes and tissue-specific genes are organized in human protein-protein interaction networks. We constructed protein-protein interaction networks for different tissue types using two gene expression datasets and one protein-protein interaction database. We then calculated three network indices of topological importance, the degree, closeness, and betweenness centralities, to measure the network position of proteins encoded by house-keeping and tissue-specific genes, and quantified their local connectivity structure. Compared to a random selection of proteins, house-keeping gene-encoded proteins tended to have a greater number of directly interacting neighbors and occupy network positions in several shortest paths of interaction between protein pairs, whereas tissue-specific gene-encoded proteins did not. In addition, house-keeping gene-encoded proteins tended to connect with other house-keeping gene-encoded proteins in all tissue types, whereas tissue-specific gene-encoded proteins also tended to connect with other tissue-specific gene-encoded proteins, but only in approximately half of the tissue types examined. Our analysis showed that house-keeping gene-encoded proteins tend to occupy important network positions, while those encoded by tissue-specific genes do not. The biological implications of our findings were discussed and we proposed a hypothesis regarding how cells organize their protein tools in protein-protein interaction networks. Our results led us to speculate that house-keeping gene-encoded proteins might form a core in human protein-protein interaction networks, while clusters of tissue-specific gene-encoded proteins are attached to the core at more peripheral positions of the networks.

Draft genome sequence of Actinotignum schaalii DSM 15541T: Genetic insights into the lifestyle, cell fitness and virulence.

PubMed

Yassin, Atteyet F; Langenberg, Stefan; Huntemann, Marcel; Clum, Alicia; Pillay, Manoj; Palaniappan, Krishnaveni; Varghese, Neha; Mikhailova, Natalia; Mukherjee, Supratim; Reddy, T B K; Daum, Chris; Shapiro, Nicole; Ivanova, Natalia; Woyke, Tanja; Kyrpides, Nikos C

2017-01-01

The permanent draft genome sequence of Actinotignum schaalii DSM 15541T is presented. The annotated genome includes 2,130,987 bp, with 1777 protein-coding and 58 rRNA-coding genes. Genome sequence analysis revealed absence of genes encoding for: components of the PTS systems, enzymes of the TCA cycle, glyoxylate shunt and gluconeogensis. Genomic data revealed that A. schaalii is able to oxidize carbohydrates via glycolysis, the nonoxidative pentose phosphate and the Entner-Doudoroff pathways. Besides, the genome harbors genes encoding for enzymes involved in the conversion of pyruvate to lactate, acetate and ethanol, which are found to be the end products of carbohydrate fermentation. The genome contained the gene encoding Type I fatty acid synthase required for de novo FAS biosynthesis. The plsY and plsX genes encoding the acyltransferases necessary for phosphatidic acid biosynthesis were absent from the genome. The genome harbors genes encoding enzymes responsible for isoprene biosynthesis via the mevalonate (MVA) pathway. Genes encoding enzymes that confer resistance to reactive oxygen species (ROS) were identified. In addition, A. schaalii harbors genes that protect the genome against viral infections. These include restriction-modification (RM) systems, type II toxin-antitoxin (TA), CRISPR-Cas and abortive infection system. A. schaalii genome also encodes several virulence factors that contribute to adhesion and internalization of this pathogen such as the tad genes encoding proteins required for pili assembly, the nanI gene encoding exo-alpha-sialidase, genes encoding heat shock proteins and genes encoding type VII secretion system. These features are consistent with anaerobic and pathogenic lifestyles. Finally, resistance to ciprofloxacin occurs by mutation in chromosomal genes that encode the subunits of DNA-gyrase (GyrA) and topisomerase IV (ParC) enzymes, while resistant to metronidazole was due to the frxA gene, which encodes NADPH-flavin oxidoreductase.

Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation

PubMed Central

2012-01-01

Background The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Superfamily (MFS) in S. cerevisiae were scrutinized for a role in ethanol stress resistance. Results A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased 3 H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. Conclusions PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to improve alcoholic fermentation performance for sustainable bio-ethanol production. PMID:22839110

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pestov, Nikolay B., E-mail: korn@mail.ibch.ru; Dmitriev, Ruslan I.; Kostina, Maria B.

Highlights: Black-Right-Pointing-Pointer Full-length secretory pathway Ca-ATPase (SPCA2) cloned from rat duodenum. Black-Right-Pointing-Pointer ATP2C2 gene (encoding SPCA2) exists only in genomes of Tetrapoda. Black-Right-Pointing-Pointer Rat and pig SPCA2 are expressed in intestines, lung and some secretory glands. Black-Right-Pointing-Pointer Subcellular localization of SPCA2 may depend on tissue type. Black-Right-Pointing-Pointer In rat duodenum, SPCA2 is localized in plasma membrane-associated compartments. -- Abstract: Secretory pathway Ca-ATPases are less characterized mammalian calcium pumps than plasma membrane Ca-ATPases and sarco-endoplasmic reticulum Ca-ATPases. Here we report analysis of molecular evolution, alternative splicing, tissue-specific expression and subcellular localization of the second isoform of the secretory pathway Ca-ATPase (SPCA2),more » the product of the ATP2C2 gene. The primary structure of SPCA2 from rat duodenum deduced from full-length transcript contains 944 amino acid residues, and exhibits 65% sequence identity with known SPCA1. The rat SPCA2 sequence is also highly homologous to putative human protein KIAA0703, however, the latter seems to have an aberrant N-terminus originating from intron 2. The tissue-specificity of SPCA2 expression is different from ubiquitous SPCA1. Rat SPCA2 transcripts were detected predominantly in gastrointestinal tract, lung, trachea, lactating mammary gland, skin and preputial gland. In the newborn pig, the expression profile is very similar with one remarkable exception: porcine bulbourethral gland gave the strongest signal. Upon overexpression in cultured cells, SPCA2 shows an intracellular distribution with remarkable enrichment in Golgi. However, in vivo SPCA2 may be localized in compartments that differ among various tissues: it is intracellular in epidermis, but enriched in plasma membranes of the intestinal epithelium. Analysis of SPCA2 sequences from various vertebrate species argue that ATP2C2 gene radiated from ATP2C1 (encoding SPCA1) during adaptation of tetrapod ancestors to terrestrial habitats.« less

SEMI-ROLLED LEAF1 Encodes a Putative Glycosylphosphatidylinositol-Anchored Protein and Modulates Rice Leaf Rolling by Regulating the Formation of Bulliform Cells1[W][OA

PubMed Central

Xiang, Jing-Jing; Zhang, Guang-Heng; Qian, Qian; Xue, Hong-Wei

2012-01-01

Leaf rolling is an important agronomic trait in rice (Oryza sativa) breeding and moderate leaf rolling maintains the erectness of leaves and minimizes shadowing between leaves, leading to improved photosynthetic efficiency and grain yields. Although a few rolled-leaf mutants have been identified and some genes controlling leaf rolling have been isolated, the molecular mechanisms of leaf rolling still need to be elucidated. Here we report the isolation and characterization of SEMI-ROLLED LEAF1 (SRL1), a gene involved in the regulation of leaf rolling. Mutants srl1-1 (point mutation) and srl1-2 (transferred DNA insertion) exhibit adaxially rolled leaves due to the increased numbers of bulliform cells at the adaxial cell layers, which could be rescued by complementary expression of SRL1. SRL1 is expressed in various tissues and is expressed at low levels in bulliform cells. SRL1 protein is located at the plasma membrane and predicted to be a putative glycosylphosphatidylinositol-anchored protein. Moreover, analysis of the gene expression profile of cells that will become epidermal cells in wild type but probably bulliform cells in srl1-1 by laser-captured microdissection revealed that the expression of genes encoding vacuolar H+-ATPase (subunits A, B, C, and D) and H+-pyrophosphatase, which are increased during the formation of bulliform cells, were up-regulated in srl1-1. These results provide the transcript profile of rice leaf cells that will become bulliform cells and demonstrate that SRL1 regulates leaf rolling through inhibiting the formation of bulliform cells by negatively regulating the expression of genes encoding vacuolar H+-ATPase subunits and H+-pyrophosphatase, which will help to understand the mechanism regulating leaf rolling. PMID:22715111

The Arabidopsis SOS5 Locus Encodes a Putative Cell Surface Adhesion Protein and Is Required for Normal Cell Expansion

PubMed Central

Shi, Huazhong; Kim, YongSig; Guo, Yan; Stevenson, Becky; Zhu, Jian-Kang

2003-01-01

Cell surface proteoglycans have been implicated in many aspects of plant growth and development, but genetic evidence supporting their function has been lacking. Here, we report that the Salt Overly Sensitive5 (SOS5) gene encodes a putative cell surface adhesion protein and is required for normal cell expansion. The sos5 mutant was isolated in a screen for Arabidopsis salt-hypersensitive mutants. Under salt stress, the root tips of sos5 mutant plants swell and root growth is arrested. The root-swelling phenotype is caused by abnormal expansion of epidermal, cortical, and endodermal cells. The SOS5 gene was isolated through map-based cloning. The predicted SOS5 protein contains an N-terminal signal sequence for plasma membrane localization, two arabinogalactan protein–like domains, two fasciclin-like domains, and a C-terminal glycosylphosphatidylinositol lipid anchor signal sequence. The presence of fasciclin-like domains, which typically are found in animal cell adhesion proteins, suggests a role for SOS5 in cell-to-cell adhesion in plants. The SOS5 protein was present at the outer surface of the plasma membrane. The cell walls are thinner in the sos5 mutant, and those between neighboring epidermal and cortical cells in sos5 roots appear less organized. SOS5 is expressed ubiquitously in all plant organs and tissues, including guard cells in the leaf. PMID:12509519

A mutation in the insulin receptor gene that impairs transport of the receptor to the plasma membrane and causes insulin-resistant diabetes.

PubMed Central

Accili, D; Frapier, C; Mosthaf, L; McKeon, C; Elbein, S C; Permutt, M A; Ramos, E; Lander, E; Ullrich, A; Taylor, S I

1989-01-01

Insulin binds to a receptor on the cell surface, thereby triggering a biological response within the target cell. Mutations in the insulin receptor gene can render the cell resistant to the biological action of insulin. We have studied a family in which two sisters have a genetic form of insulin-resistant diabetes mellitus. The technique of homozygosity mapping has been used to demonstrate that the mutation causing diabetes in this consanguineous family is genetically linked to the insulin receptor gene. The two insulin-resistant sisters are homozygous for a mutation encoding substitution of valine for phenylalanine at position 382 in the alpha-subunit of the insulin receptor. Transfection of mutant insulin receptor cDNA into NIH3T3 cells demonstrated that the Val382 mutation impaired post-translational processing and retarded transport of the insulin receptor to the plasma membrane. Thus, the mutation causes insulin resistance by decreasing the number of insulin receptors on the surface of the patients' cells. Images PMID:2573522

Nucleotide sequences of two genomic DNAs encoding peroxidase of Arabidopsis thaliana.

PubMed

Intapruk, C; Higashimura, N; Yamamoto, K; Okada, N; Shinmyo, A; Takano, M

1991-02-15

The peroxidase (EC 1.11.1.7)-encoding gene of Arabidopsis thaliana was screened from a genomic library using a cDNA encoding a neutral isozyme of horseradish, Armoracia rusticana, peroxidase (HRP) as a probe, and two positive clones were isolated. From the comparison with the sequences of the HRP-encoding genes, we concluded that two clones contained peroxidase-encoding genes, and they were named prxCa and prxEa. Both genes consisted of four exons and three introns; the introns had consensus nucleotides, GT and AG, at the 5' and 3' ends, respectively. The lengths of each putative exon of the prxEa gene were the same as those of the HRP-basic-isozyme-encoding gene, prxC3, and coded for 349 amino acids (aa) with a sequence homology of 89% to that encoded by prxC3. The prxCa gene was very close to the HRP-neutral-isozyme-encoding gene, prxC1b, and coded for 354 aa with 91% homology to that encoded by prxC1b. The aa sequence homology was 64% between the two peroxidases encoded by prxCa and prxEa.

Modulation by geraniol of gene expression involved in lipid metabolism leading to a reduction of serum-cholesterol and triglyceride levels.

PubMed

Galle, Marianela; Kladniew, Boris Rodenak; Castro, María Agustina; Villegas, Sandra Montero; Lacunza, Ezequiel; Polo, Mónica; de Bravo, Margarita García; Crespo, Rosana

2015-07-15

Geraniol (G) is a natural isoprenoid present in the essential oils of several aromatic plants, with various biochemical and pharmacologic properties. Nevertheless, the mechanisms of action of G on cellular metabolism are largely unknown. We propose that G could be a potential agent for the treatment of hyperlipidemia that could contribute to the prevention of cardiovascular disease. The aim of the present study was to advance our understanding of its mechanism of action on cholesterol and TG metabolism. NIH mice received supplemented diets containing 25, 50, and 75 mmol G/kg chow. After a 3-week treatment, serum total-cholesterol and triglyceride levels were measured by commercial kits and lipid biosynthesis determined by the [(14)C] acetate incorporated into fatty acids plus nonsaponifiable and total hepatic lipids of the mice. The activity of the mRNA encoding HMGCR-the rate-limiting step in cholesterol biosynthesis-along with the enzyme levels and catalysis were assessed by real-time RT-PCR, Western blotting, and HMG-CoA-conversion assays, respectively. In-silico analysis of several genes involved in lipid metabolism and regulated by G in cultured cells was also performed. Finally, the mRNA levels encoded by the genes for the low-density-lipoprotein receptor (LDLR), the sterol-regulatory-element-binding transcription factor (SREBF2), the very-low-density-lipoprotein receptor (VLDLR), and the acetyl-CoA carboxylase (ACACA) were determined by real-time RT-PCR. Plasma total-cholesterol and triglyceride levels plus hepatic fatty-acid, total-lipid, and nonsaponifiable-lipid biosynthesis were significantly reduced by feeding with G. Even though an up-regulation of the mRNA encoding HMGCR occurred in the G treated mouse livers, the protein levels and specific activity of the enzyme were both inhibited. G also enhanced the mRNAs encoding the LDL and VLDL receptors and reduced ACACA mRNA, without altering the transcription of the mRNA encoding the SREBF2. The following mechanisms may have mediated the decrease in plasma lipids levels in mice: a down-regulation of hepatocyte-cholesterol synthesis occurred as a result of decreased HMGCR protein levels and catalytic activity; the levels of LDLR mRNA became elevated, thus suggesting an increase in the uptake of serum LDL, especially by the liver; and TG synthesis became reduced very likely because of a decrease in fatty-acid synthesis. Copyright © 2015 Elsevier GmbH. All rights reserved.

Identification and expression of three new Nicotiana plumbaginifolia genes which encode isoforms of a plasma-membrane H(+)-ATPase, and one of which is induced by mechanical stress.

PubMed

Oufattole, M; Arango, M; Boutry, M

2000-04-01

To analyze in detail the multigene family encoding the plasma-membrane H(+)-ATPase (pma) in Nicotiana plumbaginifolia Viv., five new pma genes (pma 5-9) were isolated. Three of these (pma 6, 8, 9) were fully characterized and classified into new and independent subfamilies. Their cell-type expression was followed by the beta-glucuronidase (gusA) reporter-gene method. While the pma8-gusA transgene was not expressed in transgenic tobacco, expression of the two other transgenes (pma6- and pma9-gusA) was found to be restricted to particular cell types. In the vegetative tissues, pma6-gusA expression was limited to the head cells of the leaf short trichomes, involved in secretion, and to the cortical parenchyma of the young nodes where the developing leaves and axillary flowering stalks join the stem. In the latter tissues, gene expression was enhanced by mechanical stress, suggesting that H(+)-ATPase might be involved in the strength of the tissues and their resistance to mechanical trauma. The pma9-gusA transgene was mainly expressed in the apical meristem of adventitious roots and axillary buds as well as in the phloem tissues of the stem, in which expression depended on the developmental stage. In flowers, pma9-gusA expression was limited to the mature pollen grains and the young fertilized ovules, while that of pma6-gusA was identified in most of the organs. Reverse transcription-polymerase chain reaction of leaf and stem RNA confirmed the expression of pma 6 and 9, while pma8 was found to be expressed in both organs at a lower level. In conclusion, although pma 6 and 9 had a more restricted expression pattern than the previously characterized pma genes, they were nevertheless expressed in cell types in which H(+)-ATPase had not been previously detected.

The 'spectraplakins': cytoskeletal giants with characteristics of both spectrin and plakin families.

PubMed

Röper, Katja; Gregory, Stephen L; Brown, Nicholas H

2002-11-15

Recent studies have characterised a family of giant cytoskeletal crosslinkers encoded by the short stop gene in Drosophila and the dystonin/BPAG1 and MACF1 genes in mammals. We refer to the products of these genes as spectraplakins to highlight the fact that they share features with both the spectrin and plakin superfamilies. These genes produce a variety of large proteins, up to almost 9000 residues long, which can potentially extend 0.4 micro m across a cell. Spectraplakins can interact with all three elements of the cytoskeleton: actin, microtubules and intermediate filaments. The analysis of mutant phenotypes in BPAG1 in mouse and short stop in Drosophila demonstrates that spectraplakins have diverse roles. These include linking the plasma membrane and the cytoskeleton, linking together different elements of the cytoskeleton and organising membrane domains.

Mutation in the factor VII hepatocyte nuclear factor 4α-binding site contributes to factor VII deficiency.

PubMed

Zheng, Xing-Wu; Kudaravalli, Rama; Russell, Theresa T; DiMichele, Donna M; Gibb, Constance; Russell, J Eric; Margaritis, Paris; Pollak, Eleanor S

2011-10-01

Severe coagulant factor VII (FVII) deficiency in postpubertal dizygotic twin males results from two point mutations in the FVII gene, a promoter region T→C transition at -60 and a His-to-Arg substitution at amino acid 348; both mutations prevent persistence of plasma functional FVII. This report documents longitudinal laboratory measurements from infancy to adulthood of FVII coagulant activity (FVII:C) in the twin FVII-deficient patients; it also details specific biochemical analyses of the -60 T→C mutation. The results revealed FVII:C levels of less than 1% in infancy that remain severely decreased through puberty and into adulthood. In-vitro analyses utilizing hepatocyte nuclear factor 4α (HNF4α) co-transfection and a chromatin immunoprecipitation assay indicate that the -60 T→C mutation severely diminishes functional interaction between the FVII promoter and transcription factor HNF4α. The importance of interaction between the FVII gene and HNF4α in normal FVII expression provides an in-vivo illustration of the regulated expression of an autosomal gene encoding a coagulation protein. The constancy of FVII:C and peripubertal patient symptomatology reported here illustrates androgen-independent expression in contrast to expression with an analogous mutation in the promoter region of the gene encoding coagulation FIX.

Degradation of Benzene by Pseudomonas veronii 1YdBTEX2 and 1YB2 Is Catalyzed by Enzymes Encoded in Distinct Catabolism Gene Clusters.

PubMed

de Lima-Morales, Daiana; Chaves-Moreno, Diego; Wos-Oxley, Melissa L; Jáuregui, Ruy; Vilchez-Vargas, Ramiro; Pieper, Dietmar H

2016-01-01

Pseudomonas veronii 1YdBTEX2, a benzene and toluene degrader, and Pseudomonas veronii 1YB2, a benzene degrader, have previously been shown to be key players in a benzene-contaminated site. These strains harbor unique catabolic pathways for the degradation of benzene comprising a gene cluster encoding an isopropylbenzene dioxygenase where genes encoding downstream enzymes were interrupted by stop codons. Extradiol dioxygenases were recruited from gene clusters comprising genes encoding a 2-hydroxymuconic semialdehyde dehydrogenase necessary for benzene degradation but typically absent from isopropylbenzene dioxygenase-encoding gene clusters. The benzene dihydrodiol dehydrogenase-encoding gene was not clustered with any other aromatic degradation genes, and the encoded protein was only distantly related to dehydrogenases of aromatic degradation pathways. The involvement of the different gene clusters in the degradation pathways was suggested by real-time quantitative reverse transcription PCR. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Remodeling of Hepatic Metabolism and Hyperaminoacidemia in Mice Deficient in Proglucagon-Derived Peptides

PubMed Central

Watanabe, Chika; Seino, Yusuke; Miyahira, Hiroki; Yamamoto, Michiyo; Fukami, Ayako; Ozaki, Nobuaki; Takagishi, Yoshiko; Sato, Jun; Fukuwatari, Tsutomu; Shibata, Katsumi; Oiso, Yutaka; Murata, Yoshiharu; Hayashi, Yoshitaka

2012-01-01

Glucagon is believed to be one of the most important peptides for upregulating blood glucose levels. However, homozygous glucagon–green fluorescent protein (gfp) knock-in mice (Gcggfp/gfp: GCGKO) are normoglycemic despite the absence of proglucagon-derived peptides, including glucagon. To characterize metabolism in the GCGKO mice, we analyzed gene expression and metabolome in the liver. The expression of genes encoding rate-limiting enzymes for gluconeogenesis was only marginally altered. On the other hand, genes encoding enzymes involved in conversion of amino acids to metabolites available for the tricarboxylic acid cycle and/or gluconeogenesis showed lower expression in the GCGKO liver. The expression of genes involved in the metabolism of fatty acids and nicotinamide was also altered. Concentrations of the metabolites in the GCGKO liver were altered in manners concordant with alteration in the gene expression patterns, and the plasma concentrations of amino acids were elevated in the GCGKO mice. The insulin concentration in serum and phosphorylation of Akt protein kinase in liver were reduced in GCGKO mice. These results indicated that proglucagon-derived peptides should play important roles in regulating various metabolic pathways, especially that of amino acids. Serum insulin concentration is lowered to compensate the impacts of absent proglucagon-derived peptide on glucose metabolism. On the other hand, impacts on other metabolic pathways are only partially compensated by reduced insulin action. PMID:22187375

Cloning, expression and N-terminal myristoylation of CpCPK1, a calcium-dependent protein kinase from zucchini (Cucurbita pepo L.).

PubMed

Ellard-Ivey, M; Hopkins, R B; White, T J; Lomax, T L

1999-01-01

We have isolated a full-length cDNA clone (CpCDPK1) encoding a calcium-dependent protein kinase (CDPK) gene from zucchini (Cucurbita pepo L.). The predicted amino acid sequence of the cDNA shows a remarkably high degree of similarity to members of the CDPK gene family from Arabidopsis thaliana, especially AtCPK1 and AtCPK2. Northern analysis of steady-state mRNA levels for CpCPK1 in etiolated and light-grown zucchini seedlings shows that the transcript is most abundant in etiolated hypocotyls and overall expression is suppressed by light. As described for other members of the CDPK gene family from different species, the CpCPK1 clone has a putative N-terminal myristoylation sequence. In this study, site-directed mutagenesis and an in vitro coupled transcription/translation system were used to demonstrate that the protein encoded by this cDNA is specifically myristoylated by a plant N-myristoyl transferase. This is the first demonstration of myristoylation of a CDPK protein which may contribute to the mechanism by which this protein is localized to the plasma membrane.

Inactivation of the Carney complex gene 1 (PRKAR1A) alters spatiotemporal regulation of cAMP and cAMP-dependent protein kinase: a study using genetically encoded FRET-based reporters.

PubMed

Cazabat, Laure; Ragazzon, Bruno; Varin, Audrey; Potier-Cartereau, Marie; Vandier, Christophe; Vezzosi, Delphine; Risk-Rabin, Marthe; Guellich, Aziz; Schittl, Julia; Lechêne, Patrick; Richter, Wito; Nikolaev, Viacheslav O; Zhang, Jin; Bertherat, Jérôme; Vandecasteele, Grégoire

2014-03-01

Carney complex (CNC) is a hereditary disease associating cardiac myxoma, spotty skin pigmentation and endocrine overactivity. CNC is caused by inactivating mutations in the PRKAR1A gene encoding PKA type I alpha regulatory subunit (RIα). Although PKA activity is enhanced in CNC, the mechanisms linking PKA dysregulation to endocrine tumorigenesis are poorly understood. In this study, we used Förster resonance energy transfer (FRET)-based sensors for cAMP and PKA activity to define the role of RIα in the spatiotemporal organization of the cAMP/PKA pathway. RIα knockdown in HEK293 cells increased basal as well as forskolin or prostaglandin E1 (PGE1)-stimulated total cellular PKA activity as reported by western blots of endogenous PKA targets and the FRET-based global PKA activity reporter, AKAR3. Using variants of AKAR3 targeted to subcellular compartments, we identified similar increases in the response to PGE1 in the cytoplasm and at the outer mitochondrial membrane. In contrast, at the plasma membrane, the response to PGE1 was decreased along with an increase in basal FRET ratio. These results were confirmed by western blot analysis of basal and PGE1-induced phosphorylation of membrane-associated vasodilator-stimulated phosphoprotein. Similar differences were observed between the cytoplasm and the plasma membrane in human adrenal cells carrying a RIα inactivating mutation. RIα inactivation also increased cAMP in the cytoplasm, at the outer mitochondrial membrane and at the plasma membrane, as reported by targeted versions of the cAMP indicator Epac1-camps. These results show that RIα inactivation leads to multiple, compartment-specific alterations of the cAMP/PKA pathway revealing new aspects of signaling dysregulation in tumorigenesis.

An Early Nodulin-Like Protein Accumulates in the Sieve Element Plasma Membrane of Arabidopsis1[OA

PubMed Central

Khan, Junaid A.; Wang, Qi; Sjölund, Richard D.; Schulz, Alexander; Thompson, Gary A.

2007-01-01

Membrane proteins within the sieve element-companion cell complex have essential roles in the physiological functioning of the phloem. The monoclonal antibody line RS6, selected from hybridomas raised against sieve elements isolated from California shield leaf (Streptanthus tortuosus; Brassicaceae) tissue cultures, recognizes an antigen in the Arabidopsis (Arabidopsis thaliana) ecotype Columbia that is associated specifically with the plasma membrane of sieve elements, but not companion cells, and accumulates at the earliest stages of sieve element differentiation. The identity of the RS6 antigen was revealed by reverse transcription-PCR of Arabidopsis leaf RNA using degenerate primers to be an early nodulin (ENOD)-like protein that is encoded by the expressed gene At3g20570. Arabidopsis ENOD-like proteins are encoded by a multigene family composed of several types of structurally related phytocyanins that have a similar overall domain structure of an amino-terminal signal peptide, plastocyanin-like copper-binding domain, proline/serine-rich domain, and carboxy-terminal hydrophobic domain. The amino- and carboxy-terminal domains of the 21.5-kD sieve element-specific ENOD are posttranslationally cleaved from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both floral and vegetative tissues, the sieve element-specific ENOD is expressed only within the phloem. Members of the ENOD subfamily of the cupredoxin superfamily do not appear to bind copper and have unknown functions. Phenotypic analysis of homozygous T-DNA insertion mutants for the gene At3g20570 shows minimal alteration in vegetative growth but a significant reduction in the overall reproductive potential. PMID:17293437

An early nodulin-like protein accumulates in the sieve element plasma membrane of Arabidopsis.

PubMed

Khan, Junaid A; Wang, Qi; Sjölund, Richard D; Schulz, Alexander; Thompson, Gary A

2007-04-01

Membrane proteins within the sieve element-companion cell complex have essential roles in the physiological functioning of the phloem. The monoclonal antibody line RS6, selected from hybridomas raised against sieve elements isolated from California shield leaf (Streptanthus tortuosus; Brassicaceae) tissue cultures, recognizes an antigen in the Arabidopsis (Arabidopsis thaliana) ecotype Columbia that is associated specifically with the plasma membrane of sieve elements, but not companion cells, and accumulates at the earliest stages of sieve element differentiation. The identity of the RS6 antigen was revealed by reverse transcription-PCR of Arabidopsis leaf RNA using degenerate primers to be an early nodulin (ENOD)-like protein that is encoded by the expressed gene At3g20570. Arabidopsis ENOD-like proteins are encoded by a multigene family composed of several types of structurally related phytocyanins that have a similar overall domain structure of an amino-terminal signal peptide, plastocyanin-like copper-binding domain, proline/serine-rich domain, and carboxy-terminal hydrophobic domain. The amino- and carboxy-terminal domains of the 21.5-kD sieve element-specific ENOD are posttranslationally cleaved from the precursor protein, resulting in a mature peptide of approximately 15 kD that is attached to the sieve element plasma membrane via a carboxy-terminal glycosylphosphatidylinositol membrane anchor. Many of the Arabidopsis ENOD-like proteins accumulate in gametophytic tissues, whereas in both floral and vegetative tissues, the sieve element-specific ENOD is expressed only within the phloem. Members of the ENOD subfamily of the cupredoxin superfamily do not appear to bind copper and have unknown functions. Phenotypic analysis of homozygous T-DNA insertion mutants for the gene At3g20570 shows minimal alteration in vegetative growth but a significant reduction in the overall reproductive potential.

Vaccination with an adenoviral vector that encodes and displays a retroviral antigen induces improved neutralizing antibody and CD4+ T-cell responses and confers enhanced protection.

PubMed

Bayer, Wibke; Tenbusch, Matthias; Lietz, Ruth; Johrden, Lena; Schimmer, Simone; Uberla, Klaus; Dittmer, Ulf; Wildner, Oliver

2010-02-01

We present a new type of adenoviral vector that both encodes and displays a vaccine antigen on the capsid, thus combining in itself gene-based and protein vaccination; this vector resulted in an improved vaccination outcome in the Friend virus (FV) model. For presentation of the envelope protein gp70 of Friend murine leukemia virus on the adenoviral capsid, gp70 was fused to the adenovirus capsid protein IX. When compared to vaccination with conventional FV Env- and Gag-encoding adenoviral vectors, vaccination with the adenoviral vector that encodes and displays pIX-gp70 combined with an FV Gag-encoding vector resulted in significantly improved protection against systemic FV challenge infection, with highly controlled viral loads in plasma and spleen. This improved protection correlated with improved neutralizing antibody titers and stronger CD4(+) T-cell responses. Using a vector that displays gp70 without encoding it, we found that while the antigen display on the capsid alone was sufficient to induce high levels of binding antibodies, in vivo expression was necessary for the induction of neutralizing antibodies. This new type of adenovirus-based vaccine could be a valuable tool for vaccination.

Targeted Quantitative Screening of Chromosome 18 Encoded Proteome in Plasma Samples of Astronaut Candidates.

PubMed

Kopylov, Artur T; Ilgisonis, Ekaterina V; Moysa, Alexander A; Tikhonova, Olga V; Zavialova, Maria G; Novikova, Svetlana E; Lisitsa, Andrey V; Ponomarenko, Elena A; Moshkovskii, Sergei A; Markin, Andrey A; Grigoriev, Anatoly I; Zgoda, Victor G; Archakov, Alexander I

2016-11-04

This work was aimed at estimating the concentrations of proteins encoded by human chromosome 18 (Chr 18) in plasma samples of 54 healthy male volunteers (aged 20-47). These young persons have been certified by the medical evaluation board as healthy subjects ready for space flight training. Over 260 stable isotope-labeled peptide standards (SIS) were synthesized to perform the measurements of proteins encoded by Chr 18. Selected reaction monitoring (SRM) with SIS allowed an estimate of the levels of 84 of 276 proteins encoded by Chr 18. These proteins were quantified in whole and depleted plasma samples. Concentration of the proteins detected varied from 10 -6 M (transthyretin, P02766) to 10 -11 M (P4-ATPase, O43861). A minor part of the proteins (mostly representing intracellular proteins) was characterized by extremely high inter individual variations. The results provide a background for studies of a potential biomarker in plasma among proteins encoded by Chr 18. The SRM raw data are available in ProteomeXchange repository (PXD004374).

Identification and differential induction of the expression of aquaporins by salinity in broccoli plants.

PubMed

Muries, Beatriz; Faize, Mohamed; Carvajal, Micaela; Martínez-Ballesta, María Del Carmen

2011-04-01

Plant aquaporins belong to a large superfamily of conserved proteins called the major intrinsic proteins (MIPs). There is limited information about the diversity of MIPs and their water transport capacity in broccoli (Brassica oleracea) plants. In this study, the cDNAs of isoforms of Plasma Membrane Intrinsic Proteins (PIPs), a class of aquaporins, from broccoli roots have been partially sequenced. Thus, sequencing experiments led to the identification of eight PIP1 and three PIP2 genes encoding PIPs in B. oleracea plants. The occurrence of different gene products encoding PIPs suggests that they may play different roles in plants. The screening of their expression as well as the expression of two specific PIP2 isoforms (BoPIP2;2 and BoPIP2;3), in different organs and under different salt-stress conditions in two varieties, has helped to unravel the function and the regulation of PIPs in plants. Thus, a high degree of BoPIP2;3 expression in mature leaves suggests that this BoPIP2;3 isoform plays important roles, not only in root water relations but also in the physiology and development of leaves. In addition, differences between gene and protein patterns led us to consider that mRNA synthesis is inhibited by the accumulation of the corresponding encoded protein. Therefore, transcript levels, protein abundance determination and the integrated hydraulic architecture of the roots must be considered in order to interpret the response of broccoli to salinity.

ALS3 encodes a phloem-localized ABC transporter-like protein that is required for aluminum tolerance in Arabidopsis.

PubMed

Larsen, Paul B; Geisler, Matt J B; Jones, Carol A; Williams, Kelly M; Cancel, Jesse D

2005-02-01

Aluminum (Al) toxicity in acid soils is a worldwide agricultural problem that severely limits crop productivity through inhibition of root growth. Previously, Arabidopsis mutants with increased Al sensitivity were isolated in order to identify genes important for Al tolerance in plants. One mutant, als3, exhibited extreme root growth inhibition in the presence of Al, suggesting that this mutation negatively impacts a gene required for Al tolerance. Map-based cloning of the als3-1 mutation resulted in the isolation of a novel gene that encodes a previously undescribed ABC transporter-like protein, which is highly homologous to a putative bacterial metal resistance protein, ybbM. Northern analysis for ALS3 expression revealed that it is found in all organs examined, which is consistent with the global nature of Al sensitivity displayed by als3, and that expression increases in roots following Al treatment. Based on GUS fusion and in situ hybridization analyses, ALS3 is primarily expressed in leaf hydathodes and the phloem throughout the plant, along with the root cortex following Al treatment. Immunolocalization indicates that ALS3 predominantly accumulates in the plasma membrane of cells that express ALS3. From our results, it appears that ALS3 encodes an ABC transporter-like protein that is required for Al resistance/tolerance and may function to redistribute accumulated Al away from sensitive tissues in order to protect the growing root from the toxic effects of Al.

Biarsenical labeling of vesicular stomatitis virus encoding tetracysteine-tagged m protein allows dynamic imaging of m protein and virus uncoating in infected cells.

PubMed

Das, Subash C; Panda, Debasis; Nayak, Debasis; Pattnaik, Asit K

2009-03-01

A recombinant vesicular stomatitis virus (VSV-PeGFP-M-MmRFP) encoding enhanced green fluorescent protein fused in frame with P (PeGFP) in place of P and a fusion matrix protein (monomeric red fluorescent protein fused in frame at the carboxy terminus of M [MmRFP]) at the G-L gene junction, in addition to wild-type (wt) M protein in its normal location, was recovered, but the MmRFP was not incorporated into the virions. Subsequently, we generated recombinant viruses (VSV-PeGFP-DeltaM-Mtc and VSV-DeltaM-Mtc) encoding M protein with a carboxy-terminal tetracysteine tag (Mtc) in place of the M protein. These recombinant viruses incorporated Mtc at levels similar to M in wt VSV, demonstrating recovery of infectious rhabdoviruses encoding and incorporating a tagged M protein. Virions released from cells infected with VSV-PeGFP-DeltaM-Mtc and labeled with the biarsenical red dye (ReAsH) were dually fluorescent, fluorescing green due to incorporation of PeGFP in the nucleocapsids and red due to incorporation of ReAsH-labeled Mtc in the viral envelope. Transport and subsequent association of M protein with the plasma membrane were shown to be independent of microtubules. Sequential labeling of VSV-DeltaM-Mtc-infected cells with the biarsenical dyes ReAsH and FlAsH (green) revealed that newly synthesized M protein reaches the plasma membrane in less than 30 min and continues to accumulate there for up to 2 1/2 hours. Using dually fluorescent VSV, we determined that following adsorption at the plasma membrane, the time taken by one-half of the virus particles to enter cells and to uncoat their nucleocapsids in the cytoplasm is approximately 28 min.

Relationship of low plasma klotho with poor grip strength in older community-dwelling adults: the InCHIANTI study

PubMed Central

Semba, Richard D.; Cappola, Anne R.; Sun, Kai; Bandinelli, Stefania; Dalal, Mansi; Crasto, Candace; Guralnik, Jack M.; Ferrucci, Luigi

2012-01-01

Handgrip strength is a strong indicator of total body muscle strength and is a predictor of poor outcomes in older adults. The aging suppressor gene klotho encodes a single-pass transmembrane protein that is secreted as a circulating hormone. In mice, disruption of klotho expression results in a syndrome that includes sarcopenia, atherosclerosis, osteoporosis, and shortened lifespan, and conversely, overexpression of klotho leads to a greater longevity. The objective was to determine whether plasma klotho levels are related to skeletal muscle strength in humans. We measured plasma klotho in 804 adults, ≥65 years, in the InCHIANTI study, a longitudinal population-based study of aging in Tuscany, Italy. Grip strength was positively correlated with plasma klotho at threshold <681 pg/mL. After adjusting for age, sex, education, smoking, physical activity, cognition, and chronic diseases, plasma klotho (per 1 standard deviation increase) was associated with grip strength (beta = 1.20, standard error = 0.35, P = 0.0009) in adults with plasma klotho <681 pg/mL. These results suggest that older adults with lower plasma klotho have poor skeletal muscle strength. PMID:21769735

Mutation in the Monocarboxylate Transporter 12 Gene Affects Guanidinoacetate Excretion but Does Not Cause Glucosuria.

PubMed

Dhayat, Nasser; Simonin, Alexandre; Anderegg, Manuel; Pathare, Ganesh; Lüscher, Benjamin P; Deisl, Christine; Albano, Giuseppe; Mordasini, David; Hediger, Matthias A; Surbek, Daniel V; Vogt, Bruno; Sass, Jörn Oliver; Kloeckener-Gruissem, Barbara; Fuster, Daniel G

2016-05-01

A heterozygous mutation (c.643C>A; p.Q215X) in the monocarboxylate transporter 12-encoding gene MCT12 (also known as SLC16A12) that mediates creatine transport was recently identified as the cause of a syndrome with juvenile cataracts, microcornea, and glucosuria in a single family. Whereas the MCT12 mutation cosegregated with the eye phenotype, poor correlation with the glucosuria phenotype did not support a pathogenic role of the mutation in the kidney. Here, we examined MCT12 in the kidney and found that it resides on basolateral membranes of proximal tubules. Patients with MCT12 mutation exhibited reduced plasma levels and increased fractional excretion of guanidinoacetate, but normal creatine levels, suggesting that MCT12 may function as a guanidinoacetate transporter in vivo However, functional studies in Xenopus oocytes revealed that MCT12 transports creatine but not its precursor, guanidinoacetate. Genetic analysis revealed a separate, undescribed heterozygous mutation (c.265G>A; p.A89T) in the sodium/glucose cotransporter 2-encoding gene SGLT2 (also known as SLC5A2) in the family that segregated with the renal glucosuria phenotype. When overexpressed in HEK293 cells, the mutant SGLT2 transporter did not efficiently translocate to the plasma membrane, and displayed greatly reduced transport activity. In summary, our data indicate that MCT12 functions as a basolateral exit pathway for creatine in the proximal tubule. Heterozygous mutation of MCT12 affects systemic levels and renal handling of guanidinoacetate, possibly through an indirect mechanism. Furthermore, our data reveal a digenic syndrome in the index family, with simultaneous MCT12 and SGLT2 mutation. Thus, glucosuria is not part of the MCT12 mutation syndrome. Copyright © 2016 by the American Society of Nephrology.

Molecular characterization of an adiponectin receptor homolog in the white leg shrimp, Litopenaeus vannamei

PubMed Central

Kim, Ah Ran; Alam, Md Jobaidul; Yoon, Tae-ho; Lee, Soo Rin; Park, Hyun; Kim, Doo-Nam; An, Doo-Hae; Lee, Jae-Bong; Lee, Chung Il

2016-01-01

Adiponectin (AdipoQ) and its receptors (AdipoRs) are strongly related to growth and development of skeletal muscle, as well as glucose and lipid metabolism in vertebrates. Herein we report the identification of the first full-length cDNA encoding an AdipoR homolog (Liv-AdipoR) from the decapod crustacean Litopenaeus vannamei using a combination of next generation sequencing (NGS) technology and bioinformatics analysis. The full-length Liv-AdipoR (1,245 bp) encoded a protein that exhibited the canonical seven transmembrane domains (7TMs) and the inversed topology that characterize members of the progestin and adipoQ receptor (PAQR) family. Based on the obtained sequence information, only a single orthologous AdipoR gene appears to exist in arthropods, whereas two paralogs, AdipoR1 and AdipoR2, have evolved in vertebrates. Transcriptional analysis suggested that the single Liv-AdipoR gene appears to serve the functions of two mammalian AdipoRs. At 72 h after injection of 50 pmol Liv-AdipoR dsRNA (340 bp) into L. vannamei thoracic muscle and deep abdominal muscle, transcription levels of Liv-AdipoR decreased by 93% and 97%, respectively. This confirmed optimal conditions for RNAi of Liv-AdipoR. Knockdown of Liv-AdipoR resulted in significant changes in the plasma levels of ammonia, 3-methylhistine, and ornithine, but not plasma glucose, suggesting that that Liv-AdipoR is important for maintaining muscle fibers. The chronic effect of Liv-AdipoR dsRNA injection was increased mortality. Transcriptomic analysis showed that 804 contigs were upregulated and 212 contigs were downregulated by the knockdown of Liv-AdipoR in deep abdominal muscle. The significantly upregulated genes were categorized as four main functional groups: RNA-editing and transcriptional regulators, molecular chaperones, metabolic regulators, and channel proteins. PMID:27478708

Intestinal Fluid Permeability in Atlantic Salmon (Salmo salar L.) Is Affected by Dietary Protein Source.

PubMed

Hu, Haibin; Kortner, Trond M; Gajardo, Karina; Chikwati, Elvis; Tinsley, John; Krogdahl, Åshild

2016-01-01

In Atlantic salmon (Salmo salar L.), and also in other fish species, certain plant protein ingredients can increase fecal water content creating a diarrhea-like condition which may impair gut function and reduce fish growth. The present study aimed to strengthen understanding of the underlying mechanisms by observing effects of various alternative plant protein sources when replacing fish meal on expression of genes encoding proteins playing key roles in regulation of water transport across the mucosa of the distal intestine (DI). A 48-day feeding trial was conducted with five diets: A reference diet (FM) in which fish meal (72%) was the only protein source; Diet SBMWG with a mix of soybean meal (30%) and wheat gluten (22%); Diet SPCPM with a mix of soy protein concentrate (30%) and poultry meal (6%); Diet GMWG with guar meal (30%) and wheat gluten (14.5%); Diet PM with 58% poultry meal. Compared to fish fed the FM reference diet, fish fed the soybean meal containing diet (SBMWG) showed signs of enteritis in the DI, increased fecal water content of DI chyme and higher plasma osmolality. Altered DI expression of a battery of genes encoding aquaporins, ion transporters, tight junction and adherens junction proteins suggested reduced transcellular transport of water as well as a tightening of the junction barrier in fish fed the SBMWG diet, which may explain the observed higher fecal water content and plasma osmolality. DI structure was not altered for fish fed the other experimental diets but alterations in target gene expression and fecal water content were observed, indicating that alterations in water transport components may take place without clear effects on intestinal structure.

The effect of a genetically reduced plasma membrane protonmotive force on vegetative growth of Arabidopsis.

PubMed

Haruta, Miyoshi; Sussman, Michael R

2012-03-01

The plasma membrane proton gradient is an essential feature of plant cells. In Arabidopsis (Arabidopsis thaliana), this gradient is generated by the plasma membrane proton pump encoded by a family of 11 genes (abbreviated as AHA, for Arabidopsis H(+)-ATPase), of which AHA1 and AHA2 are the two most predominantly expressed in seedlings and adult plants. Although double knockdown mutant plants containing T-DNA insertions in both genes are embryonic lethal, under ideal laboratory growth conditions, single knockdown mutant plants with a 50% reduction in proton pump concentration complete their life cycle without any observable growth alteration. However, when grown under conditions that induce stress on the plasma membrane protonmotive force (PMF), such as high external potassium to reduce the electrical gradient or high external pH to reduce the proton chemical gradient, aha2 mutant plants show a growth retardation compared with wild-type plants. In this report, we describe the results of studies that examine in greater detail AHA2's specific role in maintaining the PMF during seedling growth. By comparing the wild type and aha2 mutants, we have measured the effects of a reduced PMF on root and hypocotyl growth, ATP-induced skewed root growth, and rapid cytoplasmic calcium spiking. In addition, genome-wide gene expression profiling revealed the up-regulation of potassium transporters in aha2 mutants, indicating, as predicted, a close link between the PMF and potassium uptake at the plasma membrane. Overall, this characterization of aha2 mutants provides an experimental and theoretical framework for investigating growth and signaling processes that are mediated by PMF-coupled energetics at the cell membrane.

Genetics of Triglycerides and the Risk of Atherosclerosis.

PubMed

Dron, Jacqueline S; Hegele, Robert A

2017-07-01

Plasma triglycerides are routinely measured with a lipid profile, and elevated plasma triglycerides are commonly encountered in the clinic. The confounded nature of this trait, which is correlated with numerous other metabolic perturbations, including depressed high-density lipoprotein cholesterol (HDL-C), has thwarted efforts to directly implicate triglycerides as causal in atherogenesis. Human genetic approaches involving large-scale populations and high-throughput genomic assessment under a Mendelian randomization framework have undertaken to sort out questions of causality. We review recent large-scale meta-analyses of cohorts and population-based sequencing studies designed to address whether common and rare variants in genes whose products are determinants of plasma triglycerides are also associated with clinical cardiovascular endpoints. The studied loci include genes encoding lipoprotein lipase and proteins that interact with it, such as apolipoprotein (apo) A-V, apo C-III and angiopoietin-like proteins 3 and 4, and common polymorphisms identified in genome-wide association studies. Triglyceride-raising variant alleles of these genes showed generally strong associations with clinical cardiovascular endpoints. However, in most cases, a second lipid disturbance-usually depressed HDL-C-was concurrently associated. While the findings collectively shift our understanding towards a potential causal role for triglycerides, we still cannot rule out the possibilities that triglycerides are a component of a joint phenotype with low HDL-C or that they are but markers of deeper causal metabolic disturbances that are not routinely measured in epidemiological-scale genetic studies.

Correlation of HAMP gene polymorphisms and expression with the susceptibility and length of hospital stays in Taiwanese children with Kawasaki disease

PubMed Central

Lu, Hsing-Fang; Wong, Henry Sung-Ching; Yu, Hong-Ren; Kuo, Hsing-Chun; Huang, Fu-Chen; Lo, Mao-Hung; Hsieh, Kai-Sheng; Chen, Su-Fen; Chang, Wei-Chiao; Kuo, Ho-Chang

2017-01-01

Kawasaki disease (KD) is a form of systemic vasculitis. Regarding its pathogenesis, HAMP gene encoding hepcidin, which is significant for iron metabolism, has a vital function. In this study, we recruited a total of 381 KD patients for genotyping. Data from 997 subjects (500 subjects from cohort 1; 497 subjects from cohort 2) were used for analysis. Using TaqMan allelic discrimination, we determined five tag SNPs (rs916145, rs10421768, rs3817623, rs7251432, and rs2293689). Treatment outcome data related to such clinical phenotypes as coronary artery lesions (CAL), coronary artery aneurysms (CAA), and intravenous immunoglobulin (IVIG) effects were also collected. Furthermore, we measured plasma hepcidin levels with an enzyme-linked immunosorbent assay. We found that HAMP gene polymorphism (rs7251432, and rs2293689) was significantly correlated with KD risk and that plasma hepcidin levels both before and after IVIG treatment had a significantly positive correlation with length of hospital stays (R = 0.217, p = 0.046 and R = 0.381, p < 0.0001, respectively). In contrast, plasma hepcidin levels has a negative correlation with KD patients’ albumin levels (R = −0.27, p < 0.001) prior to IVIG treatment. This study's findings indicate that HAMP might have a role in the disease susceptibility, as well as its expressions correlated length of hospital stays, and albumin levels in Taiwanese children with KD. PMID:28881695

Characterization of the response to zinc deficiency in the cyanobacterium Anabaena sp. strain PCC 7120.

PubMed

Napolitano, Mauro; Rubio, Miguel Ángel; Santamaría-Gómez, Javier; Olmedo-Verd, Elvira; Robinson, Nigel J; Luque, Ignacio

2012-05-01

Zur regulators control zinc homeostasis by repressing target genes under zinc-sufficient conditions in a wide variety of bacteria. This paper describes how part of a survey of duplicated genes led to the identification of the open reading frame all2473 as the gene encoding the Zur regulator of the cyanobacterium Anabaena sp. strain PCC 7120. All2473 binds to DNA in a zinc-dependent manner, and its DNA-binding sequence was characterized, which allowed us to determine the relative contribution of particular nucleotides to Zur binding. A zur mutant was found to be impaired in the regulation of zinc homeostasis, showing sensitivity to elevated concentrations of zinc but not other metals. In an effort to characterize the Zur regulon in Anabaena, 23 genes containing upstream putative Zur-binding sequences were identified and found to be regulated by Zur. These genes are organized in six single transcriptional units and six operons, some of them containing multiple Zur-regulated promoters. The identities of genes of the Zur regulon indicate that Anabaena adapts to conditions of zinc deficiency by replacing zinc metalloproteins with paralogues that fulfill the same function but presumably with a lower zinc demand, and with inducing putative metallochaperones and membrane transport systems likely being involved in the scavenging of extracellular zinc, including plasma membrane ABC transport systems and outer membrane TonB-dependent receptors. Among the Zur-regulated genes, the ones showing the highest induction level encode proteins of the outer membrane, suggesting a primary role for components of this cell compartment in the capture of zinc cations from the extracellular medium.

Mollusk genes encoding lysine tRNA (UUU) contain introns.

PubMed

Matsuo, M; Abe, Y; Saruta, Y; Okada, N

1995-11-20

New intron-containing genes encoding tRNAs were discovered when genomic DNA isolated from various animal species was amplified by the polymerase chain reaction (PCR) with primers based on sequences of rabbit tRNA(Lys). From sequencing analysis of the products of PCR, we found that introns are present in several genes encoding tRNA(Lys) in mollusks, such as Loligo bleekeri (squid) and Octopus vulgaris (octopus). These introns were specific to genes encoding tRNA(Lys)(CUU) and were not present in genes encoding tRNA(Lys)(CUU). In addition, the sequences of the introns were different from one another. To confirm the results of our initial experiments, we isolated and sequenced genes encoding tRNA(Lys)(CUU) and tRNA(Lys)(UUU). The gene for tRNA(Lys)(UUU) from squid contained an intron, whose sequence was the same as that identified by PCR, and the gene formed a cluster with a corresponding pseudogene. Several DNA regions of 2.1 kb containing this cluster appeared to be tandemly arrayed in the squid genome. By contrast, the gene encoding tRNA(Lys)(CUU) did not contain an intron, as shown also by PCR. The tRNA(Lys)(UUU) that corresponded to the analyzed gene was isolated and characterized. The present study provides the first example of an intron-containing gene encoding a tRNA in mollusks and suggests the universality of introns in such genes in higher eukaryotes.

Human AZU-1 gene, variants thereof and expressed gene products

DOEpatents

Chen, Huei-Mei; Bissell, Mina

2004-06-22

A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

The mitochondrial gene encoding ribosomal protein S12 has been translocated to the nuclear genome in Oenothera.

PubMed Central

Grohmann, L; Brennicke, A; Schuster, W

1992-01-01

The Oenothera mitochondrial genome contains only a gene fragment for ribosomal protein S12 (rps12), while other plants encode a functional gene in the mitochondrion. The complete Oenothera rps12 gene is located in the nucleus. The transit sequence necessary to target this protein to the mitochondrion is encoded by a 5'-extension of the open reading frame. Comparison of the amino acid sequence encoded by the nuclear gene with the polypeptides encoded by edited mitochondrial cDNA and genomic sequences of other plants suggests that gene transfer between mitochondrion and nucleus started from edited mitochondrial RNA molecules. Mechanisms and requirements of gene transfer and activation are discussed. Images PMID:1454526

Common skate (Raja kenojei) secretes pentraxin into the cutaneous secretion: The first skin mucus lectin in cartilaginous fish.

PubMed

Tsutsui, Shigeyuki; Yamaguchi, Motoki; Hirasawa, Ai; Nakamura, Osamu; Watanabe, Tasuku

2009-08-01

A lactose-specific lectin with a molecular mass of about 25 kDa was purified from the skin mucus of a cartilaginous fish-the common skate (Raja kenojei). The complementary DNA sequence of the lectin was 1540 bp long and contained a reading frame encoding 226 amino acids, which showed approximately 38% identity to pentraxins of mammals and teleosts. Gene expression was observed in the skin, gill, stomach and intestine in the healthy skate. We also identified an isotype gene from the liver whose deduced amino-acid sequence shared 69.0% identity with the skin type gene. The antiserum detected protein in the skin, where the lectin is localized in the epidermal cells, and in the blood plasma. The lectin genes are multicopied in the common skate genome. Although pentraxins are acute phase proteins, mRNAs of both the isotypes were not upregulated after the in vivo challenge with formalin-killed Escherichia coli, which suggests that they are constantly present in the skin mucus and blood plasma to protect against pathogenic invasion. This lectin is the fifth type of lectin found in the cutaneous secretions of fish, demonstrating that skin mucus lectins have evolved with marked molecular diversity in fish.

Digital PCR analysis of maternal plasma for noninvasive detection of sickle cell anemia.

PubMed

Barrett, Angela N; McDonnell, Thomas C R; Chan, K C Allen; Chitty, Lyn S

2012-06-01

Cell-free fetal DNA (cffDNA) constitutes approximately 10% of the cell-free DNA in maternal plasma and is a suitable source of fetal genetic material for noninvasive prenatal diagnosis (NIPD). The objective of this study was to determine the feasibility of using digital PCR for NIPD in pregnancies at risk of sickle cell anemia. Minor-groove binder (MGB) TaqMan probes were designed to discriminate between wild-type hemoglobin A and mutant (hemoglobin S) alleles encoded by the HBB (hemoglobin, beta) gene in cffDNA isolated from maternal plasma samples obtained from pregnancies at risk of sickle cell anemia. The fractional fetal DNA concentration was assessed in male-bearing pregnancies with a digital PCR assay for the Y chromosome-specific marker DYS14. In pregnancies with a female fetus, a panel of biallelic insertion/deletion polymorphism (indel) markers was developed for the quantification of the fetal DNA fraction. We used digital real-time PCR to analyze the dosage of the variant encoding hemoglobin S relative to that encoding wild-type hemoglobin A. The sickle cell genotype was correctly determined in 82% (37 of 45) of male fetuses and 75% (15 of 20) of female fetuses. Mutation status was determined correctly in 100% of the cases (25 samples) with fractional fetal DNA concentrations >7%. The panel of indels was informative in 65% of the female-bearing pregnancies. Digital PCR can be used to determine the genotype of fetuses at risk for sickle cell anemia. Optimization of the fractional fetal DNA concentration is essential. More-informative indel markers are needed for this assay's comprehensive use in cases of a female fetus.

Gene-diet interactions with polymorphisms of the MGLL gene on plasma low-density lipoprotein cholesterol and size following an omega-3 polyunsaturated fatty acid supplementation: a clinical trial.

PubMed

Ouellette, Catherine; Rudkowska, Iwona; Lemieux, Simone; Lamarche, Benoit; Couture, Patrick; Vohl, Marie-Claude

2014-05-24

Omega-3 (n-3) polyunsaturated fatty acid (PUFA) consumption increases low-density lipoprotein (LDL) cholesterol (C) concentrations and particle size. Studies showed that individuals with large, buoyant LDL particles have decreased risk of cardiovascular diseases. However, a large inter-individual variability is observed in LDL particle size. Genetic factors may explain the variability of LDL-C concentrations and particle size after an n-3 PUFA supplementation. The monoglyceride lipase (MGLL) enzyme, encoded by the MGLL gene, plays an important role in lipid metabolism, especially lipoprotein metabolism. The aim of this study was to investigate if polymorphisms (SNPs) of the MGLL gene influence the variability of LDL-C and LDL particle size in response to an n-3 PUFA supplementation. 210 subjects completed the study. They consumed 5 g/d of a fish oil supplement (1.9-2.2 g eicosapentaenoic acid and 1.1 g docosaexaenoic acid) during 6 weeks. Plasma lipids were measured before and after the supplementation period and 18 SNPs of the MGLL gene, covering 100% of common genetic variations (minor allele frequency ≥0.05), have been genotyped using TaqMan technology (Life Technologies Inc., Burlington, ON, CA). Following the n-3 PUFA supplementation, 55% of subjects increased their LDL-C levels. In a model including the supplementation, genotype and supplementation*genotype effects, gene-diet interaction effects on LDL-C concentrations (rs782440, rs6776142, rs555183, rs6780384, rs6787155 and rs1466571) and LDL particle size (rs9877819 and rs13076593) were observed for the MGLL gene SNPs (p < 0.05). SNPs within the MGLL gene may modulate plasma LDL-C levels and particle size following an n-3 PUFA supplementation. This trial was registered at clinicaltrials.gov as NCT01343342.

The Effect of Selenium Supplementation on Glucose Homeostasis and the Expression of Genes Related to Glucose Metabolism.

PubMed

Jablonska, Ewa; Reszka, Edyta; Gromadzinska, Jolanta; Wieczorek, Edyta; Krol, Magdalena B; Raimondi, Sara; Socha, Katarzyna; Borawska, Maria H; Wasowicz, Wojciech

2016-12-13

The aim of the study was to evaluate the effect of selenium supplementation on the expression of genes associated with glucose metabolism in humans, in order to explain the unclear relationship between selenium and the risk of diabetes. For gene expression analysis we used archival samples of cDNA from 76 non-diabetic subjects supplemented with selenium in the previous study. The supplementation period was six weeks and the daily dose of selenium was 200 µg (as selenium yeast). Blood for mRNA isolation was collected at four time points: before supplementation, after two and four weeks of supplementation, and after four weeks of washout. The analysis included 15 genes encoding selected proteins involved in insulin signaling and glucose metabolism. In addition, HbA1c and fasting plasma glucose were measured at three and four time points, respectively. Selenium supplementation was associated with a significantly decreased level of HbA1c but not fasting plasma glucose (FPG) and significant down-regulation of seven genes: INSR , ADIPOR1 , LDHA , PDHA , PDHB , MYC , and HIF1AN . These results suggest that selenium may affect glycemic control at different levels of regulation, linked to insulin signaling, glycolysis, and pyruvate metabolism. Further research is needed to investigate mechanisms of such transcriptional regulation and its potential implication in direct metabolic effects.

Perturbations of carotenoid and tetrapyrrole biosynthetic pathways result in differential alterations in chloroplast function and plastid signaling.

PubMed

Park, Joon-Heum; Jung, Sunyo

2017-01-22

In this study, we used the biosynthetic inhibitors of carotenoid and tetrapyrrole biosynthetic pathways, norflurazon (NF) and oxyfluorfen (OF), as tools to gain insight into mechanisms of photooxidation in rice plants. NF resulted in bleaching symptom on leaves of the treated plants, whereas OF treatment developed a fast symptom of an apparent necrotic phenotype. Both plants exhibited decreases in photosynthetic efficiency, as indicated by F v /F m . NF caused severe disruption in thylakoid membranes, whereas OF-treated plants exhibited disruption of chloroplast envelope and plasma membrane. Levels of Lhca and Lhcb proteins in photosystem I (PSI) and PSII were reduced by photooxidative stress in NF- and OF-treated plants, with a greater decrease in NF plants. The down-regulation of nuclear-encoded photosynthesis genes Lhcb and rbcS was also found in both NF- and OF-treated plants, whereas plastid-encoded photosynthetic genes including RbcL, PsaC, and PsbD accumulated normally in NF plants but decreased drastically in OF plants. This proposes that the plastids in NF plants retain their potential to develop thylakoid membranes and that photobleaching is mainly controlled by nuclear genes. Distinct photooxidation patterns between NF- and OF-treated plants developed differential signaling, which might enable the plant to coordinate the expression of photosynthetic genes from the nuclear and plastidic genomes. Copyright © 2016 Elsevier Inc. All rights reserved.

Influence of IL1B, IL6 and IL10 gene variants and plasma fatty acid interaction on metabolic syndrome risk in a cross-sectional population-based study.

PubMed

Maintinguer Norde, Marina; Oki, Erica; Ferreira Carioca, Antonio Augusto; Teixeira Damasceno, Nágila Raquel; Fisberg, Regina Mara; Lobo Marchioni, Dirce Maria; Rogero, Marcelo Macedo

2018-04-01

Metabolic syndrome (MetS) is a cluster of interrelated risk factors for type 2 diabetes mellitus, and cardiovascular disease, with underlying inflammatory pathophysiology. Genetic variations and diet are well-known risk factor for MetS, but the interaction between these two factors is less explored. The aim of the study was to evaluate the influence of interaction between SNP of inflammatory genes (encoding interleukin (IL)-6, IL-1β and IL-10) and plasma fatty acids on the odds of MetS, in a population-based cross-sectional study. Among participants of the Health Survey - São Paulo, 301 adults (19-59 y) from whom a blood sample was collected were included. Individuals with and without MetS were compared according to their plasma inflammatory biomarkers, fatty acid profile, and genotype frequency of the IL1B (rs16944, rs1143623, rs1143627, rs1143634 and rs1143643), IL6 (rs1800795, rs1800796 and rs1800797) and IL10 (rs1554286, rs1800871, rs1800872, rs1800890 and rs3024490) genes SNP. The influence of gene-fatty acids interaction on MetS risk was investigated. IL6 gene SNP rs1800795 G allele was associated with higher odds for MetS (OR = 1.88; p = 0.017). Gene-fatty acid interaction was found between the IL1B gene SNP rs116944 and stearic acid (p inter = 0.043), and between rs1143634 and EPA (p inter = 0.017). For the IL10 gene SNP rs1800896, an interaction was found for arachidonic acid (p inter = 0.007) and estimated D5D activity (p inter = 0.019). The IL6 gene SNP rs1800795 G allele is associated with increased odds for MetS. Plasma fatty acid profile interacts with the IL1B and IL10 gene variants to modulate the odds for MetS. This and other interactions of risk factors can account for the unexplained heritability of MetS, and their elucidation can lead to new strategies for genome-customized prevention of MetS. Copyright © 2017 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Cloning and characterization of aquaglyceroporin genes from rainbow smelt (Osmerus mordax) and transcript expression in response to cold temperature.

PubMed

Hall, Jennifer R; Clow, Kathy A; Rise, Matthew L; Driedzic, William R

2015-09-01

Aquaglyceroporins (GLPs) are integral membrane proteins that facilitate passive movement of water, glycerol and urea across cellular membranes. In this study, GLP-encoding genes were characterized in rainbow smelt (Osmerus mordax mordax), an anadromous teleost that accumulates high glycerol and modest urea levels in plasma and tissues as an adaptive cryoprotectant mechanism in sub-zero temperatures. We report the gene and promoter sequences for two aqp10b paralogs (aqp10ba, aqp10bb) that are 82% identical at the predicted amino acid level, and aqp9b. Aqp10bb and aqp9b have the 6 exon structure common to vertebrate GLPs. Aqp10ba has 8 exons; there are two additional exons at the 5' end, and the promoter sequence is different from aqp10bb. Molecular phylogenetic analysis suggests that the aqp10b paralogs arose from a gene duplication event specific to the smelt lineage. Smelt GLP transcripts are ubiquitously expressed; however, aqp10ba transcripts were highest in kidney, aqp10bb transcripts were highest in kidney, intestine, pyloric caeca and brain, and aqp9b transcripts were highest in spleen, liver, red blood cells and kidney. In cold-temperature challenge experiments, plasma glycerol and urea levels were significantly higher in cold- compared to warm-acclimated smelt; however, GLP transcript levels were generally either significantly lower or remained constant. The exception was significantly higher aqp10ba transcript levels in kidney. High aqp10ba transcripts in smelt kidney that increase significantly in response to cold temperature in congruence with plasma urea suggest that this gene duplicate may have evolved to allow the re-absorption of urea to concomitantly conserve nitrogen and prevent freezing. Copyright © 2015 Elsevier Inc. All rights reserved.

Identification of a Nicotiana plumbaginifolia plasma membrane H(+)-ATPase gene expressed in the pollen tube.

PubMed

Lefebvre, Benoit; Arango, Miguel; Oufattole, Mohammed; Crouzet, Jérôme; Purnelle, Bénédicte; Boutry, Marc

2005-08-01

In Nicotiana plumbaginifolia, plasma membrane H(+)-ATPases (PMAs) are encoded by a gene family of nine members. Here, we report on the characterization of a new isogene, NpPMA5 (belonging to subfamily IV), and the determination of its expression pattern using the beta-glucuronidase (gusA) reporter gene. pNpPMA5-gusA was expressed in cotyledons, in vascular tissues of the stem (mainly in nodal zones), and in the flower and fruit. In the flower, high expression was found in the pollen tube after in vitro or in vivo germination. Northern blotting analysis confirmed that NpPMA5 was expressed in the pollen tube contrary to NpPMA2 (subfamily I) or NpPMA4 (subfamily II), two genes highly expressed in other tissues. The subcellular localization of PM H(+)-ATPase in the pollen tube was analyzed by immunocytodecoration. As expected, this enzyme was localized to the plasma membrane. However, neither the tip nor the base of the pollen tube was labeled, showing an asymmetrical distribution of this enzyme. This observation supports the hypothesis that the PM H(+)-ATPase is involved in creating the pH gradient that is observed along the pollen tube and is implicated in cell elongation. Compared to other plant PM H(+)-ATPases, the C-terminal region of NpPMA5 is shorter by 26 amino acid residues and is modified in the last 6 residues, due to a sequence rearrangement, which was also found in the orthologous gene of Nicotiana glutinosa, a Nicotiana species distant from N. plumbaginifolia and Petunia hybrida and Lycopersicon esculentum, other Solanacae species. This modification alters part of the PM H(+)-ATPase regulatory domain and raises the question whether this isoform is still regulated.

Accumulation of 5-oxoproline in myocardial dysfunction and the protective effects of OPLAH.

PubMed

van der Pol, Atze; Gil, Andres; Silljé, Herman H W; Tromp, Jasper; Ovchinnikova, Ekaterina S; Vreeswijk-Baudoin, Inge; Hoes, Martijn; Domian, Ibrahim J; van de Sluis, Bart; van Deursen, Jan M; Voors, Adriaan A; van Veldhuisen, Dirk J; van Gilst, Wiek H; Berezikov, Eugene; van der Harst, Pim; de Boer, Rudolf A; Bischoff, Rainer; van der Meer, Peter

2017-11-08

In response to heart failure (HF), the heart reacts by repressing adult genes and expressing fetal genes, thereby returning to a more fetal-like gene profile. To identify genes involved in this process, we carried out transcriptional analysis on murine hearts at different stages of development and on hearts from adult mice with HF. Our screen identified Oplah , encoding for 5-oxoprolinase, a member of the γ-glutamyl cycle that functions by scavenging 5-oxoproline. OPLAH depletion occurred as a result of cardiac injury, leading to elevated 5-oxoproline and oxidative stress, whereas OPLAH overexpression improved cardiac function after ischemic injury. In HF patients, we observed elevated plasma 5-oxoproline, which was associated with a worse clinical outcome. Understanding and modulating fetal-like genes in the failing heart may lead to potential diagnostic, prognostic, and therapeutic options in HF. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

PPIA rs6850: A > G single-nucleotide polymorphism is associated with raised plasma cyclophilin A levels in patients with coronary artery disease.

PubMed

Vinitha, A; Kutty, V Raman; Vivekanand, A; Reshmi, G; Divya, G; Sumi, S; Santosh, K R; Pratapachandran, N S; Ajit, Mullassari S; Kartha, C C; Ramachandran, Surya

2016-01-01

Plasma level of cyclophilin A is a promising marker of vascular disease in patients with type 2 diabetes. Genetic variants in the peptidylprolyl isomerase A gene, encoding human cyclophilin may alter protein synthesis thus affecting its activity, function, and circulating plasma levels. We examined the effect of single-nucleotide polymorphisms (SNPs) within the PPIA gene on plasma levels of cyclophilin A and coupled this with status of vascular disease in patients with and without type 2 diabetes in 212 South Indian subjects. The regulatory region of PPIA gene was sequenced for SNPs. The association of SNPs with known blood markers of type 2 diabetes and coronary artery disease such as HbA1c, low- and high-density lipoproteins, triglycerides, fasting and postprandial blood sugar levels, and cyclophilin A were probed. We identified three SNPs namely, rs6850: A > G; (AG/-) c.*227_*228delAG and (-/T) c.*318_*319insT. Welchs two-sample t test indicated an association of SNP rs6850: A > G, located at the 5' UTR region with increased plasma levels of cyclophilin A in patients with coronary artery disease and with coronary artery disease associated with diabetes. The presence of rs6850: A > G variant was significantly associated with coronary artery disease irrespective of whether the patients had diabetes or not. In silico analysis of the sequence using different tools and matrix libraries did not predict any significant differential binding sites for rs6850: A > G, c.*227_*228delAG and c.*318_*319insT. Our results indicate that the SNP rs6850: A > G is associated with increased risk for elevated plasma levels of cyclophilin A and coronary artery disease in patients with and without type 2 diabetes.

Associations between oxytocin receptor gene (OXTR) methylation, plasma oxytocin, and attachment across adulthood.

PubMed

Ebner, Natalie C; Lin, Tian; Muradoglu, Melis; Weir, Devon H; Plasencia, Gabriela M; Lillard, Travis S; Pournajafi-Nazarloo, Hossein; Cohen, Ronald A; Sue Carter, C; Connelly, Jessica J

2018-02-02

The neuropeptide oxytocin (OT) has been implicated in a wide range of affiliative processes. OT exerts its functions via OT receptors, which are encoded by the oxytocin receptor gene (OXTR). Epigenetic modification of OXTR through the process of DNA methylation has been associated with individual differences in behavioral phenotypes. Specifically, lower levels of OXTR methylation have been linked to better social and affective functioning. However, research on epigenetic mechanisms of OXTR is scarce in non-clinical populations, and even less is known about epigenetic variability across adulthood. The present study assessed methylation levels at OXTR CpG site -934 and plasma OT levels in 22 young (20-31 years, M = 23.6) and 34 older (63-80 years, M = 71.4) participants. Lower levels of OXTR methylation and higher plasma OT levels were associated with less self-reported attachment anxiety in young but not older participants, with largely independent contributions of OXTR methylation and plasma OT levels. In contrast, in the overall sample, lower levels of OXTR methylation were associated with higher self-reported attachment avoidance. Age analysis suggested that these results were largely driven by young adults. Plasma OT levels were unrelated to attachment avoidance. Taken together, these findings support the emerging notion in the literature that epigenetic properties of OXTR, in addition to endogenous OT levels, are related to adult attachment. Further, the age effects observed in the associations between OXTR methylation, plasma OT, and adult attachment emphasize the importance of adopting a developmental perspective when studying properties of the OT system and their relation to affiliative processes. Findings contribute to growing evidence suggesting that epigenetic modification of genes regulating OT pathways and endogenous OT levels are associated with the way people form and maintain intimate social relationships. Copyright © 2018 Elsevier B.V. All rights reserved.

The organization of the fuc regulon specifying L-fucose dissimilation in Escherichia coli K12 as determined by gene cloning.

PubMed

Chen, Y M; Zhu, Y; Lin, E C

1987-12-01

In Escherichia coli the six known genes specifying the utilization of L-fucose as carbon and energy source cluster at 60.2 min and constitute a regulon. These genes include fucP (encoding L-fucose permease), fucI (encoding L-fucose isomerase), fucK (encoding L-fuculose kinase), fucA (encoding L-fuculose 1-phosphate aldolase), fucO (encoding L-1,2-propanediol oxidoreductase), and fucR (encoding the regulatory protein). In this study the fuc genes were cloned and their positions on the chromosome were established by restriction endonuclease and complementation analyses. Clockwise, the gene order is: fucO-fucA-fucP-fucI-fucK-fucR. The operons comprising the structural genes and the direction of transcription were determined by complementation analysis and Southern blot hybridization. The fucPIK and fucA operons are transcribed clockwise. The fucO operon is transcribed counterclockwise. The fucR gene product activates the three structural operons in trans.

The PaAlr1 magnesium transporter is required for ascospore development in Podospora anserina.

PubMed

Grognet, Pierre; Lalucque, Hervé; Silar, Philippe

2012-10-01

The PaAlr1 gene encoding a putative plasma membrane magnesium (Mg) transporter in Podospora anserina was inactivated. The PaAlr1(Δ) mutants showed sensitivity to deprivation and excess Mg(2+) and Ca(2+). They also exhibited an autonomous ascospore maturation defect. Mutant ascospores were arrested at an early stage when they contained two nuclei. These data emphasize the role of Mg ions during sexual development in a filamentous fungus. Copyright © 2012 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

A role for the peroxisomal 3-ketoacyl-CoA thiolase B enzyme in the control of PPARα-mediated upregulation of SREBP-2 target genes in the liver.

PubMed

Fidaleo, Marco; Arnauld, Ségolène; Clémencet, Marie-Claude; Chevillard, Grégory; Royer, Marie-Charlotte; De Bruycker, Melina; Wanders, Ronald J A; Athias, Anne; Gresti, Joseph; Clouet, Pierre; Degrace, Pascal; Kersten, Sander; Espeel, Marc; Latruffe, Norbert; Nicolas-Francès, Valérie; Mandard, Stéphane

2011-05-01

Peroxisomal 3-ketoacyl-CoA thiolase B (Thb) catalyzes the final step in the peroxisomal β-oxidation of straight-chain acyl-CoAs and is under the transcription control of the nuclear hormone receptor PPARα. PPARα binds to and is activated by the synthetic compound Wy14,643 (Wy). Here, we show that the magnitude of Wy-mediated induction of peroxisomal β-oxidation of radiolabeled (1-(14)C) palmitate was significantly reduced in mice deficient for Thb. In contrast, mitochondrial β-oxidation was unaltered in Thb(-/-) mice. Given that Wy-treatment induced Acox1 and MFP-1/-2 activity at a similar level in both genotypes, we concluded that the thiolase step alone was responsible for the reduced peroxisomal β-oxidation of fatty acids. Electron microscopic analysis and cytochemical localization of catalase indicated that peroxisome proliferation in the liver after Wy-treatment was normal in Thb(-/-) mice. Intriguingly, micro-array analysis revealed that mRNA levels of genes encoding cholesterol biosynthesis enzymes were upregulated by Wy in Wild-Type (WT) mice but not in Thb(-/-) mice, which was confirmed at the protein level for the selected genes. The non-induction of genes encoding cholesterol biosynthesis enzymes by Wy in Thb(-/-) mice appeared to be unrelated to defective SREBP-2 or PPARα signaling. No difference was observed in the plasma lathosterol/cholesterol ratio (a marker for de novo cholesterol biosynthesis) between Wy-treated WT and Thb(-/-) mice, suggesting functional compensation. Overall, we conclude that ThA and SCPx/SCP2 thiolases cannot fully compensate for the absence of ThB. In addition, our data indicate that ThB is involved in the regulation of genes encoding cholesterol biosynthesis enzymes in the liver, suggesting that the peroxisome could be a promising candidate for the correction of cholesterol imbalance in dyslipidemia. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Pharmacogenetics of novel oral anticoagulants: a review of identified gene variants & future perspectives.

PubMed

Ašić, Adna; Marjanović, Damir; Mirat, Jure; Primorac, Dragan

2018-05-16

Novel oral anticoagulants (NOACs) are becoming a therapy of choice in everyday clinical practice after almost 50 years during which warfarin and related coumarin derivatives were used as the main anticoagulants. Advantages of NOACs over standard anticoagulants include their predictable pharmacodynamics and pharmacokinetics, stable plasma concentrations and less drug-drug and food-drug interactions. However, pharmacogenetics has its place in administration of NOACs, as considerable interindividual variations have been detected. In this review, previous findings in pharmacogenetics of dabigatran, rivaroxaban, apixaban and edoxaban are summarized, along with recommendations for studying genes encoding metabolically important enzymes for four selected NOACs. Future directions include identification of clinically relevant SNPs, and change in optimum dosage for patients who are carriers of significant variants.

Differential Responses of Plasma Adropin Concentrations To Dietary Glucose or Fructose Consumption In Humans.

PubMed

Butler, Andrew A; St-Onge, Marie-Pierre; Siebert, Emily A; Medici, Valentina; Stanhope, Kimber L; Havel, Peter J

2015-10-05

Adropin is a peptide hormone encoded by the Energy Homeostasis Associated (ENHO) gene whose physiological role in humans remains incompletely defined. Here we investigated the impact of dietary interventions that affect systemic glucose and lipid metabolism on plasma adropin concentrations in humans. Consumption of glucose or fructose as 25% of daily energy requirements (E) differentially affected plasma adropin concentrations (P < 0.005) irrespective of duration, sex or age. Glucose consumption reduced plasma adropin from 3.55 ± 0.26 to 3.28 ± 0.23 ng/ml (N = 42). Fructose consumption increased plasma adropin from 3.63 ± 0.29 to 3.93 ± 0.34 ng/ml (N = 45). Consumption of high fructose corn syrup (HFCS) as 25% E had no effect (3.43 ± 0.32 versus 3.39 ± 0.24 ng/ml, N = 26). Overall, the effect of glucose, HFCS and fructose on circulating adropin concentrations were similar to those observed on postprandial plasma triglyceride concentrations. Furthermore, increases in plasma adropin levels with fructose intake were most robust in individuals exhibiting hypertriglyceridemia. Individuals with low plasma adropin concentrations also exhibited rapid increases in plasma levels following consumption of breakfasts supplemented with lipids. These are the first results linking plasma adropin levels with dietary sugar intake in humans, with the impact of fructose consumption linked to systemic triglyceride metabolism. In addition, dietary fat intake may also increase circulating adropin concentrations.

Evolutionary Characteristics of Missing Proteins: Insights into the Evolution of Human Chromosomes Related to Missing-Protein-Encoding Genes.

PubMed

Xu, Aishi; Li, Guang; Yang, Dong; Wu, Songfeng; Ouyang, Hongsheng; Xu, Ping; He, Fuchu

2015-12-04

Although the "missing protein" is a temporary concept in C-HPP, the biological information for their "missing" could be an important clue in evolutionary studies. Here we classified missing-protein-encoding genes into two groups, the genes encoding PE2 proteins (with transcript evidence) and the genes encoding PE3/4 proteins (with no transcript evidence). These missing-protein-encoding genes distribute unevenly among different chromosomes, chromosomal regions, or gene clusters. In the view of evolutionary features, PE3/4 genes tend to be young, spreading at the nonhomology chromosomal regions and evolving at higher rates. Interestingly, there is a higher proportion of singletons in PE3/4 genes than the proportion of singletons in all genes (background) and OTCSGs (organ, tissue, cell type-specific genes). More importantly, most of the paralogous PE3/4 genes belong to the newly duplicated members of the paralogous gene groups, which mainly contribute to special biological functions, such as "smell perception". These functions are heavily restricted into specific type of cells, tissues, or specific developmental stages, acting as the new functional requirements that facilitated the emergence of the missing-protein-encoding genes during evolution. In addition, the criteria for the extremely special physical-chemical proteins were first set up based on the properties of PE2 proteins, and the evolutionary characteristics of those proteins were explored. Overall, the evolutionary analyses of missing-protein-encoding genes are expected to be highly instructive for proteomics and functional studies in the future.

XTH31, Encoding an in Vitro XEH/XET-Active Enzyme, Regulates Aluminum Sensitivity by Modulating in Vivo XET Action, Cell Wall Xyloglucan Content, and Aluminum Binding Capacity in Arabidopsis[W

PubMed Central

Zhu, Xiao Fang; Shi, Yuan Zhi; Lei, Gui Jie; Fry, Stephen C.; Zhang, Bao Cai; Zhou, Yi Hua; Braam, Janet; Jiang, Tao; Xu, Xiao Yan; Mao, Chuan Zao; Pan, Yuan Jiang; Yang, Jian Li; Wu, Ping; Zheng, Shao Jian

2012-01-01

Xyloglucan endohydrolase (XEH) and xyloglucan endotransglucosylase (XET) activities, encoded by xyloglucan endotransglucosylase-hydrolase (XTH) genes, are involved in cell wall extension by cutting or cutting and rejoining xyloglucan chains, respectively. However, the physiological significance of this biochemical activity remains incompletely understood. Here, we find that an XTH31 T-DNA insertion mutant, xth31, is more Al resistant than the wild type. XTH31 is bound to the plasma membrane and the encoding gene is expressed in the root elongation zone and in nascent leaves, suggesting a role in cell expansion. XTH31 transcript accumulation is strongly downregulated by Al treatment. XTH31 expression in yeast yields a protein with an in vitro XEH:XET activity ratio of >5000:1. xth31 accumulates significantly less Al in the root apex and cell wall, shows remarkably lower in vivo XET action and extractable XET activity, has a lower xyloglucan content, and exhibits slower elongation. An exogenous supply of xyloglucan significantly ameliorates Al toxicity by reducing Al accumulation in the roots, owing to the formation of an Al-xyloglucan complex in the medium, as verified by an obvious change in chemical shift of 27Al-NMR. Taken together, the data indicate that XTH31 affects Al sensitivity by modulating cell wall xyloglucan content and Al binding capacity. PMID:23204407

[Genetic instability of probiotic characteristics in the Bifidobacterium longum subsp. longum B379M strain during cultivation and maintenance].

PubMed

Averina, O V; Nezametdinova, V Z; Alekseeva, M G; Danilenko, V N

2012-11-01

The stability of inheriting several genes in the Russian commercial strain Bifidobacterium longum subsp. longum B379M during cultivation and maintenance under laboratory conditions has been studied. The examined genes code for probiotic characteristics, such as utilization of several sugars (lacA2 gene, encoding beta-galactosidase; ara gene, encoding arabinosidase; and galA gene, encoding arabinogalactan endo-beta-galactosidase); synthesis of bacteriocins (lans gene, encoding lanthionine synthetase); and mobile gene tet(W), conferring resistance to the antibiotic tetracycline. The other gene families studied include the genes responsible for signal transduction and adaptation to stress conditions in the majority of bacteria (serine/threonine protein kinases and the toxin-antitoxin systems of MazEF and RelBE types) and transcription regulators (genes encoding WhiB family proteins). Genomic DNA was analyzed by PCR using specially selected primers. A loss of the genes galA and tet(W) has been shown. It is proposed to expand the requirements on probiotic strains, namely, to control retention of the key probiotic genes using molecular biological methods.

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, Eric E.; Roessler, Paul G.

1999-01-01

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities.

Human Genomic Signatures of Brain Oscillations During Memory Encoding.

PubMed

Berto, Stefano; Wang, Guang-Zhong; Germi, James; Lega, Bradley C; Konopka, Genevieve

2018-05-01

Memory encoding is an essential step for all learning. However, the genetic and molecular mechanisms underlying human memory encoding remain poorly understood, and how this molecular framework permits the emergence of specific patterns of brain oscillations observed during mnemonic processing is unknown. Here, we directly compare intracranial electroencephalography recordings from the neocortex in individuals performing an episodic memory task with human gene expression from the same areas. We identify genes correlated with oscillatory memory effects across 6 frequency bands. These genes are enriched for autism-related genes and have preferential expression in neurons, in particular genes encoding synaptic proteins and ion channels, supporting the idea that the genes regulating voltage gradients are involved in the modulation of oscillatory patterns during successful memory encoding across brain areas. Memory-related genes are distinct from those correlated with other forms of cognitive processing and resting state fMRI. These data are the first to identify correlations between gene expression and active human brain states as well as provide a molecular window into memory encoding oscillations in the human brain.

Is hepatic lipid metabolism of beef cattle influenced by breed and dietary silage level?

PubMed Central

2014-01-01

Background In ruminants, unsaturated dietary fatty acids are biohydrogenated in the rumen and are further metabolised in various tissues, including liver, which has an important role in lipid and lipoprotein metabolism. Therefore, manipulation of muscle fatty acid composition should take into account liver metabolism. In the present study, the influence of breed and diet on liver lipid composition and gene expression was investigated in order to clarify the role of this organ in the lipid metabolism of ruminants. Forty purebred young bulls from two phylogenetically distant autochthonous cattle breeds, Alentejana and Barrosã, were assigned to two different diets (low vs. high silage) and slaughtered at 18 months of age. Liver fatty acid composition, mRNA levels of enzymes and transcription factors involved in lipid metabolism, as well as the plasma lipid profile, were assessed. Results In spite of similar plasma non-esterified fatty acids levels, liver triacylglycerols content was higher in Barrosã than in Alentejana bulls. Moreover, the fatty acid composition of liver was clearly distinct from the remaining tissues involved in fatty acid metabolism of ruminants, as shown by Principal Components Analysis. The hepatic tissue is particularly rich in α-linolenic acid and their products of desaturation and elongation. Results indicate that DGAT1, ELOVL2, FADS1 and FADS2 genes influence the fatty acid composition of the liver the most. Moreover, genes such as DGAT1 and ELOVL2 appear to be more sensitive to genetic background than to dietary manipulation, whereas genes encoding for desaturases, such as FADS1, appear to be modulated by dietary silage level. Conclusions Our results indicate that liver plays an important role in the biosynthesis of n-3 LC-PUFA. It is also suggested that dietary silage level influences the hepatic fatty acid metabolism in a breed-dependent manner, through changes in the expression of genes encoding for enzymes associated with the desaturation and elongation pathway. The importance of devising custom-made feeding strategies taking into account the genetic background is, therefore, stressed by the results from this experiment. PMID:24621212

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, E.E.; Roessler, P.G.

1999-07-27

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities. 8 figs.

Novel NCC mutants and functional analysis in a new cohort of patients with Gitelman syndrome.

PubMed

Glaudemans, Bob; Yntema, Helger G; San-Cristobal, Pedro; Schoots, Jeroen; Pfundt, Rolph; Kamsteeg, Erik-J; Bindels, René J; Knoers, Nine V A M; Hoenderop, Joost G; Hoefsloot, Lies H

2012-03-01

Gitelman syndrome (GS) is an autosomal recessive disorder characterized by hypokalemic metabolic alkalosis in conjunction with significant hypomagnesemia and hypocalciuria. The GS phenotype is caused by mutations in the solute carrier family 12, member 3 (SLC12A3) gene that encodes the thiazide-sensitive NaCl cotransporter (NCC). We analyzed DNA samples of 163 patients with a clinical suspicion of GS by direct sequencing of all 26 exons of the SLC12A3 gene. In total, 114 different mutations were identified, 31 of which have not been reported before. These novel variants include 3 deletions, 18 missense, 6 splice site and 4 nonsense mutations. We selected seven missense mutations to investigate their effect on NCC activity and plasma membrane localization by using the Xenopus laevis oocyte expression system. The Thr392Ile mutant did not display transport activity (probably class 2 mutation), while the Asn442Ser and Gln1030Arg NCC mutants showed decreased plasma membrane localization and consequently function, likely due to impaired trafficking (class 3 mutation). Even though the NaCl uptake was hampered for NCC mutants Glu121Asp, Pro751Leu, Ser475Cys and Tyr489His, the transporters reached the plasma membrane (class 4 mutation), suggesting an effect on NCC regulation or ion affinity. The present study shows the identification of 38 novel mutations in the SLC12A3 gene and provides insight into the mechanisms that regulate NCC.

Evolutionary analysis of hydrophobin gene family in two wood-degrading basidiomycetes, Phlebia brevispora and Heterobasidion annosum s.l.

PubMed Central

2013-01-01

Background Hydrophobins are small secreted cysteine-rich proteins that play diverse roles during different phases of fungal life cycle. In basidiomycetes, hydrophobin-encoding genes often form large multigene families with up to 40 members. The evolutionary forces driving hydrophobin gene expansion and diversification in basidiomycetes are poorly understood. The functional roles of individual genes within such gene families also remain unclear. The relationship between the hydrophobin gene number, the genome size and the lifestyle of respective fungal species has not yet been thoroughly investigated. Here, we present results of our survey of hydrophobin gene families in two species of wood-degrading basidiomycetes, Phlebia brevispora and Heterobasidion annosum s.l. We have also investigated the regulatory pattern of hydrophobin-encoding genes from H. annosum s.s. during saprotrophic growth on pine wood as well as on culture filtrate from Phlebiopsis gigantea using micro-arrays. These data are supplemented by results of the protein structure modeling for a representative set of hydrophobins. Results We have identified hydrophobin genes from the genomes of two wood-degrading species of basidiomycetes, Heterobasidion irregulare, representing one of the microspecies within the aggregate H. annosum s.l., and Phlebia brevispora. Although a high number of hydrophobin-encoding genes were observed in H. irregulare (16 copies), a remarkable expansion of these genes was recorded in P. brevispora (26 copies). A significant expansion of hydrophobin-encoding genes in other analyzed basidiomycetes was also documented (1–40 copies), whereas contraction through gene loss was observed among the analyzed ascomycetes (1–11 copies). Our phylogenetic analysis confirmed the important role of gene duplication events in the evolution of hydrophobins in basidiomycetes. Increased number of hydrophobin-encoding genes appears to have been linked to the species’ ecological strategy, with the non-pathogenic fungi having increased numbers of hydrophobins compared with their pathogenic counterparts. However, there was no significant relationship between the number of hydrophobin-encoding genes and genome size. Furthermore, our results revealed significant differences in the expression levels of the 16 H. annosum s.s. hydrophobin-encoding genes which suggest possible differences in their regulatory patterns. Conclusions A considerable expansion of the hydrophobin-encoding genes in basidiomycetes has been observed. The distribution and number of hydrophobin-encoding genes in the analyzed species may be connected to their ecological preferences. Results of our analysis also have shown that H. annosum s.l. hydrophobin-encoding genes may be under positive selection. Our gene expression analysis revealed differential expression of H. annosum s.s. hydrophobin genes under different growth conditions, indicating their possible functional diversification. PMID:24188142

Glutathione S-transferase-encoding gene as a potential probe for environmental bacterial isolates capable of degrading polycyclic aromatic hydrocarbons.

PubMed Central

Lloyd-Jones, G; Lau, P C

1997-01-01

Homologs of the glutathione S-transferase (GST)-encoding gene were identified in a collection of aromatic hydrocarbon-degrading Sphingomonas spp. isolated from New Zealand, Antarctica, and the United States by using PCR primers designed from the GST-encoding gene of Sphingomonas paucimobilis EPA505. Sequence analysis of PCR fragments generated from these isolates and of the GST gene amplified from DNA extracted from polycyclic aromatic hydrocarbon (PAH)-contaminated soil revealed a high degree of conservation, which may make the GST-encoding gene a potentially useful marker for PAH-degrading bacteria. PMID:9251217

Enterotoxin-encoding genes in Staphylococcus spp. from bulk goat milk.

PubMed

Lyra, Daniele G; Sousa, Francisca G C; Borges, Maria F; Givisiez, Patrícia E N; Queiroga, Rita C R E; Souza, Evandro L; Gebreyes, Wondwossen A; Oliveira, Celso J B

2013-02-01

Although Staphylococcus aureus has been implicated as the main Staphylococcus species causing human food poisoning, recent studies have shown that coagulase-negative Staphylococcus could also harbor enterotoxin-encoding genes. Such organisms are often present in goat milk and are the most important mastitis-causing agents. Therefore, this study aimed to investigate the occurrence of enterotoxin-encoding genes among coagulase-positive (CoPS) and coagulase-negative (CoNS) staphylococci isolated from raw goat milk produced in the semi-arid region of Paraiba, the most important region for goat milk production in Brazil. Enterotoxin-encoding genes were screened in 74 staphylococci isolates (30 CoPS and 44 CoNS) by polymerase chain reaction targeting the genes sea, seb, sec, sed, see, seg, seh, and sei. Enterotoxin-encoding genes were found in nine (12.2%) isolates, and four different genes (sea, sec, seg, and sei) were identified amongst the isolates. The most frequent genes were seg and sei, which were often found simultaneously in 44.5% of the isolates. The gene sec was the most frequent among the classical genes, and sea was found only in one isolate. All CoPS isolates (n=7) harboring enterotoxigenic genes were identified as S. aureus. The two coagulase-negative isolates were S. haemolyticus and S. hominis subsp. hominis and they harbored sei and sec genes, respectively. A higher frequency of enterotoxin-encoding genes was observed amongst CoPS (23.3%) than CoNS (4.5%) isolates (p<0.05), reinforcing the importance of S. aureus as a potential foodborne agent. However, the potential risk posed by CoNS in goat milk should not be ignored because it has a higher occurrence in goat milk and enterotoxin-encoding genes were detected in some isolates.

Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease.

PubMed

Zanoni, Paolo; Khetarpal, Sumeet A; Larach, Daniel B; Hancock-Cerutti, William F; Millar, John S; Cuchel, Marina; DerOhannessian, Stephanie; Kontush, Anatol; Surendran, Praveen; Saleheen, Danish; Trompet, Stella; Jukema, J Wouter; De Craen, Anton; Deloukas, Panos; Sattar, Naveed; Ford, Ian; Packard, Chris; Majumder, Abdullah al Shafi; Alam, Dewan S; Di Angelantonio, Emanuele; Abecasis, Goncalo; Chowdhury, Rajiv; Erdmann, Jeanette; Nordestgaard, Børge G; Nielsen, Sune F; Tybjærg-Hansen, Anne; Schmidt, Ruth Frikke; Kuulasmaa, Kari; Liu, Dajiang J; Perola, Markus; Blankenberg, Stefan; Salomaa, Veikko; Männistö, Satu; Amouyel, Philippe; Arveiler, Dominique; Ferrieres, Jean; Müller-Nurasyid, Martina; Ferrario, Marco; Kee, Frank; Willer, Cristen J; Samani, Nilesh; Schunkert, Heribert; Butterworth, Adam S; Howson, Joanna M M; Peloso, Gina M; Stitziel, Nathan O; Danesh, John; Kathiresan, Sekar; Rader, Daniel J

2016-03-11

Scavenger receptor BI (SR-BI) is the major receptor for high-density lipoprotein (HDL) cholesterol (HDL-C). In humans, high amounts of HDL-C in plasma are associated with a lower risk of coronary heart disease (CHD). Mice that have depleted Scarb1 (SR-BI knockout mice) have markedly elevated HDL-C levels but, paradoxically, increased atherosclerosis. The impact of SR-BI on HDL metabolism and CHD risk in humans remains unclear. Through targeted sequencing of coding regions of lipid-modifying genes in 328 individuals with extremely high plasma HDL-C levels, we identified a homozygote for a loss-of-function variant, in which leucine replaces proline 376 (P376L), in SCARB1, the gene encoding SR-BI. The P376L variant impairs posttranslational processing of SR-BI and abrogates selective HDL cholesterol uptake in transfected cells, in hepatocyte-like cells derived from induced pluripotent stem cells from the homozygous subject, and in mice. Large population-based studies revealed that subjects who are heterozygous carriers of the P376L variant have significantly increased levels of plasma HDL-C. P376L carriers have a profound HDL-related phenotype and an increased risk of CHD (odds ratio = 1.79, which is statistically significant). Copyright © 2016, American Association for the Advancement of Science.

The pmr gene, encoding a Ca2+-ATPase, is required for calcium and manganese homeostasis and normal development of hyphae and conidia in Neurospora crassa.

PubMed

Bowman, Barry J; Abreu, Stephen; Johl, Jessica K; Bowman, Emma Jean

2012-11-01

The pmr gene is predicted to encode a Ca(2+)-ATPase in the secretory pathway. We examined two strains of Neurospora crassa that lacked PMR: the Δpmr strain, in which pmr was completely deleted, and pmr(RIP), in which the gene was extensively mutated. Both strains had identical, complex phenotypes. Compared to the wild type, these strains required high concentrations of calcium or manganese for optimal growth and had highly branched, slow-growing hyphae. They conidiated poorly, and the shape and size of the conidia were abnormal. Calcium accumulated in the Δpmr strains to only 20% of the wild-type level. High concentrations of MnCl(2) (1 to 5 mM) in growth medium partially suppressed the morphological defects but did not alter the defect in calcium accumulation. The Δpmr Δnca-2 double mutant (nca-2 encodes a Ca(2+)-ATPase in the plasma membrane) accumulated 8-fold more calcium than the wild type, and the morphology of the hyphae was more similar to that of wild-type hyphae. Previous experiments failed to show a function for nca-1, which encodes a SERCA-type Ca(2+)-ATPase in the endoplasmic reticulum (B. J. Bowman, S. Abreu, E. Margolles-Clark, M. Draskovic, and E. J. Bowman, Eukaryot. Cell 10:654-661, 2011). The pmr(RIP) Δnca-1 double mutant accumulated small amounts of calcium, like the Δpmr strain, but exhibited even more extreme morphological defects. Thus, PMR can apparently replace NCA-1 in the endoplasmic reticulum, but NCA-1 cannot replace PMR. The morphological defects in the Δpmr strain are likely caused, in part, by insufficient concentrations of calcium and manganese in the Golgi compartment; however, PMR is also needed to accumulate normal levels of calcium in the whole cell.

Fatty acid-inducible ANGPTL4 governs lipid metabolic response to exercise

PubMed Central

Catoire, Milène; Alex, Sheril; Paraskevopulos, Nicolas; Mattijssen, Frits; Evers-van Gogh, Inkie; Schaart, Gert; Jeppesen, Jacob; Kneppers, Anita; Mensink, Marco; Voshol, Peter J.; Olivecrona, Gunilla; Tan, Nguan Soon; Hesselink, Matthijs K. C.; Berbée, Jimmy F.; Rensen, Patrick C. N.; Kalkhoven, Eric; Schrauwen, Patrick; Kersten, Sander

2014-01-01

Physical activity increases energy metabolism in exercising muscle. Whether acute exercise elicits metabolic changes in nonexercising muscles remains unclear. We show that one of the few genes that is more highly induced in nonexercising muscle than in exercising human muscle during acute exercise encodes angiopoietin-like 4 (ANGPTL4), an inhibitor of lipoprotein lipase-mediated plasma triglyceride clearance. Using a combination of human, animal, and in vitro data, we show that induction of ANGPTL4 in nonexercising muscle is mediated by elevated plasma free fatty acids via peroxisome proliferator-activated receptor-δ, presumably leading to reduced local uptake of plasma triglyceride-derived fatty acids and their sparing for use by exercising muscle. In contrast, the induction of ANGPTL4 in exercising muscle likely is counteracted via AMP-activated protein kinase (AMPK)-mediated down-regulation, promoting the use of plasma triglycerides as fuel for active muscles. Our data suggest that nonexercising muscle and the local regulation of ANGPTL4 via AMPK and free fatty acids have key roles in governing lipid homeostasis during exercise. PMID:24591600

Similar clinical and neuroimaging features in monozygotic twin pair with mutation in progranulin

PubMed Central

McDade, E.; Burrus, T.M.; Boot, B.P.; Kantarci, K.; Fields, J.; Lowe, V.J.; Peller, P.; Knopman, D.; Baker, M.; Finch, N.; Rademakers, R.; Petersen, R.

2012-01-01

Objective: To report the phenotypic characterization of monozygotic twins with mutations encoding progranulin (PGRN). Methods: We studied a twin pair with an exon 4 gene deletion in the PGRN gene. Both twins had clinical and neuropsychological examinations as well as structural MRI and fluorodeoxyglucose PET (FDG-PET) scans. PGRN gene sequencing was performed followed by progranulin ELISA in plasma. Results: Both twins manifested symptoms within 3 years of each other, with early behavioral, language, dysexecutive, and memory problems. MRI and FDG-PET imaging demonstrated a strikingly similar topography of findings with clear left hemisphere predominance. Serum progranulin levels in both were well below those from a normal population sample. Conclusions: Compared with the heterogeneity seen in many families with PGRN mutations, these monozygotic twins demonstrated strong clinical, neuroimaging, and serum progranulin level similarities, demonstrating the importance of shared genetic profiles beyond environmental influences in the symptomatic expression of the disease. PMID:22491866

Genetic forms of nephrogenic diabetes insipidus (NDI): Vasopressin receptor defect (X-linked) and aquaporin defect (autosomal recessive and dominant).

PubMed

Bichet, Daniel G; Bockenhauer, Detlef

2016-03-01

Nephrogenic diabetes insipidus (NDI), which can be inherited or acquired, is characterized by an inability to concentrate urine despite normal or elevated plasma concentrations of the antidiuretic hormone, arginine vasopressin (AVP). Polyuria with hyposthenuria and polydipsia are the cardinal clinical manifestations of the disease. About 90% of patients with congenital NDI are males with X-linked NDI who have mutations in the vasopressin V2 receptor (AVPR2) gene encoding the vasopressin V2 receptor. In less than 10% of the families studied, congenital NDI has an autosomal recessive or autosomal dominant mode of inheritance with mutations in the aquaporin-2 (AQP2) gene. When studied in vitro, most AVPR2 and AQP2 mutations lead to proteins trapped in the endoplasmic reticulum and are unable to reach the plasma membrane. Prior knowledge of AVPR2 or AQP2 mutations in NDI families and perinatal mutation testing is of direct clinical value and can avert the physical and mental retardation associated with repeated episodes of dehydration. Copyright © 2016 Elsevier Ltd. All rights reserved.

Molecular cloning of the plasma membrane H(+)-ATPase from Kluyveromyces lactis: a single nucleotide substitution in the gene confers ethidium bromide resistance and deficiency in K+ uptake.

PubMed Central

Miranda, M; Ramírez, J; Peña, A; Coria, R

1995-01-01

A Kluyveromyces lactis strain resistant to ethidium bromide and deficient in potassium uptake was isolated. Studies on the proton-pumping activity of the mutant strain showed that a decreased H(+)-ATPase specific activity was responsible for the observed phenotypes. The putative K. lactis PMA1 gene encoding the plasma membrane H(+)-ATPase was cloned by its ability to relieve the potassium transport defect of this mutant and by reversing its resistance to ethidium bromide. Its deduced amino acid sequence predicts a protein 899 residues long that is structurally colinear in its full length to H(+)-ATPases cloned from different yeasts, except for the presence of a variable N-terminal domain. By PCR-mediated amplification, we identified a transition from G to A that rendered the substitution of the fully conserved methionine at position 699 by isoleucine. We attribute to this amino acid change the low capacity of the mutant H(+)-ATPase to pump out protons. PMID:7730265

Expression of three gonadotropin subunits and gonadotropin receptor mRNA during male-to-female sex change in the cinnamon clownfish, Amphiprion melanopus.

PubMed

An, Kwang Wook; Lee, Jehee; Choi, Cheol Young

2010-08-01

To quantify the sex-change progression from male to female in the cinnamon clownfish, Amphiprion melanopus, we divided gonadal development into three stages (I, mature male; II, male at 90 days after removal of the female; and III, mature female), and the expression of GTH subunits and GTH receptors during each of these stages was investigated. The mRNA of the three GTH subunits and their receptors increased with progression from male to female. To understand the effect of gonadotropin-releasing hormone (GnRH) on this progression, we examined expression of genes encoding the GTH subunit mRNA in the pituitary and the GTH-receptor mRNA in the gonads in addition to investigating changes in plasma E(2) levels after GnRH analogue (GnRHa) injection. GnRHa treatment increased mRNA expression levels of these genes, as well as plasma E(2) levels, indicating that GnRH plays an important regulatory role in the brain-pituitary-gonad axis of immature cinnamon clownfish. Copyright (c) 2010 Elsevier Inc. All rights reserved.

Laboratory and Genetic Investigation of Mutations Accounting for Congenital Fibrinogen Disorders.

PubMed

Neerman-Arbez, Marguerite; de Moerloose, Philippe; Casini, Alessandro

2016-06-01

Congenital fibrinogen disorders are classified into two types of plasma fibrinogen defects: type I (quantitative fibrinogen deficiencies), that is, hypofibrinogenemia or afibrinogenemia, in which there are low or absent plasma fibrinogen antigen levels, respectively, and type II (qualitative fibrinogen deficiencies), that is, dysfibrinogenemia or hypodysfibrinogenemia, in which there are normal or reduced antigen levels associated with disproportionately low functional activity. These disorders are caused by mutations in the three fibrinogen-encoding genes FGA, FGB, and FGG. Afibrinogenemia is associated with mild to severe bleeding, whereas hypofibrinogenemia is often asymptomatic. For these quantitative disorders, the majority of mutations prevent protein production. However, in some cases, missense or late-truncating nonsense mutations allow synthesis of the mutant fibrinogen chain, but intracellular fibrinogen assembly and/or secretion are impaired. Qualitative fibrinogen disorders are associated with bleeding, thrombosis, or both thrombosis and bleeding, but many dysfibrinogenemias are asymptomatic. The majority of cases are caused by heterozygous missense mutations. Here, we review the laboratory and genetic diagnosis of fibrinogen gene anomalies with an updated discussion of causative mutations identified. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Three copies of a single protein II-encoding sequence in the genome of Neisseria gonorrhoeae JS3: evidence for gene conversion and gene duplication.

PubMed

van der Ley, P

1988-11-01

Gonococci express a family of related outer membrane proteins designated protein II (P.II). These surface proteins are subject to both phase variation and antigenic variation. The P.II gene repertoire of Neisseria gonorrhoeae strain JS3 was found to consist of at least ten genes, eight of which were cloned. Sequence analysis and DNA hybridization studies revealed that one particular P.II-encoding sequence is present in three distinct, but almost identical, copies in the JS3 genome. These genes encode the P.II protein that was previously identified as P.IIc. Comparison of their sequences shows that the multiple copies of this P.IIc-encoding gene might have been generated by both gene conversion and gene duplication.

Reporter gene expression in dendritic cells after gene gun administration of plasmid DNA.

PubMed

Watkins, Craig; Hopkins, John; Harkiss, Gordon

2005-07-21

Dendritic cells (DC) play an integral role in plasmid DNA vaccination. However, the interaction between plasmid DNA and DC in vivo is incompletely understood. In this report, we utilise the sheep pseudoafferent cannulation model to examine the interaction between plasmid DNA encoding enhanced green fluorescent protein (pEGFP) and afferent lymph DC (ALDC) following gene gun administration. The results show that peaks of fluorescent ALDC tended to appear around days 1-4 and 9-13, then erratically thereafter for up to 2 months. Phenotypic analysis showed that EGFP+ ALDC expressed MHC class II, WC6, CD1b, and SIRPalpha markers. Plasmid, detected by PCR, was found in lymph cells and cell-free plasma on a daily basis, and was present variably for up to 2 months. Plasmid was also detected in purified CD1b+ ALDC, but the presence of plasmid did not correlate with EGFP expression by ALDC. Free EGFP in afferent lymph plasma was detectable by luminometry only after three administrations of the plasmid. The results show that gene gun administered pEGFP persisted for extended periods after a single administration, leeching out of skin on a daily basis. The plasmid was associated with both the cellular and fluid components of afferent lymph. EGFP protein appeared in afferent lymph in a pulsatile manner, but associated only with ALDC.

Uridine Metabolism in HIV-1-Infected Patients: Effect of Infection, of Antiretroviral Therapy and of HIV-1/ART-Associated Lipodystrophy Syndrome

PubMed Central

Domingo, Pere; Torres-Torronteras, Javier; Pomar, Virginia; Giralt, Marta; Domingo, Joan Carles; Gutierrez, Maria del Mar; Gallego-Escuredo, José M.; Mateo, Maria Gracia; Cano-Soldado, Pedro; Fernandez, Irene; Pastor-Anglada, Marçal; Vidal, Francesc; Villarroya, Francesc; Andreu, Antoni; Marti, Ramon

2010-01-01

Background Uridine has been advocated for the treatment of HIV-1/HAART-associated lipodystrophy (HALS), although its metabolism in HIV-1-infected patients is poorly understood. Methods Plasma uridine concentrations were measured in 35 controls and 221 HIV-1-infected patients and fat uridine in 15 controls and 19 patients. The diagnosis of HALS was performed following the criteria of the Lipodystrophy Severity Grading Scale. Uridine was measured by a binary gradient-elution HPLC method. Analysis of genes encoding uridine metabolizing enzymes in fat was performed with TaqMan RT-PCR. Results Median plasma uridine concentrations for HIV-1-infected patients were 3.80 µmol/l (interquartile range: 1.60), and for controls 4.60 µmol/l (IQR: 1.8) (P = 0.0009). In fat, they were of 6.0 (3.67), and 2.8 (4.65) nmol/mg of protein, respectively (P = 0.0118). Patients with a mixed HALS form had a median plasma uridine level of 4.0 (IC95%: 3.40–4.80) whereas in those with isolated lipoatrophy it was 3.25 (2.55–4.15) µmol/l/l (P = 0.0066). The expression of uridine cytidine kinase and uridine phosphorylase genes was significantly decreased in all groups of patients with respect to controls. A higher expression of the mRNAs for concentrative nucleoside transporters was found in HIV-1-infected patients with respect to healthy controls. Conclusions HIV-1 infection is associated with a decrease in plasma uridine and a shift of uridine to the adipose tissue compartment. Antiretroviral therapy was not associated with plasma uridine concentrations, but pure lipoatrophic HALS was associated with significantly lower plasma uridine concentrations. PMID:21085568

Rab11-dependent Recycling of the Human Ether-a-go-go-related Gene (hERG) Channel*

PubMed Central

Chen, Jeffery; Guo, Jun; Yang, Tonghua; Li, Wentao; Lamothe, Shawn M.; Kang, Yudi; Szendrey, John A.; Zhang, Shetuan

2015-01-01

The human ether-a-go-go-related gene (hERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel (IKr). A reduction in the hERG current causes long QT syndrome, which predisposes affected individuals to ventricular arrhythmias and sudden death. We reported previously that hERG channels in the plasma membrane undergo vigorous internalization under low K+ conditions. In the present study, we addressed whether hERG internalization occurs under normal K+ conditions and whether/how internalized channels are recycled back to the plasma membrane. Using patch clamp, Western blot, and confocal imaging analyses, we demonstrated that internalized hERG channels can effectively recycle back to the plasma membrane. Low K+-enhanced hERG internalization is accompanied by an increased rate of hERG recovery in the plasma membrane upon reculture following proteinase K-mediated clearance of cell-surface proteins. The increased recovery rate is not due to enhanced protein synthesis, as hERG mRNA expression was not altered by low K+ exposure, and the increased recovery was observed in the presence of the protein biosynthesis inhibitor cycloheximide. GTPase Rab11, but not Rab4, is involved in the recycling of hERG channels. Interfering with Rab11 function not only delayed hERG recovery in cells after exposure to low K+ medium but also decreased hERG expression and function in cells under normal culture conditions. We concluded that the recycling pathway plays an important role in the homeostasis of plasma membrane-bound hERG channels. PMID:26152716

Cadmium induces cadmium-tolerant gene expression in the filamentous fungus Trichoderma harzianum.

PubMed

Cacciola, Santa O; Puglisi, Ivana; Faedda, Roberto; Sanzaro, Vincenzo; Pane, Antonella; Lo Piero, Angela R; Evoli, Maria; Petrone, Goffredo

2015-11-01

The filamentous fungus Trichoderma harzianum, strain IMI 393899, was able to grow in the presence of the heavy metals cadmium and mercury. The main objective of this research was to study the molecular mechanisms underlying the tolerance of the fungus T. harzianum to cadmium. The suppression subtractive hybridization (SSH) method was used for the characterization of the genes of T. harzianum implicated in cadmium tolerance compared with those expressed in the response to the stress induced by mercury. Finally, the effects of cadmium exposure were also validated by measuring the expression levels of the putative genes coding for a glucose transporter, a plasma membrane ATPase, a Cd(2+)/Zn(2+) transporter protein and a two-component system sensor histidine kinase YcbA, by real-time-PCR. By using the aforementioned SSH strategy, it was possible to identify 108 differentially expressed genes of the strain IMI 393899 of T. harzianum grown in a mineral substrate with the addition of cadmium. The expressed sequence tags identified by SSH technique were encoding different genes that may be involved in different biological processes, including those associated to primary and secondary metabolism, intracellular transport, transcription factors, cell defence, signal transduction, DNA metabolism, cell growth and protein synthesis. Finally, the results show that in the mechanism of tolerance to cadmium a possible signal transduction pathway could activate a Cd(2+)/Zn(2+) transporter protein and/or a plasma membrane ATPase that could be involved in the compartmentalization of cadmium inside the cell.

Regulation of cation transport in Saccharomyces cerevisiae by the salt tolerance gene HAL3.

PubMed Central

Ferrando, A; Kron, S J; Rios, G; Fink, G R; Serrano, R

1995-01-01

Dynamic regulation of ion transport is essential for homeostasis as cells confront changes in their environment. The gene HAL3 encodes a novel component of this regulatory circuit in the yeast Saccharomyces cerevisiae. Overexpression of HAL3 improves growth of wild-type cells exposed to toxic concentrations of sodium and lithium and suppresses the salt sensitivity conferred by mutation of the calcium-dependent protein phosphatase calcineurin. Null mutants of HAL3 display salt sensitivity. The sequence of HAL3 gives little clue to its function. However, alterations in intracellular cation concentrations associated with changes in HAL3 expression suggest that HAL3 activity may directly increase cytoplasmic K+ and decrease Na+ and Li+. Cation efflux in S. cerevisiae is mediated by the P-type ATPase encoded by the ENA1/PMR24 gene, a putative plasma membrane Na+ pump whose expression is salt induced. Acting in concert with calcineurin, HAL3 is necessary for full activation of ENA1 expression. This functional complementarity is also reflected in the participation of both proteins in recovery from alpha-factor-induced growth arrest. Recently, HAL3 was isolated as a gene (named SIS2) which when overexpressed partially relieves loss of transcription of G1 cyclins in mutants lacking the protein phosphatase Sit4p. Therefore, HAL3 influences cell cycle control and ion homeostasis, acting in parallel to the protein phosphatases Sit4p and calcineurin. PMID:7565698

[Expression changes of major outer membrane protein antigens in Leptospira interrogans during infection and its mechanism].

PubMed

Zheng, Linli; Ge, Yumei; Hu, Weilin; Yan, Jie

2013-03-01

To determine expression changes of major outer membrane protein(OMP) antigens of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai strain Lai during infection of human macrophages and its mechanism. OmpR encoding genes and OmpR-related histidine kinase (HK) encoding gene of L.interrogans strain Lai and their functional domains were predicted using bioinformatics technique. mRNA level changes of the leptospiral major OMP-encoding genes before and after infection of human THP-1 macrophages were detected by real-time fluorescence quantitative RT-PCR. Effects of the OmpR-encoding genes and HK-encoding gene on the expression of leptospiral OMPs during infection were determined by HK-peptide antiserum block assay and closantel inhibitive assays. The bioinformatics analysis indicated that LB015 and LB333 were referred to OmpR-encoding genes of the spirochete, while LB014 might act as a OmpR-related HK-encoding gene. After the spirochete infecting THP-1 cells, mRNA levels of leptospiral lipL21, lipL32 and lipL41 genes were rapidly and persistently down-regulated (P <0.01), whereas mRNA levels of leptospiral groEL, mce, loa22 and ligB genes were rapidly but transiently up-regulated (P<0.01). The treatment with closantel and HK-peptide antiserum partly reversed the infection-based down-regulated mRNA levels of lipL21 and lipL48 genes (P <0.01). Moreover, closantel caused a decrease of the infection-based up-regulated mRNA levels of groEL, mce, loa22 and ligB genes (P <0.01). Expression levels of L.interrogans strain Lai major OMP antigens present notable changes during infection of human macrophages. There is a group of OmpR-and HK-encoding genes which may play a major role in down-regulation of expression levels of partial OMP antigens during infection.

Rgs13 constrains early B cell responses and limits germinal center sizes.

PubMed

Hwang, Il-Young; Hwang, Kyung-Sun; Park, Chung; Harrison, Kathleen A; Kehrl, John H

2013-01-01

Germinal centers (GCs) are microanatomic structures that develop in secondary lymphoid organs in response to antigenic stimulation. Within GCs B cells clonally expand and their immunoglobulin genes undergo class switch recombination and somatic hypermutation. Transcriptional profiling has identified a number of genes that are prominently expressed in GC B cells. Among them is Rgs13, which encodes an RGS protein with a dual function. Its canonical function is to accelerate the intrinsic GTPase activity of heterotrimeric G-protein α subunits at the plasma membrane, thereby limiting heterotrimeric G-protein signaling. A unique, non-canonical function of RGS13 occurs following translocation to the nucleus, where it represses CREB transcriptional activity. The functional role of RGS13 in GC B cells is unknown. To create a surrogate marker for Rgs13 expression and a loss of function mutation, we inserted a GFP coding region into the Rgs13 genomic locus. Following immunization GFP expression rapidly increased in activated B cells, persisted in GC B cells, but declined in newly generated memory B and plasma cells. Intravital microscopy of the inguinal lymph node (LN) of immunized mice revealed the rapid appearance of GFP(+) cells at LN interfollicular regions and along the T/B cell borders, and eventually within GCs. Analysis of WT, knock-in, and mixed chimeric mice indicated that RGS13 constrains extra-follicular plasma cell generation, GC size, and GC B cell numbers. Analysis of select cell cycle and GC specific genes disclosed an aberrant gene expression profile in the Rgs13 deficient GC B cells. These results indicate that RGS13, likely acting at cell membranes and in nuclei, helps coordinate key decision points during the expansion and differentiation of naive B cells.

Transcriptomic analysis of Arabidopsis developing stems: a close-up on cell wall genes

PubMed Central

Minic, Zoran; Jamet, Elisabeth; San-Clemente, Hélène; Pelletier, Sandra; Renou, Jean-Pierre; Rihouey, Christophe; Okinyo, Denis PO; Proux, Caroline; Lerouge, Patrice; Jouanin, Lise

2009-01-01

Background Different strategies (genetics, biochemistry, and proteomics) can be used to study proteins involved in cell biogenesis. The availability of the complete sequences of several plant genomes allowed the development of transcriptomic studies. Although the expression patterns of some Arabidopsis thaliana genes involved in cell wall biogenesis were identified at different physiological stages, detailed microarray analysis of plant cell wall genes has not been performed on any plant tissues. Using transcriptomic and bioinformatic tools, we studied the regulation of cell wall genes in Arabidopsis stems, i.e. genes encoding proteins involved in cell wall biogenesis and genes encoding secreted proteins. Results Transcriptomic analyses of stems were performed at three different developmental stages, i.e., young stems, intermediate stage, and mature stems. Many genes involved in the synthesis of cell wall components such as polysaccharides and monolignols were identified. A total of 345 genes encoding predicted secreted proteins with moderate or high level of transcripts were analyzed in details. The encoded proteins were distributed into 8 classes, based on the presence of predicted functional domains. Proteins acting on carbohydrates and proteins of unknown function constituted the two most abundant classes. Other proteins were proteases, oxido-reductases, proteins with interacting domains, proteins involved in signalling, and structural proteins. Particularly high levels of expression were established for genes encoding pectin methylesterases, germin-like proteins, arabinogalactan proteins, fasciclin-like arabinogalactan proteins, and structural proteins. Finally, the results of this transcriptomic analyses were compared with those obtained through a cell wall proteomic analysis from the same material. Only a small proportion of genes identified by previous proteomic analyses were identified by transcriptomics. Conversely, only a few proteins encoded by genes having moderate or high level of transcripts were identified by proteomics. Conclusion Analysis of the genes predicted to encode cell wall proteins revealed that about 345 genes had moderate or high levels of transcripts. Among them, we identified many new genes possibly involved in cell wall biogenesis. The discrepancies observed between results of this transcriptomic study and a previous proteomic study on the same material revealed post-transcriptional mechanisms of regulation of expression of genes encoding cell wall proteins. PMID:19149885

Defective mitochondrial rRNA methyltransferase MRM2 causes MELAS-like clinical syndrome

PubMed Central

Garone, Caterina; D’Souza, Aaron R; Dallabona, Cristina; Lodi, Tiziana; Rebelo-Guiomar, Pedro; Rorbach, Joanna; Donati, Maria Alice; Procopio, Elena; Montomoli, Martino; Guerrini, Renzo; Zeviani, Massimo; Calvo, Sarah E; Mootha, Vamsi K; DiMauro, Salvatore; Ferrero, Ileana; Minczuk, Michal

2017-01-01

Abstract Defects in nuclear-encoded proteins of the mitochondrial translation machinery cause early-onset and tissue-specific deficiency of one or more OXPHOS complexes. Here, we report a 7-year-old Italian boy with childhood-onset rapidly progressive encephalomyopathy and stroke-like episodes. Multiple OXPHOS defects and decreased mtDNA copy number (40%) were detected in muscle homogenate. Clinical features combined with low level of plasma citrulline were highly suggestive of mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome, however, the common m.3243 A > G mutation was excluded. Targeted exome sequencing of genes encoding the mitochondrial proteome identified a damaging mutation, c.567 G > A, affecting a highly conserved amino acid residue (p.Gly189Arg) of the MRM2 protein. MRM2 has never before been linked to a human disease and encodes an enzyme responsible for 2’-O-methyl modification at position U1369 in the human mitochondrial 16S rRNA. We generated a knockout yeast model for the orthologous gene that showed a defect in respiration and the reduction of the 2’-O-methyl modification at the equivalent position (U2791) in the yeast mitochondrial 21S rRNA. Complementation with the mrm2 allele carrying the equivalent yeast mutation failed to rescue the respiratory phenotype, which was instead completely rescued by expressing the wild-type allele. Our findings establish that defective MRM2 causes a MELAS-like phenotype, and suggests the genetic screening of the MRM2 gene in patients with a m.3243 A > G negative MELAS-like presentation. PMID:28973171

Defective mitochondrial rRNA methyltransferase MRM2 causes MELAS-like clinical syndrome.

PubMed

Garone, Caterina; D'Souza, Aaron R; Dallabona, Cristina; Lodi, Tiziana; Rebelo-Guiomar, Pedro; Rorbach, Joanna; Donati, Maria Alice; Procopio, Elena; Montomoli, Martino; Guerrini, Renzo; Zeviani, Massimo; Calvo, Sarah E; Mootha, Vamsi K; DiMauro, Salvatore; Ferrero, Ileana; Minczuk, Michal

2017-11-01

Defects in nuclear-encoded proteins of the mitochondrial translation machinery cause early-onset and tissue-specific deficiency of one or more OXPHOS complexes. Here, we report a 7-year-old Italian boy with childhood-onset rapidly progressive encephalomyopathy and stroke-like episodes. Multiple OXPHOS defects and decreased mtDNA copy number (40%) were detected in muscle homogenate. Clinical features combined with low level of plasma citrulline were highly suggestive of mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome, however, the common m.3243 A > G mutation was excluded. Targeted exome sequencing of genes encoding the mitochondrial proteome identified a damaging mutation, c.567 G > A, affecting a highly conserved amino acid residue (p.Gly189Arg) of the MRM2 protein. MRM2 has never before been linked to a human disease and encodes an enzyme responsible for 2'-O-methyl modification at position U1369 in the human mitochondrial 16S rRNA. We generated a knockout yeast model for the orthologous gene that showed a defect in respiration and the reduction of the 2'-O-methyl modification at the equivalent position (U2791) in the yeast mitochondrial 21S rRNA. Complementation with the mrm2 allele carrying the equivalent yeast mutation failed to rescue the respiratory phenotype, which was instead completely rescued by expressing the wild-type allele. Our findings establish that defective MRM2 causes a MELAS-like phenotype, and suggests the genetic screening of the MRM2 gene in patients with a m.3243 A > G negative MELAS-like presentation. © The Author 2017. Published by Oxford University Press.

Accumulation of N-Acetylglucosamine Oligomers in the Plant Cell Wall Affects Plant Architecture in a Dose-Dependent and Conditional Manner1[W][OPEN

PubMed Central

Vanholme, Bartel; Vanholme, Ruben; Turumtay, Halbay; Goeminne, Geert; Cesarino, Igor; Goubet, Florence; Morreel, Kris; Rencoret, Jorge; Bulone, Vincent; Hooijmaijers, Cortwa; De Rycke, Riet; Gheysen, Godelieve; Ralph, John; De Block, Marc; Meulewaeter, Frank; Boerjan, Wout

2014-01-01

To study the effect of short N-acetylglucosamine (GlcNAc) oligosaccharides on the physiology of plants, N-ACETYLGLUCOSAMINYLTRANSFERASE (NodC) of Azorhizobium caulinodans was expressed in Arabidopsis (Arabidopsis thaliana). The corresponding enzyme catalyzes the polymerization of GlcNAc and, accordingly, β-1,4-GlcNAc oligomers accumulated in the plant. A phenotype characterized by difficulties in developing an inflorescence stem was visible when plants were grown for several weeks under short-day conditions before transfer to long-day conditions. In addition, a positive correlation between the oligomer concentration and the penetrance of the phenotype was demonstrated. Although NodC overexpression lines produced less cell wall compared with wild-type plants under nonpermissive conditions, no indications were found for changes in the amount of the major cell wall polymers. The effect on the cell wall was reflected at the transcriptome level. In addition to genes encoding cell wall-modifying enzymes, a whole set of genes encoding membrane-coupled receptor-like kinases were differentially expressed upon GlcNAc accumulation, many of which encoded proteins with an extracellular Domain of Unknown Function26. Although stress-related genes were also differentially expressed, the observed response differed from that of a classical chitin response. This is in line with the fact that the produced chitin oligomers were too small to activate the chitin receptor-mediated signal cascade. Based on our observations, we propose a model in which the oligosaccharides modify the architecture of the cell wall by acting as competitors in carbohydrate-carbohydrate or carbohydrate-protein interactions, thereby affecting noncovalent interactions in the cell wall or at the interface between the cell wall and the plasma membrane. PMID:24664205

The role of production of adipsin and leptin in the development of insulin resistance in patients with abdominal obesity.

PubMed

Vasilenko, M A; Kirienkova, E V; Skuratovskaia, D A; Zatolokin, P A; Mironyuk, N I; Litvinova, L S

2017-07-01

We investigated the tissue-specific features of the production of adipokines (leptin and adipsin) by adipose tissue in obese patients depending on the degree of obesity and the state of carbohydrate metabolism. An increase in the content of adipsin and leptin in the blood plasma was found. In patients with varying degrees of obesity with and without type 2 diabetes mellitus (DM 2), we determined the level of tissue-specific expression of LEP and CFD genes encoding leptin and adipsin, respectively. The contribution of different adipose tissue depots to the blood plasma level of adipsin and leptin in obese patients with and without DM 2 was established. The disturbance of reciprocal relationships between adipsin and leptin in obesity is associated with the development of insulin resistance.

Development and evaluation of a real-time polymerase chain reaction assay for the rapid detection of Talaromyces marneffei MP1 gene in human plasma.

PubMed

Hien, Ha Thuc Ai; Thanh, Tran Tan; Thu, Nguyen Thi Mai; Nguyen, Ashley; Thanh, Nguyen Tat; Lan, Nguyen Phu Huong; Simmons, Cameron; Shikuma, Cecilia; Chau, Nguyen Van Vinh; Thwaites, Guy; Le, Thuy

2016-12-01

Penicilliosis caused by Talaromyces marneffei is a common AIDS-defining illness in South and Southeast Asia. Diagnosis is based on culture which can take up to 14 days for identification, leading to treatment delay and increased mortality. We developed a TaqMan real-time PCR assay targeting the MP1 gene encoding an abundant cell wall protein specific to T. marneffei. The assay's performance was evaluated in MP1-containing plasmids, clinical isolates, and plasma from HIV-infected patients with and without penicilliosis. The assay consistently detected 10 copies of MP1-containing plasmids per reaction and 100 T. marneffei yeast cells per millilitre plasma. There were no amplification with seven other Penicillium species and six other HIV-associated fungal pathogens tested. The assay was evaluated in 70 patients with AIDS: 50 patients with culture-confirmed penicilliosis and 20 patients with opportunistic infections other than penicilliosis. The diagnostic sensitivity was 70.4% (19/27, 95% CI: 51.5-84.1%) and 52.2% (12/23, 95% CI: 33.0-70.8%) in plasma samples collected prior to and within 48 h of antifungal therapy respectively. The diagnostic specificity was 100% (20/20, 95% CI: 83.9-100%). This assay provides a useful tool for the rapid diagnosis of T. marneffei infection and has the potential to improve the management of patients with penicilliosis. © 2016 The Authors. Mycoses Published by Blackwell Verlag GmbH.

Magnetogenetics: Remote Control of Cellular Signaling with Magnetic Fields

NASA Astrophysics Data System (ADS)

Sauer, Jeremy P.

Means for temporally regulating gene expression and cellular activity are invaluable for elucidating the underlying physiological processes and have therapeutic implications. Here we report the development of a system for remote regulation of gene expression by low frequency radiowaves (RF) or by a static magnetic field. We accomplished this by first adding iron oxide nanoparticles - either exogenously or as genetically encoded ferritin/ferric oxyhydroxide particle. These particles have been designed with affinity to the plasma membrane ion channel Transient Receptor Potential Vanilloid 1 (TRPV1) by a conjugated antibody. Application of a magnetic field stimulates the particle to gate the ion channel and this, in turn, initiates calcium-dependent transgene expression. We first demonstrated in vitro that TRPV1 can be actuated to cause calcium flux into the cell by directly applying a localized magnetic field. In mice expressing these genetically encoded components, application of external magnetic field caused remote stimulation of insulin transgene expression and significantly lowered blood glucose. In addition, we are investigating mechanisms by which iron oxide nanoparticles can absorb RF, and transduce this energy to cause channel opening. This robust, repeatable method for remote cellular regulation in vivo may ultimately have applications in basic science, as well as in technology and therapeutics.

Molecular characterization of cDNAs encoding G protein alpha and beta subunits and study of their temporal and spatial expression patterns in Nicotiana plumbaginifolia Viv.

PubMed

Kaydamov, C; Tewes, A; Adler, K; Manteuffel, R

2000-04-25

We have isolated cDNA sequences encoding alpha and beta subunits of potential G proteins from a cDNA library prepared from somatic embryos of Nicotiana plumbaginifolia Viv. at early developmental stages. The predicted NPGPA1 and NPGPB1 gene products are 75-98% identical to the known respective plant alpha and beta subunits. Southern hybridizations indicate that NPGPA1 is probably a single-copy gene, whereas at least two copies of NPGPB1 exist in the N. plumbaginifolia genome. Northern analyses reveal that both NPGPA1 and NPGPB1 mRNA are expressed in all embryogenic stages and plant tissues examined and their expression is obviously regulated by the plant hormone auxin. Immunohistological localization of NPGPalpha1 and NPGPbeta1 preferentially on plasma and endoplasmic reticulum membranes and their immunochemical detection exclusively in microsomal cell fractions implicate membrane association of both proteins. The temporal and spatial expression patterns of NPGPA1 and NPGPB1 show conformity as well as differences. This could account for not only cooperative, but also individual activities of both subunits during embryogenesis and plant development.

Efficient lowering of triglyceride levels in mice by human apoAV protein variants associated with hypertriglyceridemia.

PubMed

Vaessen, Stefan F C; Sierts, Jeroen A; Kuivenhoven, Jan Albert; Schaap, Frank G

2009-02-06

Variation in the apolipoprotein A5 (APOA5) gene has consistently been associated with increased plasma triglyceride (TG) levels in epidemiological studies. In vivo functionality of these variations, however, has thus far not been tested. Using adenoviral over-expression, we evaluated plasma expression levels and TG-lowering efficacies of wild-type human apoAV, two human apoAV variants associated with increased TG (S19W, G185C) and one variant (Q341H) that is predicted to have altered protein function. Injection of mice with adenovirus encoding wild-type or mutant apoAV resulted in an identical dose-dependent elevation of human apoAV levels in plasma. The increase in apoAV levels resulted in pronounced lowering of plasma TG levels at two viral dosages. Unexpectedly, the TG-lowering efficacy of all three apoAV variants was similar to wild-type apoAV. In addition, no effect on TG-hydrolysis-related plasma parameters (free fatty acids, glycerol and post-heparin lipoprotein lipase activity) was apparent upon expression of all apoAV variants. In conclusion, our data indicate that despite their association with hypertriglyceridemia and/or predicted protein dysfunction, the 19W, 185C and 341H apoAV variants are equally effective in reducing plasma TG levels in mice.

Trichoderma genes

DOEpatents

Foreman, Pamela [Los Altos, CA; Goedegebuur, Frits [Vlaardingen, NL; Van Solingen, Pieter [Naaldwijk, NL; Ward, Michael [San Francisco, CA

2012-06-19

Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

The rice blast resistance gene Ptr encodes an atypical protein required for broad spectrum disease resistance

USDA-ARS?s Scientific Manuscript database

Plant resistance (R) genes typically encode proteins with nucleotide binding site-leucine rich repeat (NLR) domains. We identified a novel, broad-spectrum rice blast R gene, Ptr, encoding a non-NLR protein with four Armadillo repeats. Ptr was originally identified by fast neutron mutagenesis as a ...

Selenium Pretreatment Alleviated LPS-Induced Immunological Stress Via Upregulation of Several Selenoprotein Encoding Genes in Murine RAW264.7 Cells.

PubMed

Wang, Longqiong; Jing, Jinzhong; Yan, Hui; Tang, Jiayong; Jia, Gang; Liu, Guangmang; Chen, Xiaoling; Tian, Gang; Cai, Jingyi; Shang, Haiying; Zhao, Hua

2018-04-18

This study was conducted to profile selenoprotein encoding genes in mouse RAW264.7 cells upon lipopolysaccharide (LPS) challenge and integrate their roles into immunological regulation in response to selenium (Se) pretreatment. LPS was used to develop immunological stress in macrophages. Cells were pretreated with different levels of Se (0, 0.5, 1.0, 1.5, 2.0 μmol Se/L) for 2 h, followed by LPS (100 ng/mL) stimulation for another 3 h. The mRNA expression of 24 selenoprotein encoding genes and 9 inflammation-related genes were investigated. The results showed that LPS (100 ng/mL) effectively induced immunological stress in RAW264.7 cells with induced inflammation cytokines, IL-6 and TNF-α, mRNA expression, and cellular secretion. LPS increased (P < 0.05) mRNA profiles of 9 inflammation-related genes in cells, while short-time Se pretreatment modestly reversed (P < 0.05) the LPS-induced upregulation of 7 genes (COX-2, ICAM-1, IL-1β, IL-6, IL-10, iNOS, and MCP-1) and further increased (P < 0.05) expression of IFN-β and TNF-α in stressed cells. Meanwhile, LPS decreased (P < 0.05) mRNA levels of 18 selenoprotein encoding genes and upregulated mRNA levels of TXNRD1 and TXNRD3 in cells. Se pretreatment recovered (P < 0.05) expression of 3 selenoprotein encoding genes (GPX1, SELENOH, and SELENOW) in a dose-dependent manner and increased (P < 0.05) expression of another 5 selenoprotein encoding genes (SELENOK, SELENOM, SELENOS, SELENOT, and TXNRD2) only at a high level (2.0 μmol Se/L). Taken together, LPS-induced immunological stress in RAW264.7 cells accompanied with the global downregulation of selenoprotein encoding genes and Se pretreatment alleviated immunological stress via upregulation of a subset of selenoprotein encoding genes.

Disruption of the psbA gene by the copy correction mechanism reveals that the expression of plastid-encoded genes is regulated by photosynthesis activity.

PubMed

Khan, Muhammad Sarwar; Hameed, Waqar; Nozoe, Mikio; Shiina, Takashi

2007-05-01

The functional analysis of genes encoded by the chloroplast genome of tobacco by reverse genetics is routine. Nevertheless, for a small number of genes their deletion generates heteroplasmic genotypes, complicating their analysis. There is thus the need for additional strategies to develop deletion mutants for these genes. We have developed a homologous copy correction-based strategy for deleting/mutating genes encoded on the chloroplast genome. This system was used to produce psbA knockouts. The resulting plants are homoplasmic and lack photosystem II (PSII) activity. Further, the deletion mutants exhibit a distinct phenotype; young leaves are green, whereas older leaves are bleached, irrespective of light conditions. This suggests that senescence is promoted by the absence of psbA. Analysis of the transcript levels indicates that NEP (nuclear-encoded plastid RNA polymerase)-dependent plastid genes are up regulated in the psbA deletion mutants, whereas the bleached leaves retain plastid-encoded plastid RNA polymerase activity. Hence, the expression of NEP-dependent plastid genes may be regulated by photosynthesis, either directly or indirectly.

A genetic variant in the gene encoding adrenomedullin predicts the development of dysglycemia over 6.4 years in Chinese.

PubMed

Ong, Kwok Leung; Tso, Annette W K; Leung, Raymond Y H; Cherny, Stacey S; Sham, Pak Chung; Lam, Tai Hing; Cheung, Bernard M Y; Lam, Karen S L

2011-01-30

Adrenomedullin, a vasodilatory peptide, facilitates the differentiation of pre-adipocytes, and affects lipolysis and glucose uptake. We investigated the association of common single nucleotide polymorphisms (SNPs) in the gene encoding adrenomedullin (ADM) with dysglycemia in the Hong Kong Chinese population. Four SNPs were genotyped in 1391 subjects without dysglycemia at baseline from the Hong Kong Cardiovascular Risk Factor Prevalence Study-2, which had a median follow-up time of 6.4 years. Dysglycemia included impaired fasting glucose, impaired glucose tolerance, and diabetes according to the WHO 1998 criteria. At follow-up, 382 subjects had developed dysglycemia. In stepwise logistic regression, the SNP rs11042725 was a significant independent predictor of the development of dysglycemia (OR=1.31, P=0.012), together with baseline age (P<0.001), plasma triglycerides (P<0.001), body mass index (P=0.004), 2-h glucose after oral glucose tolerance test (P<0.001), homeostasis model assessment of insulin resistance index (P=0.045), and follow-up duration (P=0.009). The association was more significant in women (P=0.002) and in subjects without regular exercise (P=0.001). Our study suggests a potential role of genetic variants in the ADM gene in the development of dysglycemia in our local Chinese population. Copyright © 2010 Elsevier B.V. All rights reserved.

Genome-Wide Identification and Mapping of NBS-Encoding Resistance Genes in Solanum tuberosum Group Phureja

PubMed Central

Lozano, Roberto; Ponce, Olga; Ramirez, Manuel; Mostajo, Nelly; Orjeda, Gisella

2012-01-01

The majority of disease resistance (R) genes identified to date in plants encode a nucleotide-binding site (NBS) and leucine-rich repeat (LRR) domain containing protein. Additional domains such as coiled-coil (CC) and TOLL/interleukin-1 receptor (TIR) domains can also be present. In the recently sequenced Solanum tuberosum group phureja genome we used HMM models and manual curation to annotate 435 NBS-encoding R gene homologs and 142 NBS-derived genes that lack the NBS domain. Highly similar homologs for most previously documented Solanaceae R genes were identified. A surprising ∼41% (179) of the 435 NBS-encoding genes are pseudogenes primarily caused by premature stop codons or frameshift mutations. Alignment of 81.80% of the 577 homologs to S. tuberosum group phureja pseudomolecules revealed non-random distribution of the R-genes; 362 of 470 genes were found in high density clusters on 11 chromosomes. PMID:22493716

Quantitative Profiling of Brain Lipid Raft Proteome in a Mouse Model of Fragile X Syndrome

PubMed Central

Kalinowska, Magdalena; Castillo, Catherine; Francesconi, Anna

2015-01-01

Fragile X Syndrome, a leading cause of inherited intellectual disability and autism, arises from transcriptional silencing of the FMR1 gene encoding an RNA-binding protein, Fragile X Mental Retardation Protein (FMRP). FMRP can regulate the expression of approximately 4% of brain transcripts through its role in regulation of mRNA transport, stability and translation, thus providing a molecular rationale for its potential pleiotropic effects on neuronal and brain circuitry function. Several intracellular signaling pathways are dysregulated in the absence of FMRP suggesting that cellular deficits may be broad and could result in homeostatic changes. Lipid rafts are specialized regions of the plasma membrane, enriched in cholesterol and glycosphingolipids, involved in regulation of intracellular signaling. Among transcripts targeted by FMRP, a subset encodes proteins involved in lipid biosynthesis and homeostasis, dysregulation of which could affect the integrity and function of lipid rafts. Using a quantitative mass spectrometry-based approach we analyzed the lipid raft proteome of Fmr1 knockout mice, an animal model of Fragile X syndrome, and identified candidate proteins that are differentially represented in Fmr1 knockout mice lipid rafts. Furthermore, network analysis of these candidate proteins reveals connectivity between them and predicts functional connectivity with genes encoding components of myelin sheath, axonal processes and growth cones. Our findings provide insight to aid identification of molecular and cellular dysfunctions arising from Fmr1 silencing and for uncovering shared pathologies between Fragile X syndrome and other autism spectrum disorders. PMID:25849048

The candidate histocompatibility locus of a Basal chordate encodes two highly polymorphic proteins.

PubMed

Nydam, Marie L; Netuschil, Nikolai; Sanders, Erin; Langenbacher, Adam; Lewis, Daniel D; Taketa, Daryl A; Marimuthu, Arumugapradeep; Gracey, Andrew Y; De Tomaso, Anthony W

2013-01-01

The basal chordate Botryllus schlosseri undergoes a natural transplantation reaction governed by a single, highly polymorphic locus called the fuhc. Our initial characterization of this locus suggested it encoded a single gene alternatively spliced into two transcripts: a 555 amino acid-secreted form containing the first half of the gene, and a full-length, 1008 amino acid transmembrane form, with polymorphisms throughout the ectodomain determining outcome. We have now found that the locus encodes two highly polymorphic genes which are separated by a 227 bp intergenic region: first, the secreted form as previously described, and a second gene encoding a 531 amino acid membrane-bound gene containing three extracellular immunoglobulin domains. While northern blotting revealed only these two mRNAs, both PCR and mRNA-seq detect a single capped and polyadenylated transcript that encodes processed forms of both genes linked by the intergenic region, as well as other transcripts in which exons of the two genes are spliced together. These results might suggest that the two genes are expressed as an operon, during which both genes are co-transcribed and then trans-spliced into two separate messages. This type of transcriptional regulation has been described in tunicates previously; however, the membrane-bound gene does not encode a typical Splice Leader (SL) sequence at the 5' terminus that usually accompanies trans-splicing. Thus, the presence of stable transcripts encoding both genes may suggest a novel mechanism of regulation, or conversely may be rare but stable transcripts in which the two mRNAs are linked due to a small amount of read-through by RNA polymerase. Both genes are highly polymorphic and co-expressed on tissues involved in histocompatibility. In addition, polymorphisms on both genes correlate with outcome, although we have found a case in which it appears that the secreted form may be major allorecognition determinant.

Development of a gene synthesis platform for the efficient large scale production of small genes encoding animal toxins.

PubMed

Sequeira, Ana Filipa; Brás, Joana L A; Guerreiro, Catarina I P D; Vincentelli, Renaud; Fontes, Carlos M G A

2016-12-01

Gene synthesis is becoming an important tool in many fields of recombinant DNA technology, including recombinant protein production. De novo gene synthesis is quickly replacing the classical cloning and mutagenesis procedures and allows generating nucleic acids for which no template is available. In addition, when coupled with efficient gene design algorithms that optimize codon usage, it leads to high levels of recombinant protein expression. Here, we describe the development of an optimized gene synthesis platform that was applied to the large scale production of small genes encoding venom peptides. This improved gene synthesis method uses a PCR-based protocol to assemble synthetic DNA from pools of overlapping oligonucleotides and was developed to synthesise multiples genes simultaneously. This technology incorporates an accurate, automated and cost effective ligation independent cloning step to directly integrate the synthetic genes into an effective Escherichia coli expression vector. The robustness of this technology to generate large libraries of dozens to thousands of synthetic nucleic acids was demonstrated through the parallel and simultaneous synthesis of 96 genes encoding animal toxins. An automated platform was developed for the large-scale synthesis of small genes encoding eukaryotic toxins. Large scale recombinant expression of synthetic genes encoding eukaryotic toxins will allow exploring the extraordinary potency and pharmacological diversity of animal venoms, an increasingly valuable but unexplored source of lead molecules for drug discovery.

Comprehensive search for accessory proteins encoded with archaeal and bacterial type III CRISPR-cas gene cassettes reveals 39 new cas gene families.

PubMed

Shah, Shiraz A; Alkhnbashi, Omer S; Behler, Juliane; Han, Wenyuan; She, Qunxin; Hess, Wolfgang R; Garrett, Roger A; Backofen, Rolf

2018-06-19

A study was undertaken to identify conserved proteins that are encoded adjacent to cas gene cassettes of Type III CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats - CRISPR associated) interference modules. Type III modules have been shown to target and degrade dsDNA, ssDNA and ssRNA and are frequently intertwined with cofunctional accessory genes, including genes encoding CRISPR-associated Rossman Fold (CARF) domains. Using a comparative genomics approach, and defining a Type III association score accounting for coevolution and specificity of flanking genes, we identified and classified 39 new Type III associated gene families. Most archaeal and bacterial Type III modules were seen to be flanked by several accessory genes, around half of which did not encode CARF domains and remain of unknown function. Northern blotting and interference assays in Synechocystis confirmed that one particular non-CARF accessory protein family was involved in crRNA maturation. Non-CARF accessory genes were generally diverse, encoding nuclease, helicase, protease, ATPase, transporter and transmembrane domains with some encoding no known domains. We infer that additional families of non-CARF accessory proteins remain to be found. The method employed is scalable for potential application to metagenomic data once automated pipelines for annotation of CRISPR-Cas systems have been developed. All accessory genes found in this study are presented online in a readily accessible and searchable format for researchers to audit their model organism of choice: http://accessory.crispr.dk .

Modularity of Plant Metabolic Gene Clusters: A Trio of Linked Genes That Are Collectively Required for Acylation of Triterpenes in Oat[W][OA

PubMed Central

Mugford, Sam T.; Louveau, Thomas; Melton, Rachel; Qi, Xiaoquan; Bakht, Saleha; Hill, Lionel; Tsurushima, Tetsu; Honkanen, Suvi; Rosser, Susan J.; Lomonossoff, George P.; Osbourn, Anne

2013-01-01

Operon-like gene clusters are an emerging phenomenon in the field of plant natural products. The genes encoding some of the best-characterized plant secondary metabolite biosynthetic pathways are scattered across plant genomes. However, an increasing number of gene clusters encoding the synthesis of diverse natural products have recently been reported in plant genomes. These clusters have arisen through the neo-functionalization and relocation of existing genes within the genome, and not by horizontal gene transfer from microbes. The reasons for clustering are not yet clear, although this form of gene organization is likely to facilitate co-inheritance and co-regulation. Oats (Avena spp) synthesize antimicrobial triterpenoids (avenacins) that provide protection against disease. The synthesis of these compounds is encoded by a gene cluster. Here we show that a module of three adjacent genes within the wider biosynthetic gene cluster is required for avenacin acylation. Through the characterization of these genes and their encoded proteins we present a model of the subcellular organization of triterpenoid biosynthesis. PMID:23532069

The trenbolone acetate affects the immune system in rainbow trout, Oncorhynchus mykiss.

PubMed

Massart, Sophie; Redivo, Baptiste; Flamion, Enora; Mandiki, S N M; Falisse, Elodie; Milla, Sylvain; Kestemont, Patrick

2015-06-01

In aquatic systems, the presence of endocrine-disrupting chemicals (EDC) can disrupt the reproductive function but also the immune system of wildlife. Some studies have investigated the effects of androgens on the fish immune parameters but the mechanisms by which the xenoandrogens alter the immunity are not well characterized. In order to test the effects of trenbolone acetate (TbA) on fish immune system, we exposed rainbow trout male juveniles during three weeks to TbA levels at 0.1 and 1μg/L. The present results suggest that TbA impacts, in a tissue-dependent manner, the rainbow trout immunity by affecting primarily the humoral immunity. Indeed, TbA inhibited lysozyme activity in plasma and liver and enhanced the alternative complement pathway activity (ACH50) in kidney. In plasma, the modulation of the complement system was time-dependent. The mRNA expression of genes encoding some cytokines such as renal TGF-β1, TNF-α in skin and hepatic IL-1β was also altered in fish exposed to TbA. Regarding the cellular immunity, no effect was observed on the leucocyte population. However, the expression of genes involved in the development and maturation of lymphoid cells (RAG-1 and RAG-2) was decreased in TbA-treated fish. Among those effects, we suggest that the modulation of RAG-1 and mucus apolipoprotein-A1 gene expression as well as plasma and hepatic lysozyme activities are mediated through the action of the androgen receptor. All combined, we conclude that trenbolone affects the rainbow trout immunity. Copyright © 2015 Elsevier B.V. All rights reserved.

Tetramer-organizing polyproline-rich peptides differ in CHO cell-expressed and plasma-derived human butyrylcholinesterase tetramers.

PubMed

Schopfer, Lawrence M; Lockridge, Oksana

2016-06-01

Tetrameric butyrylcholinesterase (BChE) in human plasma is the product of multiple genes, namely one BCHE gene on chromosome 3q26.1 and multiple genes that encode polyproline-rich peptides. The function of the polyproline-rich peptides is to assemble BChE into tetramers. CHO cells transfected with human BChE cDNA express BChE monomers and dimers, but only low quantities of tetramers. Our goal was to identify the polyproline-rich peptides in CHO-cell derived human BChE tetramers. CHO cell-produced human BChE tetramers were purified from serum-free culture medium. Peptides embedded in the tetramerization domain were released from BChE tetramers by boiling and identified by liquid chromatography-tandem mass spectrometry. A total of 270 proline-rich peptides were sequenced, ranging in size from 6-41 residues. The peptides originated from 60 different proteins that reside in multiple cell compartments including the nucleus, cytoplasm, and endoplasmic reticulum. No single protein was the source of the polyproline-rich peptides in CHO cell-expressed human BChE tetramers. In contrast, 70% of the tetramer-organizing peptides in plasma-derived BChE tetramers originate from lamellipodin. No protein source was identified for polyproline peptides containing up to 41 consecutive proline residues. In conclusion, the use of polyproline-rich peptides as a tetramerization motif is documented only for the cholinesterases, but is expected to serve other tetrameric proteins as well. The CHO cell data suggest that the BChE tetramer-organizing peptide can arise from a variety of proteins. Copyright © 2016 Elsevier B.V. All rights reserved.

Detection with synthetic oligonucleotide probes of nucleotide sequence variations in the genes encoding enterotoxins of Escherichia coli.

PubMed Central

Nishibuchi, M; Murakami, A; Arita, M; Jikuya, H; Takano, J; Honda, T; Miwatani, T

1989-01-01

We examined variations in the genes encoding heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT) in 88 strains of Escherichia coli isolated from individuals with traveler's diarrhea to find suitable sequences for use as oligonucleotide probes. Four oligonucleotide probes of the gene encoding ST of human origin (STIb or STh), one oligonucleotide probe of the gene encoding ST of porcine origin (STIa or STp), and three oligonucleotide probes of the gene encoding LT of human origin (LTIh) were used in DNA colony hybridization tests. In 15 of 22 strains possessing the STh gene and 28 of 42 strains producing LT, the sequences of all regions tested were identical to the published sequences. One region in the STh gene examined with a 18-mer probe was relatively well conserved and was shown to be closely associated with the enterotoxicity of the E. coli strains in suckling mice. This oligonucleotide, however, hybridized with strains of Vibrio cholerae O1, V. parahaemolyticus, and Yersinia enterocolitica that gave negative results in the suckling mouse assay. PMID:2685027

Cyclic stretch-induced the cytoskeleton rearrangement and gene expression of cytoskeletal regulators in human periodontal ligament cells.

PubMed

Wu, Yaqin; Zhuang, Jiabao; Zhao, Dan; Zhang, Fuqiang; Ma, Jiayin; Xu, Chun

2017-10-01

This study aimed to explore the mechanism of the stretch-induced cell realignment and cytoskeletal rearrangement by identifying several mechanoresponsive genes related to cytoskeletal regulators in human PDL cells. After the cells were stretched by 1, 10 and 20% strains for 0.5, 1, 2, 4, 6, 12 or 24 h, the changes of the morphology and content of microfilaments were recorded and calculated. Meanwhile, the expression of 84 key genes encoding cytoskeletal regulators after 6 and 24 h stretches with 20% strain was detected by using real-time PCR array. Western blot was applied to identify the protein expression level of several cytoskeletal regulators encoded by these differentially expressed genes. The confocal fluorescent staining results confirmed that stretch-induced realignment of cells and rearrangement of microfilaments. Among the 84 genes screened, one gene was up-regulated while two genes were down-regulated after 6 h stretch. Meanwhile, three genes were up-regulated while two genes were down-regulated after 24 h stretch. These genes displaying differential expression included genes regulating polymerization/depolymerization of microfilaments (CDC42EP2, FNBP1L, NCK2, PIKFYVE, WASL), polymerization/depolymerization of microtubules (STMN1), interacting between microfilaments and microtubules (MACF1), as well as a phosphatase (PPP1R12B). Among the proteins encoded by these genes, the protein expression level of Cdc42 effector protein-2 (encoded by CDC42EP2) and Stathmin-1 (encoded by STMN1) was down-regulated, while the protein expression level of N-WASP (encoded by WASL) was up-regulated. The present study confirmed the cyclic stretch-induced cellular realignment and rearrangement of microfilaments in the human PDL cells and indicated several force-sensitive genes with regard to cytoskeletal regulators.

A High-Resolution Gene Map of the Chloroplast Genome of the Red Alga Porphyra purpurea.

PubMed Central

Reith, M; Munholland, J

1993-01-01

Extensive DNA sequencing of the chloroplast genome of the red alga Porphyra purpurea has resulted in the detection of more than 125 genes. Fifty-eight (approximately 46%) of these genes are not found on the chloroplast genomes of land plants. These include genes encoding 17 photosynthetic proteins, three tRNAs, and nine ribosomal proteins. In addition, nine genes encoding proteins related to biosynthetic functions, six genes encoding proteins involved in gene expression, and at least five genes encoding miscellaneous proteins are among those not known to be located on land plant chloroplast genomes. The increased coding capacity of the P. purpurea chloroplast genome, along with other characteristics such as the absence of introns and the conservation of ancestral operons, demonstrate the primitive nature of the P. purpurea chloroplast genome. In addition, evidence for a monophyletic origin of chloroplasts is suggested by the identification of two groups of genes that are clustered in chloroplast genomes but not in cyanobacteria. PMID:12271072

Genome-Wide Architecture of Disease Resistance Genes in Lettuce

PubMed Central

Christopoulou, Marilena; Wo, Sebastian Reyes-Chin; Kozik, Alex; McHale, Leah K.; Truco, Maria-Jose; Wroblewski, Tadeusz; Michelmore, Richard W.

2015-01-01

Genome-wide motif searches identified 1134 genes in the lettuce reference genome of cv. Salinas that are potentially involved in pathogen recognition, of which 385 were predicted to encode nucleotide binding-leucine rich repeat receptor (NLR) proteins. Using a maximum-likelihood approach, we grouped the NLRs into 25 multigene families and 17 singletons. Forty-one percent of these NLR-encoding genes belong to three families, the largest being RGC16 with 62 genes in cv. Salinas. The majority of NLR-encoding genes are located in five major resistance clusters (MRCs) on chromosomes 1, 2, 3, 4, and 8 and cosegregate with multiple disease resistance phenotypes. Most MRCs contain primarily members of a single NLR gene family but a few are more complex. MRC2 spans 73 Mb and contains 61 NLRs of six different gene families that cosegregate with nine disease resistance phenotypes. MRC3, which is 25 Mb, contains 22 RGC21 genes and colocates with Dm13. A library of 33 transgenic RNA interference tester stocks was generated for functional analysis of NLR-encoding genes that cosegregated with disease resistance phenotypes in each of the MRCs. Members of four NLR-encoding families, RGC1, RGC2, RGC21, and RGC12 were shown to be required for 16 disease resistance phenotypes in lettuce. The general composition of MRCs is conserved across different genotypes; however, the specific repertoire of NLR-encoding genes varied particularly of the rapidly evolving Type I genes. These tester stocks are valuable resources for future analyses of additional resistance phenotypes. PMID:26449254

Cloning, characterization, expression analysis and inhibition studies of a novel gene encoding Bowman-Birk type protease inhibitor from rice bean

USDA-ARS?s Scientific Manuscript database

This paper presents the first study describing the isolation, cloning and characterization of a full length gene encoding Bowman-Birk protease inhibitor (RbTI) from rice bean (Vigna umbellata). A full-length protease inhibitor gene with complete open reading frame of 327bp encoding 109 amino acids w...

Cytochrome b5 gene and protein of Candida tropicalis and methods relating thereto

DOEpatents

Craft, David L.; Madduri, Krishna M.; Loper, John C.

2003-01-01

A novel gene has been isolated which encodes cytochrome b5 (CYTb5) protein of the .omega.-hydroxylase complex of C. tropicalis 20336. Vectors including this gene, and transformed host cells are provided. Methods of increasing the production of a CYTb5 protein are also provided which involve transforming a host cell with a gene encoding this protein and culturing the cells. Methods of increasing the production of a dicarboxylic acid are also provided which involve increasing in the host cell the number of genes encoding this protein.

Overexpression of a Plasma Membrane-Localized SbSRP-Like Protein Enhances Salinity and Osmotic Stress Tolerance in Transgenic Tobacco

PubMed Central

Udawat, Pushpika; Jha, Rajesh K.; Mishra, Avinash; Jha, Bhavanath

2017-01-01

An obligate halophyte, Salicornia brachiata grows in salt marshes and is considered to be a potential resource of salt- and drought-responsive genes. It is important to develop an understanding of the mechanisms behind enhanced salt tolerance. To increase this understanding, a novel SbSRP gene was cloned, characterized, over-expressed, and functionally validated in the model plant Nicotiana tabacum. The genome of the halophyte S. brachiata contains two homologs of an intronless SbSRP gene of 1,262 bp in length that encodes for a stress-related protein. An in vivo localization study confirmed that SbSRP is localized on the plasma membrane. Transgenic tobacco plants (T1) that constitutively over-express the SbSRP gene showed improved salinity and osmotic stress tolerance. In comparison to Wild Type (WT) and Vector Control (VC) plants, transgenic lines showed elevated relative water and chlorophyll content, lower malondialdehyde content, lower electrolyte leakage and higher accumulation of proline, free amino acids, sugars, polyphenols, and starch under abiotic stress treatments. Furthermore, a lower build-up of H2O2 content and superoxide-radicals was found in transgenic lines compared to WT and VC plants under stress conditions. Transcript expression of Nt-APX (ascorbate peroxidase), Nt-CAT (catalase), Nt-SOD (superoxide dismutase), Nt-DREB (dehydration responsive element binding factor), and Nt-AP2 (apetala2) genes was higher in transgenic lines under stress compared to WT and VC plants. The results suggested that overexpression of membrane-localized SbSRP mitigates salt and osmotic stress in the transgenic tobacco plant. It was hypothesized that SbSRP can be a transporter protein to transmit the environmental stimuli downward through the plasma membrane. However, a detailed study is required to ascertain its exact role in the abiotic stress tolerance mechanism. Overall, SbSRP is a potential candidate to be used for engineering salt and osmotic tolerance in crops. PMID:28473839

Glutathione S-transferase genes and the risk of type 2 diabetes mellitus: Role of sexual dimorphism, gene-gene and gene-smoking interactions in disease susceptibility.

PubMed

Azarova, Iuliia; Bushueva, Olga; Konoplya, Alexander; Polonikov, Alexey

2018-05-01

Compromised defense against reactive oxygen species (ROS) is considered important in the pathogenesis of type 2 diabetes mellitus (T2DM); therefore, genes encoding antioxidant defense enzymes may contribute to disease susceptibility. This study investigated whether polymorphisms in genes encoding glutathione S-transferase M1 (GSTM1), T1 (GSTT1), and P1 (GSTP1) jointly contribute to the risk of T2DM. In all, 1120 unrelated Russian subjects (600 T2DM patients, 520 age- and sex-matched healthy subjects), were recruited to the study. Genotyping was performed by multiplex polymerase chain reaction (PCR; del/del polymorphisms of GSTM1 and GSTT1) and TaqMan-based PCR (polymorphisms I105V and A114V of GSTP1). Plasma ROS and glutathione levels in study subjects were analyzed by fluorometric and colorimetric assays, respectively. Genotype del/del GSTT1 was significantly associated with the risk of T2DM (odds ratio [OR] 1.60, 95% confidence interval [CI] 1.17-2.21, P = 0.003). Gender-stratified analysis showed that the deletion genotypes of GSTM1 (OR 1.99, 95% CI 1.30-3.05; P = 0.0002, Q = 0.016) and GSTT1 (OR 2.23, 95% CI 1.22-4.09; P = 0.008, Q = 0.0216), as well as genotype 114A/V of GSTP1 (OR 2.85, 95% CI 1.44-5.62; P = 0.005, Q = 0.02) were associated with an increased risk of T2DM exclusively in males. Three genotype combinations (i.e. GSTM1+ × GSTT1+, GSTM1+ × GSTP1 114A/A and GSTT1+ × GSTP1 114A/A) showed significant associations with a decreased risk of T2DM in males. This study demonstrates, for the first time, that genes encoding glutathione S-transferases jointly contribute to the risk of T2DM, and that their effects on disease susceptibility are gender specific. © 2017 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and John Wiley & Sons Australia, Ltd.

Polymorphisms in methotrexate transporters and their relationship to plasma methotrexate levels, toxicity of high-dose methotrexate, and outcome of pediatric acute lymphoblastic leukemia.

PubMed

Liu, Shu-Guang; Gao, Chao; Zhang, Rui-Dong; Zhao, Xiao-Xi; Cui, Lei; Li, Wei-Jing; Chen, Zhen-Ping; Yue, Zhi-Xia; Zhang, Yuan-Yuan; Wu, Min-Yuan; Wang, Jian-Xiang; Li, Zhi-Gang; Zheng, Hu-Yong

2017-06-06

High-dose methotrexate (HDMTX) plays an important role in the treatment of acute lymphoblastic leukemia (ALL) although there is great inter-patient variability in the efficacy and toxicity of MTX. The relationship between polymorphisms in genes encoding MTX transporters and MTX response is controversial. In the present study, 322 Chinese children with standard- and intermediate-risk ALL were genotyped for 12 polymorphisms. SLCO1B1 rs10841753 showed a significant association with plasma MTX levels at 48 h (P = 0.017). Patients who had the ABCB1 rs1128503 C allele had longer duration of hospitalization than did those with the TT genotype (P = 0.006). No association was found between oral mucositis and any polymorphism. Long-term outcome was worse in patients with the SLCO1B1 rs4149056 CC genotype than in patients with TT or TC (5-year event-free survival [EFS] 33.3 ± 19.2% vs. 90.5 ± 1.7%, P < 0.001), and was worse in patients with the SCL19A1 rs2838958 AA genotype than in patients with AG or GG (5-year EFS 78.5 ± 4.6% vs. 92.2 ± 1.8%, P = 0.008). Multiple Cox regression analyses revealed associations of minimal residual disease (MRD) at day 33 (hazard ratio 3.458; P = 0.002), MRD at day 78 (hazard ratio 6.330; P = 0.001), SLCO1B1 rs4149056 (hazard ratio 12.242; P < 0.001), and SCL19A1 rs2838958 (hazard ratio 2.324; P = 0.019) with EFS. Our findings show that polymorphisms in genes encoding MTX transporters substantially influence the kinetics and response to HDMTX therapy in childhood ALL.

Adaptation of Saccharomyces cerevisiae to the Herbicide 2,4-Dichlorophenoxyacetic Acid, Mediated by Msn2p- and Msn4p-Regulated Genes: Important Role of SPI1

PubMed Central

Simões, T.; Teixeira, M. C.; Fernandes, A. R.; Sá-Correia, Isabel

2003-01-01

The possible roles of 13 Msn2p- and Msn4p-regulated genes in the adaptation of Saccharomyces cerevisiae to the herbicide 2,4-d-dichlorophenoxyacetic acid (2,4-D) were examined. Single deletion of genes involved in defense against oxidizing agents (CTT1, GRX1, and GRX2/TTR1) or encoding chaperones of the HSP70 family (SSA1, SSA4, and SSE2) showed a slight effect. A more significant role was observed for the heat shock genes HSP78, HSP26, HSP104, HSP12, and HSP42, most of which encode molecular chaperones. However, the SPI1 gene, encoding a member of the glycosylphosphatidylinositol-anchored cell wall protein family, emerged as the major determinant of 2,4-D resistance. SPI1 expression reduced the loss of viability of an unadapted yeast population suddenly exposed to the herbicide, allowing earlier growth resumption. Significantly, yeast adaptation to 2,4-D involves the rapid and transient Msn2p- and Msn4p-mediated activation (fivefold) of SPI1 transcription. SPI1 mRNA levels were reduced to values slightly above those in unstressed cells when the adapted population started duplication in the presence of 2,4-D. Since SPI1 deletion leads to the higher β-1,3-glucanase sensitivity of 2,4-D-stressed cells, it was hypothesized that adaptation may involve an Spi1p-mediated increase in the diffusional restriction of the liposoluble acid form of the herbicide across the cell envelope. Such a cell response would avoid a futile cycle due to acid reentry into the cell counteracting the active export of the anionic form, presumably through an inducible plasma membrane transporter(s). Consistent with this concept, the concentration of 14C-labeled 2,4-D in 2,4-D-energized adapted Δspi1 mutant cells and the consequent intracellular acidification are higher than in wild-type cells. PMID:12839777

Perturbations of carotenoid and tetrapyrrole biosynthetic pathways result in differential alterations in chloroplast function and plastid signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Joon-Heum; Jung, Sunyo

In this study, we used the biosynthetic inhibitors of carotenoid and tetrapyrrole biosynthetic pathways, norflurazon (NF) and oxyfluorfen (OF), as tools to gain insight into mechanisms of photooxidation in rice plants. NF resulted in bleaching symptom on leaves of the treated plants, whereas OF treatment developed a fast symptom of an apparent necrotic phenotype. Both plants exhibited decreases in photosynthetic efficiency, as indicated by F{sub v}/F{sub m}. NF caused severe disruption in thylakoid membranes, whereas OF-treated plants exhibited disruption of chloroplast envelope and plasma membrane. Levels of Lhca and Lhcb proteins in photosystem I (PSI) and PSII were reduced bymore » photooxidative stress in NF- and OF-treated plants, with a greater decrease in NF plants. The down-regulation of nuclear-encoded photosynthesis genes Lhcb and rbcS was also found in both NF- and OF-treated plants, whereas plastid-encoded photosynthetic genes including RbcL, PsaC, and PsbD accumulated normally in NF plants but decreased drastically in OF plants. This proposes that the plastids in NF plants retain their potential to develop thylakoid membranes and that photobleaching is mainly controlled by nuclear genes. Distinct photooxidation patterns between NF- and OF-treated plants developed differential signaling, which might enable the plant to coordinate the expression of photosynthetic genes from the nuclear and plastidic genomes. - Highlights: • Two modes of photooxidation by carotenoid and tetrapyrrole biosynthetic inhibitors. • We examine differential alterations in chloroplast function and plastid signaling. • NF and OF cause differential alterations in chloroplast ultrastructure and function. • Photooxidation coordinates photosynthetic gene expression from nucleus and plastid.« less

Plant glycosylphosphatidylinositol (GPI) anchored proteins at the plasma membrane-cell wall nexus.

PubMed

Yeats, Trevor H; Bacic, Antony; Johnson, Kim L

2018-04-18

Approximately 1% of plant proteins are predicted to be post-translationally modified with a glycosylphosphatidylinositol (GPI) anchor that tethers the polypeptide to the outer leaflet of the plasma membrane. While the synthesis and structure of GPI anchors is largely conserved across eukaryotes, the repertoire of functional domains present in the GPI-anchored proteome has diverged substantially. In plants, this includes a large fraction of the GPI-anchored proteome being further modified with plant-specific arabinogalactan (AG) O-glycans. The importance of the GPI-anchored proteome to plant development is underscored by the fact that GPI biosynthetic null mutants exhibit embryo lethality. Mutations in genes encoding specific GPI-anchored proteins (GAPs) further supports their contribution to diverse biological processes occurring at the interface of the plasma membrane and cell wall, including signaling, cell wall metabolism, cell wall polymer cross-linking, and plasmodesmatal transport. Here, we review the literature concerning plant GPI-anchored proteins in the context of their potential to act as molecular hubs that mediate interactions between the plasma membrane and the cell wall and their potential to transduce the signal into the protoplast and thereby activate signal transduction pathways. This article is protected by copyright. All rights reserved.

Genome complexity in the coelacanth is reflected in its adaptive immune system

USGS Publications Warehouse

Saha, Nil Ratan; Ota, Tatsuya; Litman, Gary W.; Hansen, John; Parra, Zuly; Hsu, Ellen; Buonocore, Francesco; Canapa, Adriana; Cheng, Jan-Fang; Amemiya, Chris T.

2014-01-01

We have analyzed the available genome and transcriptome resources from the coelacanth in order to characterize genes involved in adaptive immunity. Two highly distinctive IgW-encoding loci have been identified that exhibit a unique genomic organization, including a multiplicity of tandemly repeated constant region exons. The overall organization of the IgW loci precludes typical heavy chain class switching. A locus encoding IgM could not be identified either computationally or by using several different experimental strategies. Four distinct sets of genes encoding Ig light chains were identified. This includes a variant sigma-type Ig light chain previously identified only in cartilaginous fishes and which is now provisionally denoted sigma-2. Genes encoding α/β and γ/δ T-cell receptors, and CD3, CD4, and CD8 co-receptors also were characterized. Ig heavy chain variable region genes and TCR components are interspersed within the TCR α/δ locus; this organization previously was reported only in tetrapods and raises questions regarding evolution and functional cooption of genes encoding variable regions. The composition, organization and syntenic conservation of the major histocompatibility complex locus have been characterized. We also identified large numbers of genes encoding cytokines and their receptors, and other genes associated with adaptive immunity. In terms of sequence identity and organization, the adaptive immune genes of the coelacanth more closely resemble orthologous genes in tetrapods than those in teleost fishes, consistent with current phylogenomic interpretations. Overall, the work reported described herein highlights the complexity inherent in the coelacanth genome and provides a rich catalog of immune genes for future investigations.

The Bacillus subtilis ywjI (glpX) gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the class III Fbp enzyme.

PubMed

Jules, Matthieu; Le Chat, Ludovic; Aymerich, Stéphane; Le Coq, Dominique

2009-05-01

We present here experimental evidence that the Bacillus subtilis ywjI gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the fbp-encoded class III enzyme, and constitutes with the upstream gene, murAB, an operon transcribed at the same level under glycolytic or gluconeogenic conditions.

The Bacillus subtilis ywjI (glpX) Gene Encodes a Class II Fructose-1,6-Bisphosphatase, Functionally Equivalent to the Class III Fbp Enzyme▿

PubMed Central

Jules, Matthieu; Le Chat, Ludovic; Aymerich, Stéphane; Le Coq, Dominique

2009-01-01

We present here experimental evidence that the Bacillus subtilis ywjI gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the fbp-encoded class III enzyme, and constitutes with the upstream gene, murAB, an operon transcribed at the same level under glycolytic or gluconeogenic conditions. PMID:19270101

Analysis of CFB, a cytokinin-responsive gene of Arabidopsis thaliana encoding a novel F-box protein regulating sterol biosynthesis.

PubMed

Brenner, Wolfram G; Leuendorf, Jan Erik; Cortleven, Anne; Martin, Laetitia B B; Schaller, Hubert; Schmülling, Thomas

2017-05-17

Protein degradation by the ubiquitin-26S proteasome pathway is important for the regulation of cellular processes, but the function of most F-box proteins relevant to substrate recognition is unknown. We describe the analysis of the gene Cytokinin-induced F-box encoding (CFB, AT3G44326), identified in a meta-analysis of cytokinin-related transcriptome studies as one of the most robust cytokinin response genes. F-box domain-dependent interaction with the E3 ubiquitin ligase complex component ASK1 classifies CFB as a functional F-box protein. Apart from F-box and transmembrane domains, CFB contains no known functional domains. CFB is expressed in all plant tissues, predominantly in root tissue. A ProCFB:GFP-GUS fusion gene showed strongest expression in the lateral root cap and during lateral root formation. CFB-GFP fusion proteins were mainly localized in the nucleus and the cytosol but also at the plasma membrane. cfb mutants had no discernible phenotype, but CFB overexpressing plants showed several defects, such as a white upper inflorescence stem, similar to the hypomorphic cycloartenol synthase mutant cas1-1. Both CFB overexpressing plants and cas1-1 mutants accumulated the CAS1 substrate 2,3-oxidosqualene in the white stem tissue, the latter even more after cytokinin treatment, indicating impairment of CAS1 function. This suggests that CFB may link cytokinin and the sterol biosynthesis pathway. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Rare loss-of-function mutations in ANGPTL family members contribute to plasma triglyceride levels in humans

PubMed Central

Romeo, Stefano; Yin, Wu; Kozlitina, Julia; Pennacchio, Len A.; Boerwinkle, Eric; Hobbs, Helen H.; Cohen, Jonathan C.

2008-01-01

The relative activity of lipoprotein lipase (LPL) in different tissues controls the partitioning of lipoprotein-derived fatty acids between sites of fat storage (adipose tissue) and oxidation (heart and skeletal muscle). Here we used a reverse genetic strategy to test the hypothesis that 4 angiopoietin-like proteins (ANGPTL3, -4, -5, and -6) play key roles in triglyceride (TG) metabolism in humans. We re-sequenced the coding regions of the genes encoding these proteins and identified multiple rare nonsynonymous (NS) sequence variations that were associated with low plasma TG levels but not with other metabolic phenotypes. Functional studies revealed that all mutant alleles of ANGPTL3 and ANGPTL4 that were associated with low plasma TG levels interfered either with the synthesis or secretion of the protein or with the ability of the ANGPTL protein to inhibit LPL. A total of 1% of the Dallas Heart Study population and 4% of those participants with a plasma TG in the lowest quartile had a rare loss-of-function mutation in ANGPTL3, ANGPTL4, or ANGPTL5. Thus, ANGPTL3, ANGPTL4, and ANGPTL5, but not ANGPTL6, play nonredundant roles in TG metabolism, and multiple alleles at these loci cumulatively contribute to variability in plasma TG levels in humans. PMID:19075393

Targeting the AtCWIN1 Gene to Explore the Role of Invertases in Sucrose Transport in Roots and during Botrytis cinerea Infection

PubMed Central

Veillet, Florian; Gaillard, Cécile; Coutos-Thévenot, Pierre; La Camera, Sylvain

2016-01-01

Cell wall invertases (CWIN) cleave sucrose into glucose and fructose in the apoplast. CWINs are key regulators of carbon partitioning and source/sink relationships during growth, development and under biotic stresses. In this report, we monitored the expression/activity of Arabidopsis cell wall invertases in organs behaving as source, sink, or subjected to a source/sink transition after infection with the necrotrophic fungus Botrytis cinerea. We showed that organs with different source/sink status displayed differential CWIN activities, depending on carbohydrate needs or availabilities in the surrounding environment, through a transcriptional and posttranslational regulation. Loss-of-function mutation of the Arabidopsis cell wall invertase 1 gene, AtCWIN1, showed that the corresponding protein was the main contributor to the apoplastic sucrose cleaving activity in both leaves and roots. The CWIN-deficient mutant cwin1-1 exhibited a reduced capacity to actively take up external sucrose in roots, indicating that this process is mainly dependent on the sucrolytic activity of AtCWIN1. Using T-DNA and CRISPR/Cas9 mutants impaired in hexose transport, we demonstrated that external sucrose is actively absorbed in the form of hexoses by a sugar/H+ symport system involving the coordinated activity of AtCWIN1 with several Sugar Transporter Proteins (STP) of the plasma membrane, i.e., STP1 and STP13. Part of external sucrose was imported without apoplastic cleavage into cwin1-1 seedling roots, highlighting an alternative AtCWIN1-independent pathway for the assimilation of external sucrose. Accordingly, we showed that several genes encoding sucrose transporters of the plasma membrane were expressed. We also detected transcript accumulation of vacuolar invertase (VIN)-encoding genes and high VIN activities. Upon infection, AtCWIN1 was responsible for all the Botrytis-induced apoplastic invertase activity. We detected a transcriptional activation of several AtSUC and AtVIN genes accompanied with an enhanced vacuolar invertase activity, suggesting that the AtCWIN1-independent pathway is efficient upon infection. In absence of AtCWIN1, we postulate that intracellular sucrose hydrolysis is sufficient to provide intracellular hexoses to maintain sugar homeostasis in host cells and to fuel plant defenses. Finally, we demonstrated that Botrytis cinerea possesses its own functional sucrolytic machinery and hexose uptake system, and does not rely on the host apoplastic invertases. PMID:28066461

Differential Responses of Plasma Adropin Concentrations To Dietary Glucose or Fructose Consumption In Humans

PubMed Central

Butler, Andrew A.; St-Onge, Marie-Pierre; Siebert, Emily A.; Medici, Valentina; Stanhope, Kimber L.; Havel, Peter J.

2015-01-01

Adropin is a peptide hormone encoded by the Energy Homeostasis Associated (ENHO) gene whose physiological role in humans remains incompletely defined. Here we investigated the impact of dietary interventions that affect systemic glucose and lipid metabolism on plasma adropin concentrations in humans. Consumption of glucose or fructose as 25% of daily energy requirements (E) differentially affected plasma adropin concentrations (P < 0.005) irrespective of duration, sex or age. Glucose consumption reduced plasma adropin from 3.55 ± 0.26 to 3.28 ± 0.23 ng/ml (N = 42). Fructose consumption increased plasma adropin from 3.63 ± 0.29 to 3.93 ± 0.34 ng/ml (N = 45). Consumption of high fructose corn syrup (HFCS) as 25% E had no effect (3.43 ± 0.32 versus 3.39 ± 0.24 ng/ml, N = 26). Overall, the effect of glucose, HFCS and fructose on circulating adropin concentrations were similar to those observed on postprandial plasma triglyceride concentrations. Furthermore, increases in plasma adropin levels with fructose intake were most robust in individuals exhibiting hypertriglyceridemia. Individuals with low plasma adropin concentrations also exhibited rapid increases in plasma levels following consumption of breakfasts supplemented with lipids. These are the first results linking plasma adropin levels with dietary sugar intake in humans, with the impact of fructose consumption linked to systemic triglyceride metabolism. In addition, dietary fat intake may also increase circulating adropin concentrations. PMID:26435060

Molecular comparison of the structural proteins encoding gene clusters of two related Lactobacillus delbrueckii bacteriophages.

PubMed Central

Vasala, A; Dupont, L; Baumann, M; Ritzenthaler, P; Alatossava, T

1993-01-01

Virulent phage LL-H and temperate phage mv4 are two related bacteriophages of Lactobacillus delbrueckii. The gene clusters encoding structural proteins of these two phages have been sequenced and further analyzed. Six open reading frames (ORF-1 to ORF-6) were detected. Protein sequencing and Western immunoblotting experiments confirmed that ORF-3 (g34) encoded the main capsid protein Gp34. The presence of a putative late promoter in front of the phage LL-H g34 gene was suggested by primer extension experiments. Comparative sequence analysis between phage LL-H and phage mv4 revealed striking similarities in the structure and organization of this gene cluster, suggesting that the genes encoding phage structural proteins belong to a highly conservative module. Images PMID:8497043

The identification of aluminium-resistance genes provides opportunities for enhancing crop production on acid soils.

PubMed

Ryan, P R; Tyerman, S D; Sasaki, T; Furuichi, T; Yamamoto, Y; Zhang, W H; Delhaize, E

2011-01-01

Acid soils restrict plant production around the world. One of the major limitations to plant growth on acid soils is the prevalence of soluble aluminium (Al(3+)) ions which can inhibit root growth at micromolar concentrations. Species that show a natural resistance to Al(3+) toxicity perform better on acid soils. Our understanding of the physiology of Al(3+) resistance in important crop plants has increased greatly over the past 20 years, largely due to the application of genetics and molecular biology. Fourteen genes from seven different species are known to contribute to Al(3+) tolerance and resistance and several additional candidates have been identified. Some of these genes account for genotypic variation within species and others do not. One mechanism of resistance which has now been identified in a range of species relies on the efflux of organic anions such as malate and citrate from roots. The genes controlling this trait are members of the ALMT and MATE families which encode membrane proteins that facilitate organic anion efflux across the plasma membrane. Identification of these and other resistance genes provides opportunities for enhancing the Al(3+) resistance of plants by marker-assisted breeding and through biotechnology. Most attempts to enhance Al(3+) resistance in plants with genetic engineering have targeted genes that are induced by Al(3+) stress or that are likely to increase organic anion efflux. In the latter case, studies have either enhanced organic anion synthesis or increased organic anion transport across the plasma membrane. Recent developments in this area are summarized and the structure-function of the TaALMT1 protein from wheat is discussed.

Bacillus subtilis 168 Contains Two Differentially Regulated Genes Encoding l-Asparaginase

PubMed Central

Fisher, Susan H.; Wray, Lewis V.

2002-01-01

Expression of the two Bacillus subtilis genes encoding l-asparaginase is controlled by independent regulatory factors. The ansZ gene (formerly yccC) was shown by mutational analysis to encode a functional l-asparaginase, the expression of which is activated during nitrogen-limited growth by the TnrA transcription factor. Gel mobility shift and DNase I footprinting experiments indicate that TnrA regulates ansZ expression by binding to a DNA site located upstream of the ansZ promoter. The expression of the ansA gene, which encodes the second l-asparaginase, was found to be induced by asparagine. The ansA repressor, AnsR, was shown to negatively regulate its own expression. PMID:11914346

Bacillus subtilis 168 contains two differentially regulated genes encoding L-asparaginase.

PubMed

Fisher, Susan H; Wray, Lewis V

2002-04-01

Expression of the two Bacillus subtilis genes encoding L-asparaginase is controlled by independent regulatory factors. The ansZ gene (formerly yccC) was shown by mutational analysis to encode a functional L-asparaginase, the expression of which is activated during nitrogen-limited growth by the TnrA transcription factor. Gel mobility shift and DNase I footprinting experiments indicate that TnrA regulates ansZ expression by binding to a DNA site located upstream of the ansZ promoter. The expression of the ansA gene, which encodes the second L-asparaginase, was found to be induced by asparagine. The ansA repressor, AnsR, was shown to negatively regulate its own expression.

Recombinant DNA encoding a desulfurization biocatalyst

DOEpatents

Rambosek, John; Piddington, Chris S.; Kovacevich, Brian R.; Young, Kevin D.; Denome, Sylvia A.

1994-01-01

This invention relates to a recombinant DNA molecule containing a gene or genes which encode a biocatalyst capable of desulfurizing a fossil fuel which contains organic sulfur molecules. For example, the present invention encompasses a recombinant DNA molecule containing a gene or genes of a strain of Rhodococcus rhodochrous.

Cell Surface Expression of Human Ether-a-go-go-related Gene (hERG) Channels Is Regulated by Caveolin-3 Protein via the Ubiquitin Ligase Nedd4-2*

PubMed Central

Guo, Jun; Wang, Tingzhong; Li, Xian; Shallow, Heidi; Yang, Tonghua; Li, Wentao; Xu, Jianmin; Fridman, Michael D.; Yang, Xiaolong; Zhang, Shetuan

2012-01-01

The human ether-a-go-go-related gene (hERG) encodes the rapidly activating delayed rectifier potassium channel (IKr) which plays an important role in cardiac repolarization. A reduction or increase in hERG current can cause long or short QT syndrome, respectively, leading to fatal cardiac arrhythmias. The channel density in the plasma membrane is a key determinant of the whole cell current amplitude. To gain insight into the molecular mechanisms for the regulation of hERG density at the plasma membrane, we used whole cell voltage clamp, Western blotting, and immunocytochemical methods to investigate the effects of an integral membrane protein, caveolin-3 (Cav3) on hERG expression levels. Our data demonstrate that Cav3, hERG, and ubiquitin-ligase Nedd4-2 interact with each other and form a complex. Expression of Cav3 thus enhances the hERG-Nedd4-2 interaction, leading to an increased ubiquitination and degradation of mature, plasma-membrane localized hERG channels. Disrupting Nedd4-2 interaction with hERG by mutations eliminates the effects of Cav3 on hERG channels. Knockdown of endogenous Cav3 or Nedd4-2 in cultured neonatal rat ventricular myocytes using siRNA led to an increase in native IKr. Our data demonstrate that hERG expression in the plasma membrane is regulated by Cav3 via Nedd4-2. These findings extend our understanding of the regulation of hERG channels and cardiac electrophysiology. PMID:22879586

Increased plasma chemokine levels in children with Prader-Willi syndrome.

PubMed

Butler, Merlin G; Hossain, Waheeda; Sulsona, Carlos; Driscoll, Daniel J; Manzardo, Ann M

2015-03-01

Prader-Willi syndrome (PWS) is caused by loss of paternally expressed genes from the 15q11-q13 region and reportedly rearranged as a cause of autism. Additionally, increased inflammatory markers and features of autism are reported in PWS. Cytokines encoded by genes involved with inflammation, cell proliferation, migration, and adhesion play a role in neurodevelopment and could be disturbed in PWS as abnormal plasma cytokine levels are reported in autism. We analyzed 41 plasma cytokines in a cohort of well-characterized children with PWS between 5 and 11 years of age and unaffected unrelated siblings using multiplex sandwich immunoassays with the Luminex magnetic-bead based platform. Data were analyzed using ANOVA testing for effects of diagnosis, gender, body mass index (BMI) and age on the 24 cytokines meeting laboratory criteria for inclusion. No significant effects were observed for age, gender or BMI. The log-transformed levels of the 24 analyzable cytokines were examined simultaneously using MANOVA adjusting for age and gender and a main effect of diagnosis was found (P-value <0.03). Four of 24 plasma cytokine levels (MCP1, MDC, Eotaxin, RANTES) were significantly higher in children with PWS compared with controls and classified as bioinflammatory chemokines supporting a disturbed immune response unrelated to obesity status. BMI was not statistically different in the two subject groups (PWS or unaffected unrelated siblings) and chemokine levels were not correlated with percentage of total body fat. Additional studies are required to identify whether possible early immunological disturbances and chemokine inflammatory processes found in PWS may contribute to neurodevelopment and behavioral features. © 2015 Wiley Periodicals, Inc.

A highly divergent gene cluster in honey bees encodes a novel silk family.

PubMed

Sutherland, Tara D; Campbell, Peter M; Weisman, Sarah; Trueman, Holly E; Sriskantha, Alagacone; Wanjura, Wolfgang J; Haritos, Victoria S

2006-11-01

The pupal cocoon of the domesticated silk moth Bombyx mori is the best known and most extensively studied insect silk. It is not widely known that Apis mellifera larvae also produce silk. We have used a combination of genomic and proteomic techniques to identify four honey bee fiber genes (AmelFibroin1-4) and two silk-associated genes (AmelSA1 and 2). The four fiber genes are small, comprise a single exon each, and are clustered on a short genomic region where the open reading frames are GC-rich amid low GC intergenic regions. The genes encode similar proteins that are highly helical and predicted to form unusually tight coiled coils. Despite the similarity in size, structure, and composition of the encoded proteins, the genes have low primary sequence identity. We propose that the four fiber genes have arisen from gene duplication events but have subsequently diverged significantly. The silk-associated genes encode proteins likely to act as a glue (AmelSA1) and involved in silk processing (AmelSA2). Although the silks of honey bees and silkmoths both originate in larval labial glands, the silk proteins are completely different in their primary, secondary, and tertiary structures as well as the genomic arrangement of the genes encoding them. This implies independent evolutionary origins for these functionally related proteins.

Photocontrol of the expression of genes encoding chlorophyll a/b binding proteins and small subunit of ribulose-1,5-bisphosphate carboxylase in etiolated seedlings of Lycopersicon esculentum (L. ) and Nicotiana tabacum (L. )

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wehmeyer, B.; Cashmore, A.R.; Schaefer, E.

Phytochrome and the blue ultraviolet-A photoreceptor control light-induced expression of genes encoding the chlorophyll a/b binding protein of photosystem II and photosystem I and the genes for the small subunit of the ribulose-1,5-bisphosphate carboxylase in etiolated seedlings of Lycopersicon esculentum (tomato) and Nicotiana tabacum (tobacco). A high irradiance response also controls the induction of these genes. Genes encoding photosystem II- and I-associated chlorophyll a/b binding proteins both exhibit a transient rapid increase in expression in response to light pulse or to continuous irradiation. In contrast, genes encoding the small subunit exhibit a continuous increase in expression in response to light.more » These distinct expression characteristics are shown to reflect differences at the level of transcription.« less

The evolution of genes encoding for green fluorescent proteins: insights from cephalochordates (amphioxus)

NASA Astrophysics Data System (ADS)

Yue, Jia-Xing; Holland, Nicholas D.; Holland, Linda Z.; Deheyn, Dimitri D.

2016-06-01

Green Fluorescent Protein (GFP) was originally found in cnidarians, and later in copepods and cephalochordates (amphioxus) (Branchiostoma spp). Here, we looked for GFP-encoding genes in Asymmetron, an early-diverged cephalochordate lineage, and found two such genes closely related to some of the Branchiostoma GFPs. Dim fluorescence was found throughout the body in adults of Asymmetron lucayanum, and, as in Branchiostoma floridae, was especially intense in the ripe ovaries. Spectra of the fluorescence were similar between Asymmetron and Branchiostoma. Lineage-specific expansion of GFP-encoding genes in the genus Branchiostoma was observed, largely driven by tandem duplications. Despite such expansion, purifying selection has strongly shaped the evolution of GFP-encoding genes in cephalochordates, with apparent relaxation for highly duplicated clades. All cephalochordate GFP-encoding genes are quite different from those of copepods and cnidarians. Thus, the ancestral cephalochordates probably had GFP, but since GFP appears to be lacking in more early-diverged deuterostomes (echinoderms, hemichordates), it is uncertain whether the ancestral cephalochordates (i.e. the common ancestor of Asymmetron and Branchiostoma) acquired GFP by horizontal gene transfer (HGT) from copepods or cnidarians or inherited it from the common ancestor of copepods and deuterostomes, i.e. the ancestral bilaterians.

Identification and characterization of the gltK gene encoding a membrane-associated glucose transport protein of pseudomonas aeruginosa.

PubMed

Adewoye, L O; Worobec, E A

2000-08-08

The Pseudomonas aeruginosa oprB gene encodes the carbohydrate-selective OprB porin, which translocates substrate molecules across the outer membrane to the periplasmic glucose-binding protein. We identified and cloned two open reading frames (ORFs) flanking the oprB gene but are not in operonic arrangement with the oprB gene. The downstream ORF encodes a putative polypeptide homologous to members of a family of transcriptional repressors, whereas the oprB gene is preceded by an ORF encoding a putative product, which exhibits strong homology to several carbohydrate transport ATP-binding cassette (ABC) proteins. The genomic copy of the upstream ORF was mutagenized by homologous recombination. Analysis of the deletion mutant in comparison with the wild type revealed a significant reduction in [14C] glucose transport activity in the mutant strain, suggesting that this ORF likely encodes the inner membrane component of the glucose ABC transporter. It is thus designated gltK gene to reflect its homology to the Pseudomona fluorescens mtlK and its involvement in the high-affinity glucose transport system. Multiple alignment analysis revealed that the P. aeruginosa gltK gene product is a member of the MalK subfamily of ABC proteins.

Comparative transcriptomic evidence for Tween80-enhanced biodegradation of phenanthrene by Sphingomonas sp. GY2B.

PubMed

Liu, Shasha; Guo, Chuling; Lin, Weijia; Wu, Fengji; Lu, Guining; Lu, Jing; Dang, Zhi

2017-12-31

Previous study of the effects of surfactants on the biodegradation of phenanthrene focused on investigating alterations of the cell characteristics of Sphingomonas sp. GY2B. However, genes regulation associated with biodegradation and biological processes in response to the presence of surfactants, remains unclear. In this study, comparative transcriptome analysis was conducted to observe the gene expression of GY2B during phenanthrene biodegradation in the presence and absence of Tween80. A diverse set of genes was regulated by Tween80, leading to increased biodegradation of phenanthrene by GY2B: (i) Tween80 increased expression of genes related to H + transport in the plasma membrane to provide a driving force (i.e., ATP) for accelerating transmembrane transport of phenanthrene with increasing Tween80 concentrations, thereby enhancing the uptake and degradation of phenanthrene by GY2B; (ii) Tween80 (1 and 8 CMC) promoted intracellular biodegradation of phenanthrene by stimulating expression of genes encoding dioxygenases and monooxygenase, increasing expression of genes involved in intracellular metabolic processes (e.g., TCA cycle); and (iii) Tween80 likely increased GY2B vitality and growth by inducing expression of genes associated with ABC transporters and protein transport, regulating genes involved in other biological processes (e.g., transcription, translation). Copyright © 2017. Published by Elsevier B.V.

Prevalence, antibiotic resistance, virulence traits and genetic lineages of Staphylococcus aureus in healthy sheep in Tunisia.

PubMed

Gharsa, Haythem; Ben Slama, Karim; Lozano, Carmen; Gómez-Sanz, Elena; Klibi, Naouel; Ben Sallem, Rym; Gómez, Paula; Zarazaga, Myriam; Boudabous, Abdellatif; Torres, Carmen

2012-05-04

Nasal swabs of 163 healthy sheep were obtained from two farms and one abattoir in Tunisia during 2010. Samples were inoculated in Baird Parker agar and ORSAB medium for Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) recovery, respectively. MRSA was detected in 5 of these 163 samples (3%) in ORSAB medium, and one isolate per sample was further studied. MRSA isolates were mecA-positive, typed as ST153-CC80-t044-agrIII, and contained blaZ, ant(6)-Ia, aph(3')-IIIa, erm(C), tet(K), and fusB genes encoding penicillin, streptomycin, kanamycin, erythromycin, tetracycline and fusidic acid resistance, respectively. These MRSA isolates showed indistinguishable or closely related PFGE-patterns and harboured the lukF/lukS gene encoding the Panton-Valentine leukocidin and the luk-ED, hla, hld, and hlg(v) genes. Methicillin-susceptible S. aureus (MSSA) were recovered in 68 of the 163 samples (41.7%) and one isolate per sample was characterized. Most of MSSA (82.4%) showed susceptibility to the tested antibiotics with exceptions: penicillin (6%, carrying blaZ gene), tetracycline (19%, carrying tet(K) gene) and fusidic acid (9%). The following toxin-genes were identified among MSSA: tst (53 isolates), luk-M (52), luk-ED, hla, hlb, hld and hlg(v) (67), hlg (1), sec (49), sel (52), and the egc-cluster-like sen-sem-sei-seo-seg (1). Ten spa-types (two of them new ones) and nine sequence types (six new ones) were detected among the 73 S. aureus isolates, and they were ascribed to agr types I and III. All MRSA and MSSA isolates were able to coagulate bovine plasma and MRSA harboured the immune-evasion-gene-cluster type E. Conclusions. Nares of healthy sheep could be a reservoir of PVL-positive community-associated-MRSA and also of TSST-positive S. aureus isolates, with potential implications in public health. Copyright © 2011 Elsevier B.V. All rights reserved.

Recombinant DNA encoding a desulfurization biocatalyst

DOEpatents

Rambosek, J.; Piddington, C.S.; Kovacevich, B.R.; Young, K.D.; Denome, S.A.

1994-10-18

This invention relates to a recombinant DNA molecule containing a gene or genes which encode a biocatalyst capable of desulfurizing a fossil fuel which contains organic sulfur molecules. For example, the present invention encompasses a recombinant DNA molecule containing a gene or genes of a strain of Rhodococcus rhodochrous. 13 figs.

Structure, Function, Interaction, Co-evolution of Rice Blast Resistance Genes

USDA-ARS?s Scientific Manuscript database

Rice blast disease caused by the fungal pathogen Magnaporthe oryzae is one of the most destructive rice diseases worldwide. Resistance (R) genes to blast encode proteins that detect pathogen signaling molecules encoded by M. oryzae avirulence (AVR) genes. R genes can be a single or a member of clu...

Development of Murine Lupus Involves the Combined Genetic Contribution of the SLAM and FcγR Intervals within the Nba2 Autoimmune Susceptibility Locus

PubMed Central

Jørgensen, Trine N.; Alfaro, Jennifer; Enriquez, Hilda L.; Jiang, Chao; Loo, William M.; Atencio, Stephanie; Bupp, Melanie R. Gubbels; Mailloux, Christina M.; Metzger, Troy; Flannery, Shannon; Rozzo, Stephen J.; Kotzin, Brian L.; Rosemblatt, Mario; Bono, María Rosa; Erickson, Loren D.

2010-01-01

Autoantibodies are of central importance in the pathogenesis of Ab-mediated autoimmune disorders. The murine lupus susceptibility locus Nba2 on chromosome 1 and the syntenic human locus are associated with a loss of immune tolerance that leads to antinuclear Ab production. To identify gene intervals within Nba2 that control the development of autoantibody-producing B cells and to determine the cellular components through which Nba2 genes accomplish this, we generated congenic mice expressing various Nba2 intervals where genes for the FcγR, SLAM, and IFN-inducible families are encoded. Analysis of congenic strains demonstrated that the FcγR and SLAM intervals independently controlled the severity of autoantibody production and renal disease, yet are both required for lupus susceptibility. Deregulated homeostasis of terminally differentiated B cells was found to be controlled by the FcγR interval where FcγRIIb-mediated apoptosis of germinal center B cells and plasma cells was impaired. Increased numbers of activated plasmacytoid dendritic cells that were distinctly CD19+ and promoted plasma cell differentiation via the proinflammatory cytokines IL-10 and IFNα were linked to the SLAM interval. These findings suggest that SLAM and FcγR intervals act cooperatively to influence the clinical course of disease through supporting the differentiation and survival of autoantibody-producing cells. PMID:20018631

Molecular genetics of Erwinia amylovora involved in the development of fire blight.

PubMed

Oh, Chang-Sik; Beer, Steven V

2005-12-15

The bacterial plant pathogen, Erwinia amylovora, causes the devastating disease known as fire blight in some Rosaceous plants like apple, pear, quince, raspberry and several ornamentals. Knowledge of the factors affecting the development of fire blight has mushroomed in the last quarter century. On the molecular level, genes encoding a Hrp type III secretion system, genes encoding enzymes involved in synthesis of extracellular polysaccharides and genes facilitating the growth of E. amylovora in its host plants have been characterized. The Hrp pathogenicity island, delimited by genes suggesting horizontal gene transfer, is composed of four distinct regions, the hrp/hrc region, the HEE (Hrp effectors and elicitors) region, the HAE (Hrp-associated enzymes) region, and the IT (Island transfer) region. The Hrp pathogenicity island encodes a Hrp type III secretion system (TTSS), which delivers several proteins from bacteria to plant apoplasts or cytoplasm. E. amylovora produces two exopolysaccharides, amylovoran and levan, which cause the characteristic fire blight wilting symptom in host plants. In addition, other genes, and their encoded proteins, have been characterized as virulence factors of E. amylovora that encode enzymes facilitating sorbitol metabolism, proteolytic activity and iron harvesting. This review summarizes our understanding of the genes and gene products of E. amylovora that are involved in the development of the fire blight disease.

The Drosophila pigmentation gene pink (p) encodes a homologue of human Hermansky-Pudlak syndrome 5 (HPS5).

PubMed

Falcón-Pérez, Juan M; Romero-Calderón, Rafael; Brooks, Elizabeth S; Krantz, David E; Dell'Angelica, Esteban C

2007-02-01

Lysosome-related organelles comprise a group of specialized intracellular compartments that include melanosomes and platelet dense granules (in mammals) and eye pigment granules (in insects). In humans, the biogenesis of these organelles is defective in genetic disorders collectively known as Hermansky-Pudlak syndrome (HPS). Patients with HPS-2, and two murine HPS models, carry mutations in genes encoding subunits of adaptor protein (AP)-3. Other genes mutated in rodent models include those encoding VPS33A and Rab38. Orthologs of all of these genes in Drosophila melanogaster belong to the 'granule group' of eye pigmentation genes. Other genes associated with HPS encode subunits of three complexes of unknown function, named biogenesis of lysosome-related organelles complex (BLOC)-1, -2 and -3, for which the Drosophila counterparts had not been characterized. Here, we report that the gene encoding the Drosophila ortholog of the HPS5 subunit of BLOC-2 is identical to the granule group gene pink (p), which was first studied in 1910 but had not been identified at the molecular level. The phenotype of pink mutants was exacerbated by mutations in AP-3 subunits or in the orthologs of VPS33A and Rab38. These results validate D. melanogaster as a genetic model to study the function of the BLOCs.

Isolation and characterization of cDNA clones for human erythrocyte beta-spectrin.

PubMed Central

Prchal, J T; Morley, B J; Yoon, S H; Coetzer, T L; Palek, J; Conboy, J G; Kan, Y W

1987-01-01

Spectrin is an important structural component of the membrane skeleton that underlies and supports the erythrocyte plasma membrane. It is composed of nonidentical alpha (Mr 240,000) and beta (Mr 220,000) subunits, each of which contains multiple homologous 106-amino acid segments. We report here the isolation and characterization of a human erythroid-specific beta-spectrin cDNA clone that encodes parts of the beta-9 through beta-12 repeat segments. This cDNA was used as a hybridization probe to assign the beta-spectrin gene to human chromosome 14 and to begin molecular analysis of the gene and its mRNA transcripts. RNA transfer blot analysis showed that the reticulocyte beta-spectrin mRNA is 7.8 kilobases in length. Southern blot analysis of genomic DNA revealed the presence of restriction fragment length polymorphisms (RFLPs) within the beta-spectrin gene locus. The isolation of human spectrin cDNA probes and the identification of closely linked RFLPs will facilitate analysis of mutant spectrin genes causing congenital hemolytic anemias associated with quantitative and qualitative spectrin abnormalities. Images PMID:3478706

Novel Mutation in the ATP-Binding Cassette Transporter A3 (ABCA3) Encoding Gene Causes Respiratory Distress Syndrome in A Term Newborn in Southwest Iran

PubMed Central

Rezaei, Farideh; Shafiei, Mohammad; Shariati, Gholamreza; Dehdashtian, Ali; Mohebbi, Maryam; Galehdari, Hamid

2016-01-01

Introduction ABCA3 glycoprotein belongs to the ATP-binding cassette (ABC) superfamily of transporters, which utilize the energy derived from hydrolysis of ATP for the translocation of a wide variety of substrates across the plasma membrane. Mutations in the ABCA3 gene are knowingly causative for fatal surfactant deficiency, particularly respiratory distress syndrome (RDS) in term babies. Case Presentation In this study, Sanger sequencing of the whole ABCA3 gene (NCBI NM_001089) was performed in a neonatal boy with severe RDS. A homozygous mutation has been identified in the patient. Parents were heterozygous for the same missense mutation GGA > AGA at position 202 in exon 6 of the ABCA3 gene (c.604G > A; p.G202R). Furthermore, 70 normal individuals have been analyzed for the mentioned change with negative results. Conclusions Regarding Human Genome Mutation Database (HGMD) and other literature recherche, the detected change is a novel mutation and has not been reported before. Bioinformatics mutation predicting tools prefer it as pathogenic. PMID:27437095

Determination of CFTR densities in erythrocyte plasma membranes using recognition imaging

NASA Astrophysics Data System (ADS)

Ebner, Andreas; Nikova, Dessy; Lange, Tobias; Häberle, Johannes; Falk, Sabine; Dübbers, Angelika; Bruns, Reimer; Hinterdorfer, Peter; Oberleithner, Hans; Schillers, Hermann

2008-09-01

CFTR (cystic fibrosis transmembrane conductance regulator) is a cAMP-regulated chloride (Cl-) channel that plays an important role in salt and fluid movement across epithelia. Cystic fibrosis (CF), the most common genetic disease among Caucasians, is caused by mutations in the gene encoding CFTR. The most predominant mutation, F508del, disturbs CFTR protein trafficking, resulting in a reduced number of CFTR in the plasma membrane. Recent studies indicate that CFTR is not only found in epithelia but also in human erythrocytes. Although considerable attempts have been made to quantify CFTR in cells, conclusions on numbers of CFTR molecules localized in the plasma membrane have been drawn indirectly. AFM has the power to provide the needed information, since both sub-molecular spatial resolution and direct protein recognition via antibody-antigen interaction can be observed. We performed a quantification study of the CFTR copies in erythrocyte membranes at the single molecule level, and compared the difference between healthy donors and CF patients. We detected that the number of CFTR molecules is reduced by 70% in erythrocytes of cystic fibrosis patients.

Chlorella viruses contain genes encoding a complete polyamine biosynthetic pathway

PubMed Central

Baumann, Sascha; Sander, Adrianne; Gurnon, James R.; Yanai-Balser, Giane; VanEtten, James L.; Piotrowski, Markus

2007-01-01

Two genes encoding the putative polyamine biosynthetic enzymes agmatine iminohydrolase (AIH) and N-carbamoylputrescine amidohydrolase (CPA) were cloned from the chloroviruses PBCV-1, NY-2A and MT325. They were expressed in Escherichia coli to form C-terminal (His)6-tagged proteins and the recombinant proteins were purified by Ni2+- binding affinity chromatography. The biochemical properties of the two enzymes are similar to AIH and CPA enzymes from Arabidopsis thaliana and Pseudomonas aeruginosa. Together with the previously known virus genes encoding ornithine/arginine decarboxlyase (ODC/ADC) and homospermidine synthase, the chloroviruses have genes that encode a complete set of functional enzymes that synthesize the rare polyamine homospermidine from arginine via agmatine, N-carbamoylputrescine and putrescine. The PBCV-1 aih and cpa genes are expressed early during virus infection together with the odc/adc gene, suggesting that biosynthesis of putrescine is important in early stages of viral replication. The aih and cpa genes are widespread in the chlorella viruses. PMID:17101165

A Comprehensive Analysis of Nuclear-Encoded Mitochondrial Genes in Schizophrenia.

PubMed

Gonçalves, Vanessa F; Cappi, Carolina; Hagen, Christian M; Sequeira, Adolfo; Vawter, Marquis P; Derkach, Andriy; Zai, Clement C; Hedley, Paula L; Bybjerg-Grauholm, Jonas; Pouget, Jennie G; Cuperfain, Ari B; Sullivan, Patrick F; Christiansen, Michael; Kennedy, James L; Sun, Lei

2018-05-01

The genetic risk factors of schizophrenia (SCZ), a severe psychiatric disorder, are not yet fully understood. Multiple lines of evidence suggest that mitochondrial dysfunction may play a role in SCZ, but comprehensive association studies are lacking. We hypothesized that variants in nuclear-encoded mitochondrial genes influence susceptibility to SCZ. We conducted gene-based and gene-set analyses using summary association results from the Psychiatric Genomics Consortium Schizophrenia Phase 2 (PGC-SCZ2) genome-wide association study comprising 35,476 cases and 46,839 control subjects. We applied the MAGMA method to three sets of nuclear-encoded mitochondrial genes: oxidative phosphorylation genes, other nuclear-encoded mitochondrial genes, and genes involved in nucleus-mitochondria crosstalk. Furthermore, we conducted a replication study using the iPSYCH SCZ sample of 2290 cases and 21,621 control subjects. In the PGC-SCZ2 sample, 1186 mitochondrial genes were analyzed, among which 159 had p values < .05 and 19 remained significant after multiple testing correction. A meta-analysis of 818 genes combining the PGC-SCZ2 and iPSYCH samples resulted in 104 nominally significant and nine significant genes, suggesting a polygenic model for the nuclear-encoded mitochondrial genes. Gene-set analysis, however, did not show significant results. In an in silico protein-protein interaction network analysis, 14 mitochondrial genes interacted directly with 158 SCZ risk genes identified in PGC-SCZ2 (permutation p = .02), and aldosterone signaling in epithelial cells and mitochondrial dysfunction pathways appeared to be overrepresented in this network of mitochondrial and SCZ risk genes. This study provides evidence that specific aspects of mitochondrial function may play a role in SCZ, but we did not observe its broad involvement even using a large sample. Copyright © 2018 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

[Effect of FABP2 gene G54A polymorphism on lipid and glucose metabolism in simple obesity children].

PubMed

Xu, Yunpeng; Rao, Xiaojiao; Hao, Min; Hou, Lijuan; Zhu, Xiaobo; Chang, Xiaotong

2016-01-01

To explore the relationship between intestinal fatty acid binding protein (FABP2) gene G54A polymorphism and simple childhood obesity, the effect of mutant 54A FABP2 gene on serum lipids and glucose metabolism. The total of 83 subjects with overweight/obesity and 100 subjects with healthy/normal weight were involved in this study. The G54A FABP2 gene allele and genotype frequencies between control group and overweight/obesity group were detected using polymerase chain reaction (PCR) -restriction fragment length polymorphism (RFLP) technology, and DNA sequences were confirmed by DNA sequencing. The automatic biochemical analyzer was used to detect fasting blood glucose (FBG), triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) levels. Plasma insulin (Ins) was detected by radiation immune method, free fatty acids (FFA) was tested by ELISA method, insulin resistance index ( HOMA-IR ) was also calculated. The correlation between FABP2 G54A polymorphism and the development of children' obesity was analyzed. The relation between FABP2 G54A polymorphism and abnormal blood lipid and insulin resistance was assessed. The results of study on FABP2 gene polymorphism revealed as followed. In overweight/obese groups, the frequencies of GG, GA, AA genotypes was 33.7%, 49.4% and 16.9%, respectively. In control group, the frequencies of GG, GA, AA genotypes was 51. 0% , 40. 0% and 9. 0% , respectively. The differences between two groups was statistically significant (Χ2 = 6.27, P < 0.05). In overweight/obesity group, the frequencies of alleles were 58.4% for 54G and 41.6% for 54A. In control group, the frequencies of alleles were 71.0% for 54G and 29.0% for 54A. There was significant differences (Χ2 = 6.32, P < 0.05). The plasma biochemical variables results showed that compared with the normal control group, plasma TG (P < 0.01), Ins (P < 0.05), HOMA-IR (P < 0.05) were elevated in overweight/obesity group, the difference between two groups was statistically significant. At the same time, in overweight/obesity group, the carriers of AA homozygous genotypes had significantly higher plasma TG levels than those with GG wild genotypes (P < 0.05). A increased tendency of plasma Ins, FFA levels and HOMA-IR was found in the carriers with AA homozygous genotypes, but no differences compared with those with GG wild genotypes. Compared with those with GG wild genotypes, related plasma biochemical variables in the carriers with GA heterozygous genotypes had no differences (P > 0.05). The FABP2 gene G54A polymorphism is related to simple children obesity and lipid metabolism abnormality. The allele encoding in FABP2 gene may be a potential factor contributing to promoting lipid metabolism abnormality of and insulin resistance.

The ribosomal protein genes and Minute loci of Drosophila melanogaster

PubMed Central

Marygold, Steven J; Roote, John; Reuter, Gunter; Lambertsson, Andrew; Ashburner, Michael; Millburn, Gillian H; Harrison, Paul M; Yu, Zhan; Kenmochi, Naoya; Kaufman, Thomas C; Leevers, Sally J; Cook, Kevin R

2007-01-01

Background Mutations in genes encoding ribosomal proteins (RPs) have been shown to cause an array of cellular and developmental defects in a variety of organisms. In Drosophila melanogaster, disruption of RP genes can result in the 'Minute' syndrome of dominant, haploinsufficient phenotypes, which include prolonged development, short and thin bristles, and poor fertility and viability. While more than 50 Minute loci have been defined genetically, only 15 have so far been characterized molecularly and shown to correspond to RP genes. Results We combined bioinformatic and genetic approaches to conduct a systematic analysis of the relationship between RP genes and Minute loci. First, we identified 88 genes encoding 79 different cytoplasmic RPs (CRPs) and 75 genes encoding distinct mitochondrial RPs (MRPs). Interestingly, nine CRP genes are present as duplicates and, while all appear to be functional, one member of each gene pair has relatively limited expression. Next, we defined 65 discrete Minute loci by genetic criteria. Of these, 64 correspond to, or very likely correspond to, CRP genes; the single non-CRP-encoding Minute gene encodes a translation initiation factor subunit. Significantly, MRP genes and more than 20 CRP genes do not correspond to Minute loci. Conclusion This work answers a longstanding question about the molecular nature of Minute loci and suggests that Minute phenotypes arise from suboptimal protein synthesis resulting from reduced levels of cytoribosomes. Furthermore, by identifying the majority of haplolethal and haplosterile loci at the molecular level, our data will directly benefit efforts to attain complete deletion coverage of the D. melanogaster genome. PMID:17927810

Genetic modifiers of muscular dystrophy act on sarcolemmal resealing and recovery from injury

PubMed Central

Quattrocelli, Mattia; Capote, Joanna; Ohiri, Joyce C.; Warner, James L.; Vo, Andy H.; Hadhazy, Michele; Demonbreun, Alexis R.; Spencer, Melissa J.; McNally, Elizabeth M.

2017-01-01

Genetic disruption of the dystrophin complex produces muscular dystrophy characterized by a fragile muscle plasma membrane leading to excessive muscle degeneration. Two genetic modifiers of Duchenne Muscular Dystrophy implicate the transforming growth factor β (TGFβ) pathway, osteopontin encoded by the SPP1 gene and latent TGFβ binding protein 4 (LTBP4). We now evaluated the functional effect of these modifiers in the context of muscle injury and repair to elucidate their mechanisms of action. We found that excess osteopontin exacerbated sarcolemmal injury, and correspondingly, that loss of osteopontin reduced injury extent both in isolated myofibers and in muscle in vivo. We found that ablation of osteopontin was associated with reduced expression of TGFβ and TGFβ-associated pathways. We identified that increased TGFβ resulted in reduced expression of Anxa1 and Anxa6, genes encoding key components of the muscle sarcolemma resealing process. Genetic manipulation of Ltbp4 in dystrophic muscle also directly modulated sarcolemmal resealing, and Ltbp4 alleles acted in concert with Anxa6, a distinct modifier of muscular dystrophy. These data provide a model in which a feed forward loop of TGFβ and osteopontin directly impacts the capacity of muscle to recover from injury, and identifies an intersection of genetic modifiers on muscular dystrophy. PMID:29065150

Genetic modifiers of muscular dystrophy act on sarcolemmal resealing and recovery from injury.

PubMed

Quattrocelli, Mattia; Capote, Joanna; Ohiri, Joyce C; Warner, James L; Vo, Andy H; Earley, Judy U; Hadhazy, Michele; Demonbreun, Alexis R; Spencer, Melissa J; McNally, Elizabeth M

2017-10-01

Genetic disruption of the dystrophin complex produces muscular dystrophy characterized by a fragile muscle plasma membrane leading to excessive muscle degeneration. Two genetic modifiers of Duchenne Muscular Dystrophy implicate the transforming growth factor β (TGFβ) pathway, osteopontin encoded by the SPP1 gene and latent TGFβ binding protein 4 (LTBP4). We now evaluated the functional effect of these modifiers in the context of muscle injury and repair to elucidate their mechanisms of action. We found that excess osteopontin exacerbated sarcolemmal injury, and correspondingly, that loss of osteopontin reduced injury extent both in isolated myofibers and in muscle in vivo. We found that ablation of osteopontin was associated with reduced expression of TGFβ and TGFβ-associated pathways. We identified that increased TGFβ resulted in reduced expression of Anxa1 and Anxa6, genes encoding key components of the muscle sarcolemma resealing process. Genetic manipulation of Ltbp4 in dystrophic muscle also directly modulated sarcolemmal resealing, and Ltbp4 alleles acted in concert with Anxa6, a distinct modifier of muscular dystrophy. These data provide a model in which a feed forward loop of TGFβ and osteopontin directly impacts the capacity of muscle to recover from injury, and identifies an intersection of genetic modifiers on muscular dystrophy.

The Full-Size ABCG Transporters Nb-ABCG1 and Nb-ABCG2 Function in Pre- and Postinvasion Defense against Phytophthora infestans in Nicotiana benthamiana

PubMed Central

Shibata, Yusuke; Ojika, Makoto; Sugiyama, Akifumi; Yazaki, Kazufumi; Jones, David A.; Kawakita, Kazuhito

2016-01-01

The sesquiterpenoid capsidiol is the major phytoalexin produced by Nicotiana and Capsicum species. Capsidiol is produced in plant tissues attacked by pathogens and plays a major role in postinvasion defense by inhibiting pathogen growth. Using virus-induced gene silencing-based screening, we identified two Nicotiana benthamiana (wild tobacco) genes encoding functionally redundant full-size ABCG (PDR-type) transporters, Nb-ABCG1/PDR1 and Nb-ABCG2/PDR2, which are essential for resistance to the potato late blight pathogen Phytophthora infestans. Silencing of Nb-ABCG1/2 compromised secretion of capsidiol, revealing Nb-ABCG1/2 as probable exporters of capsidiol. Accumulation of plasma membrane-localized Nb-ABCG1 and Nb-ABCG2 was observed at the site of pathogen penetration. Silencing of EAS (encoding 5-epi-aristolochene synthase), a gene for capsidiol biosynthesis, reduced resistance to P. infestans, but penetration by P. infestans was not affected. By contrast, Nb-ABCG1/2-silenced plants showed reduced penetration defense, indicating that Nb-ABCG1/2 are involved in preinvasion defense against P. infestans. Plastidic GGPPS1 (geranylgeranyl diphosphate synthase) was also found to be required for preinvasion defense, thereby suggesting that plastid-produced diterpene(s) are the antimicrobial compounds active in preinvasion defense. These findings suggest that N. benthamiana ABCG1/2 are involved in the export of both antimicrobial diterpene(s) for preinvasion defense and capsidiol for postinvasion defense against P. infestans. PMID:27102667

Identifying metabolic enzymes with multiple types of association evidence

PubMed Central

Kharchenko, Peter; Chen, Lifeng; Freund, Yoav; Vitkup, Dennis; Church, George M

2006-01-01

Background Existing large-scale metabolic models of sequenced organisms commonly include enzymatic functions which can not be attributed to any gene in that organism. Existing computational strategies for identifying such missing genes rely primarily on sequence homology to known enzyme-encoding genes. Results We present a novel method for identifying genes encoding for a specific metabolic function based on a local structure of metabolic network and multiple types of functional association evidence, including clustering of genes on the chromosome, similarity of phylogenetic profiles, gene expression, protein fusion events and others. Using E. coli and S. cerevisiae metabolic networks, we illustrate predictive ability of each individual type of association evidence and show that significantly better predictions can be obtained based on the combination of all data. In this way our method is able to predict 60% of enzyme-encoding genes of E. coli metabolism within the top 10 (out of 3551) candidates for their enzymatic function, and as a top candidate within 43% of the cases. Conclusion We illustrate that a combination of genome context and other functional association evidence is effective in predicting genes encoding metabolic enzymes. Our approach does not rely on direct sequence homology to known enzyme-encoding genes, and can be used in conjunction with traditional homology-based metabolic reconstruction methods. The method can also be used to target orphan metabolic activities. PMID:16571130

Identification and Characterization of LFD-2, a Predicted Fringe Protein Required for Membrane Integrity during Cell Fusion in Neurospora crassa

PubMed Central

Palma-Guerrero, Javier; Zhao, Jiuhai; Gonçalves, A. Pedro; Starr, Trevor L.

2015-01-01

The molecular mechanisms of membrane merger during somatic cell fusion in eukaryotic species are poorly understood. In the filamentous fungus Neurospora crassa, somatic cell fusion occurs between genetically identical germinated asexual spores (germlings) and between hyphae to form the interconnected network characteristic of a filamentous fungal colony. In N. crassa, two proteins have been identified to function at the step of membrane fusion during somatic cell fusion: PRM1 and LFD-1. The absence of either one of these two proteins results in an increase of germling pairs arrested during cell fusion with tightly appressed plasma membranes and an increase in the frequency of cell lysis of adhered germlings. The level of cell lysis in ΔPrm1 or Δlfd-1 germlings is dependent on the extracellular calcium concentration. An available transcriptional profile data set was used to identify genes encoding predicted transmembrane proteins that showed reduced expression levels in germlings cultured in the absence of extracellular calcium. From these analyses, we identified a mutant (lfd-2, for late fusion defect-2) that showed a calcium-dependent cell lysis phenotype. lfd-2 encodes a protein with a Fringe domain and showed endoplasmic reticulum and Golgi membrane localization. The deletion of an additional gene predicted to encode a low-affinity calcium transporter, fig1, also resulted in a strain that showed a calcium-dependent cell lysis phenotype. Genetic analyses showed that LFD-2 and FIG1 likely function in separate pathways to regulate aspects of membrane merger and repair during cell fusion. PMID:25595444

[High gene conversion frequency between genes encoding 2-deoxyglucose-6-phosphate phosphatase in 3 Saccharomyces species].

PubMed

Piscopo, Sara-Pier; Drouin, Guy

2014-05-01

Gene conversions are nonreciprocal sequence exchanges between genes. They are relatively common in Saccharomyces cerevisiae, but few studies have investigated the evolutionary fate of gene conversions or their functional impacts. Here, we analyze the evolution and impact of gene conversions between the two genes encoding 2-deoxyglucose-6-phosphate phosphatase in S. cerevisiae, Saccharomyces paradoxus and Saccharomyces mikatae. Our results demonstrate that the last half of these genes are subject to gene conversions among these three species. The greater similarity and the greater percentage of GC nucleotides in the converted regions, as well as the absence of long regions of adjacent common converted sites, suggest that these gene conversions are frequent and occur independently in all three species. The high frequency of these conversions probably result from the fact that they have little impact on the protein sequences encoded by these genes.

Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks.

PubMed

Damiani, Isabelle; Drain, Alice; Guichard, Marjorie; Balzergue, Sandrine; Boscari, Alexandre; Boyer, Jean-Christophe; Brunaud, Véronique; Cottaz, Sylvain; Rancurel, Corinne; Da Rocha, Martine; Fizames, Cécile; Fort, Sébastien; Gaillard, Isabelle; Maillol, Vincent; Danchin, Etienne G J; Rouached, Hatem; Samain, Eric; Su, Yan-Hua; Thouin, Julien; Touraine, Bruno; Puppo, Alain; Frachisse, Jean-Marie; Pauly, Nicolas; Sentenac, Hervé

2016-01-01

Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as "papilionoid legume-specific" were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This transcriptome provides a valuable resource to investigate root hair biology in legumes and the roles that these cells play in rhizobial symbiosis establishment. These results could also contribute to the long-term objective of transferring this symbiotic capacity to non-legume plants.

Pepper pathogenesis-related protein 4c is a plasma membrane-localized cysteine protease inhibitor that is required for plant cell death and defense signaling.

PubMed

Kim, Nak Hyun; Hwang, Byung Kook

2015-01-01

Xanthomonas campestris pv. vesicatoria (Xcv) type III effector AvrBsT triggers programmed cell death (PCD) and activates the hypersensitive response (HR) in plants. Here, we isolated and identified the plasma membrane localized pathogenesis-related (PR) protein 4c gene (CaPR4c) from pepper (Capsicum annuum) leaves undergoing AvrBsT-triggered HR cell death. CaPR4c encodes a protein with a signal peptide and a Barwin domain. Recombinant CaPR4c protein expressed in Escherichia coli exhibited cysteine protease-inhibitor activity and ribonuclease (RNase) activity. Subcellular localization analyses revealed that CaPR4c localized to the plasma membrane in plant cells. CaPR4c expression was rapidly and specifically induced by avirulent Xcv (avrBsT) infection. Transient expression of CaPR4c caused HR cell death in pepper leaves, which was accompanied by enhanced accumulation of H2 O2 and significant induction of some defense-response genes. Deletion of the signal peptide from CaPR4c abolished the induction of HR cell death, indicating a requirement for plasma membrane localization of CaPR4c for HR cell death. CaPR4c silencing in pepper disrupted both basal and AvrBsT-triggered resistance responses, and enabled Xcv proliferation in infected leaves. H2 O2 accumulation, cell-death induction, and defense-response gene expression were distinctly reduced in CaPR4c-silenced pepper. CaPR4c overexpression in transgenic Arabidopsis plants conferred greater resistance against infection by Pseudomonas syringae pv. tomato and Hyaloperonospora arabidopsidis. These results collectively suggest that CaPR4c plays an important role in plant cell death and defense signaling. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Loss-of-function mutations in APOC3, triglycerides, and coronary disease.

PubMed

Crosby, Jacy; Peloso, Gina M; Auer, Paul L; Crosslin, David R; Stitziel, Nathan O; Lange, Leslie A; Lu, Yingchang; Tang, Zheng-zheng; Zhang, He; Hindy, George; Masca, Nicholas; Stirrups, Kathleen; Kanoni, Stavroula; Do, Ron; Jun, Goo; Hu, Youna; Kang, Hyun Min; Xue, Chenyi; Goel, Anuj; Farrall, Martin; Duga, Stefano; Merlini, Pier Angelica; Asselta, Rosanna; Girelli, Domenico; Olivieri, Oliviero; Martinelli, Nicola; Yin, Wu; Reilly, Dermot; Speliotes, Elizabeth; Fox, Caroline S; Hveem, Kristian; Holmen, Oddgeir L; Nikpay, Majid; Farlow, Deborah N; Assimes, Themistocles L; Franceschini, Nora; Robinson, Jennifer; North, Kari E; Martin, Lisa W; DePristo, Mark; Gupta, Namrata; Escher, Stefan A; Jansson, Jan-Håkan; Van Zuydam, Natalie; Palmer, Colin N A; Wareham, Nicholas; Koch, Werner; Meitinger, Thomas; Peters, Annette; Lieb, Wolfgang; Erbel, Raimund; Konig, Inke R; Kruppa, Jochen; Degenhardt, Franziska; Gottesman, Omri; Bottinger, Erwin P; O'Donnell, Christopher J; Psaty, Bruce M; Ballantyne, Christie M; Abecasis, Goncalo; Ordovas, Jose M; Melander, Olle; Watkins, Hugh; Orho-Melander, Marju; Ardissino, Diego; Loos, Ruth J F; McPherson, Ruth; Willer, Cristen J; Erdmann, Jeanette; Hall, Alistair S; Samani, Nilesh J; Deloukas, Panos; Schunkert, Heribert; Wilson, James G; Kooperberg, Charles; Rich, Stephen S; Tracy, Russell P; Lin, Dan-Yu; Altshuler, David; Gabriel, Stacey; Nickerson, Deborah A; Jarvik, Gail P; Cupples, L Adrienne; Reiner, Alex P; Boerwinkle, Eric; Kathiresan, Sekar

2014-07-03

Plasma triglyceride levels are heritable and are correlated with the risk of coronary heart disease. Sequencing of the protein-coding regions of the human genome (the exome) has the potential to identify rare mutations that have a large effect on phenotype. We sequenced the protein-coding regions of 18,666 genes in each of 3734 participants of European or African ancestry in the Exome Sequencing Project. We conducted tests to determine whether rare mutations in coding sequence, individually or in aggregate within a gene, were associated with plasma triglyceride levels. For mutations associated with triglyceride levels, we subsequently evaluated their association with the risk of coronary heart disease in 110,970 persons. An aggregate of rare mutations in the gene encoding apolipoprotein C3 (APOC3) was associated with lower plasma triglyceride levels. Among the four mutations that drove this result, three were loss-of-function mutations: a nonsense mutation (R19X) and two splice-site mutations (IVS2+1G→A and IVS3+1G→T). The fourth was a missense mutation (A43T). Approximately 1 in 150 persons in the study was a heterozygous carrier of at least one of these four mutations. Triglyceride levels in the carriers were 39% lower than levels in noncarriers (P<1×10(-20)), and circulating levels of APOC3 in carriers were 46% lower than levels in noncarriers (P=8×10(-10)). The risk of coronary heart disease among 498 carriers of any rare APOC3 mutation was 40% lower than the risk among 110,472 noncarriers (odds ratio, 0.60; 95% confidence interval, 0.47 to 0.75; P=4×10(-6)). Rare mutations that disrupt APOC3 function were associated with lower levels of plasma triglycerides and APOC3. Carriers of these mutations were found to have a reduced risk of coronary heart disease. (Funded by the National Heart, Lung, and Blood Institute and others.).

Loss-of-Function Mutations in APOC3, Triglycerides, and Coronary Disease

PubMed Central

2014-01-01

Background Plasma triglyceride levels are heritable and are correlated with the risk of coronary heart disease. Sequencing of the protein-coding regions of the human genome (the exome) has the potential to identify rare mutations that have a large effect on phenotype. Methods We sequenced the protein-coding regions of 18,666 genes in each of 3734 participants of European or African ancestry in the Exome Sequencing Project. We conducted tests to determine whether rare mutations in coding sequence, individually or in aggregate within a gene, were associated with plasma triglyceride levels. For mutations associated with triglyceride levels, we subsequently evaluated their association with the risk of coronary heart disease in 110,970 persons. Results An aggregate of rare mutations in the gene encoding apolipoprotein C3 (APOC3) was associated with lower plasma triglyceride levels. Among the four mutations that drove this result, three were loss-of-function mutations: a nonsense mutation (R19X) and two splice-site mutations (IVS2+1G→A and IVS3+1G→T). The fourth was a missense mutation (A43T). Approximately 1 in 150 persons in the study was a heterozygous carrier of at least one of these four mutations. Triglyceride levels in the carriers were 39% lower than levels in noncarriers (P<1×10−20), and circulating levels of APOC3 in carriers were 46% lower than levels in noncarriers (P = 8×10−10). The risk of coronary heart disease among 498 carriers of any rare APOC3 mutation was 40% lower than the risk among 110,472 noncarriers (odds ratio, 0.60; 95% confidence interval, 0.47 to 0.75; P = 4×10−6). Conclusions Rare mutations that disrupt APOC3 function were associated with lower levels of plasma triglycerides and APOC3. Carriers of these mutations were found to have a reduced risk of coronary heart disease. (Funded by the National Heart, Lung, and Blood Institute and others.) PMID:24941081

Secretion Trap Tagging of Secreted and Membrane-Spanning Proteins Using Arabidopsis Gene Traps

Treesearch

Andrew T. Groover; Joseph R. Fontana; Juana M. Arroyo; Cristina Yordan; W. Richard McCombie; Robert A. Martienssen

2003-01-01

Secreted and membrane-spanning proteins play fundamental roles in plant development but pose challenges for genetic identification and characterization. We describe a "secretion trap" screen for gene trap insertions in genes encoding proteins routed through the secretory pathway. The gene trap transposon encodes a ß-glucuronidase reporter enzyme...

A deep auto-encoder model for gene expression prediction.

PubMed

Xie, Rui; Wen, Jia; Quitadamo, Andrew; Cheng, Jianlin; Shi, Xinghua

2017-11-17

Gene expression is a key intermediate level that genotypes lead to a particular trait. Gene expression is affected by various factors including genotypes of genetic variants. With an aim of delineating the genetic impact on gene expression, we build a deep auto-encoder model to assess how good genetic variants will contribute to gene expression changes. This new deep learning model is a regression-based predictive model based on the MultiLayer Perceptron and Stacked Denoising Auto-encoder (MLP-SAE). The model is trained using a stacked denoising auto-encoder for feature selection and a multilayer perceptron framework for backpropagation. We further improve the model by introducing dropout to prevent overfitting and improve performance. To demonstrate the usage of this model, we apply MLP-SAE to a real genomic datasets with genotypes and gene expression profiles measured in yeast. Our results show that the MLP-SAE model with dropout outperforms other models including Lasso, Random Forests and the MLP-SAE model without dropout. Using the MLP-SAE model with dropout, we show that gene expression quantifications predicted by the model solely based on genotypes, align well with true gene expression patterns. We provide a deep auto-encoder model for predicting gene expression from SNP genotypes. This study demonstrates that deep learning is appropriate for tackling another genomic problem, i.e., building predictive models to understand genotypes' contribution to gene expression. With the emerging availability of richer genomic data, we anticipate that deep learning models play a bigger role in modeling and interpreting genomics.

Inhibition of miR-33a/b in non-human primates raises plasma HDL and lowers VLDL triglycerides.

PubMed

Rayner, Katey J; Esau, Christine C; Hussain, Farah N; McDaniel, Allison L; Marshall, Stephanie M; van Gils, Janine M; Ray, Tathagat D; Sheedy, Frederick J; Goedeke, Leigh; Liu, Xueqing; Khatsenko, Oleg G; Kaimal, Vivek; Lees, Cynthia J; Fernandez-Hernando, Carlos; Fisher, Edward A; Temel, Ryan E; Moore, Kathryn J

2011-10-19

Cardiovascular disease remains the leading cause of mortality in westernized countries, despite optimum medical therapy to reduce the levels of low-density lipoprotein (LDL)-associated cholesterol. The pursuit of novel therapies to target the residual risk has focused on raising the levels of high-density lipoprotein (HDL)-associated cholesterol in order to exploit its atheroprotective effects. MicroRNAs (miRNAs) have emerged as important post-transcriptional regulators of lipid metabolism and are thus a new class of target for therapeutic intervention. MicroRNA-33a and microRNA-33b (miR-33a/b) are intronic miRNAs whose encoding regions are embedded in the sterol-response-element-binding protein genes SREBF2 and SREBF1 (refs 3-5), respectively. These miRNAs repress expression of the cholesterol transporter ABCA1, which is a key regulator of HDL biogenesis. Recent studies in mice suggest that antagonizing miR-33a may be an effective strategy for raising plasma HDL levels and providing protection against atherosclerosis; however, extrapolating these findings to humans is complicated by the fact that mice lack miR-33b, which is present only in the SREBF1 gene of medium and large mammals. Here we show in African green monkeys that systemic delivery of an anti-miRNA oligonucleotide that targets both miR-33a and miR-33b increased hepatic expression of ABCA1 and induced a sustained increase in plasma HDL levels over 12 weeks. Notably, miR-33 antagonism in this non-human primate model also increased the expression of miR-33 target genes involved in fatty acid oxidation (CROT, CPT1A, HADHB and PRKAA1) and reduced the expression of genes involved in fatty acid synthesis (SREBF1, FASN, ACLY and ACACA), resulting in a marked suppression of the plasma levels of very-low-density lipoprotein (VLDL)-associated triglycerides, a finding that has not previously been observed in mice. These data establish, in a model that is highly relevant to humans, that pharmacological inhibition of miR-33a and miR-33b is a promising therapeutic strategy to raise plasma HDL and lower VLDL triglyceride levels for the treatment of dyslipidaemias that increase cardiovascular disease risk.

Molecular interactions between tomato and the leaf mold pathogen Cladosporium fulvum.

PubMed

Rivas, Susana; Thomas, Colwyn M

2005-01-01

The interaction between tomato and the leaf mold pathogen Cladosporium fulvum is controlled in a gene-for-gene manner. This interaction has provided useful insights to the molecular basis of recognition specificity in plant disease resistance (R) proteins, disease resistance (R) gene evolution, R-protein mediated signaling, and cellular responses to pathogen attack. Tomato Cf genes encode type I membrane-associated receptor-like proteins (RLPs) comprised predominantly of extracellular leucine-rich repeats (eLRRs) and which are anchored in the plasma membrane. Cf proteins recognize fungal avirulence (Avr) peptides secreted into the leaf apoplast during infection. A direct interaction of Cf proteins with their cognate Avr proteins has not been demonstrated and the molecular mechanism of Avr protein perception is not known. Following ligand perception Cf proteins trigger a hypersensitive response (HR) and the arrest of pathogen development. Cf proteins lack an obvious signaling domain, suggesting that defense response activation is mediated through interactions with other partners. Avr protein perception results in the rapid accumulation of active oxygen species (AOS), changes in cellular ion fluxes, activation of protein kinase cascades, changes in gene expression and, possibly, targeted protein degradation. Here we review our current understanding of Cf-mediated responses in resistance to C. fulvum.

Molecular evolution of nitrogen assimilatory enzymes in marine prasinophytes.

PubMed

Ghoshroy, Sohini; Robertson, Deborah L

2015-01-01

Nitrogen assimilation is a highly regulated process requiring metabolic coordination of enzymes and pathways in the cytosol, chloroplast, and mitochondria. Previous studies of prasinophyte genomes revealed that genes encoding nitrate and ammonium transporters have a complex evolutionary history involving both vertical and horizontal transmission. Here we examine the evolutionary history of well-conserved nitrogen-assimilating enzymes to determine if a similar complex history is observed. Phylogenetic analyses suggest that genes encoding glutamine synthetase (GS) III in the prasinophytes evolved by horizontal gene transfer from a member of the heterokonts. In contrast, genes encoding GSIIE, a canonical vascular plant and green algal enzyme, were found in the Micromonas genomes but have been lost from Ostreococcus. Phylogenetic analyses placed the Micromonas GSIIs in a larger chlorophyte/vascular plant clade; a similar topology was observed for ferredoxin-dependent nitrite reductase (Fd-NiR), indicating the genes encoding GSII and Fd-NiR in these prasinophytes evolved via vertical transmission. Our results show that genes encoding the nitrogen-assimilating enzymes in Micromonas and Ostreococcus have been differentially lost and as well as recruited from different evolutionary lineages, suggesting that the regulation of nitrogen assimilation in prasinophytes will differ from other green algae.

Living colors in the gray mold pathogen Botrytis cinerea: codon-optimized genes encoding green fluorescent protein and mCherry, which exhibit bright fluorescence.

PubMed

Leroch, Michaela; Mernke, Dennis; Koppenhoefer, Dieter; Schneider, Prisca; Mosbach, Andreas; Doehlemann, Gunther; Hahn, Matthias

2011-05-01

The green fluorescent protein (GFP) and its variants have been widely used in modern biology as reporters that allow a variety of live-cell imaging techniques. So far, GFP has rarely been used in the gray mold fungus Botrytis cinerea because of low fluorescence intensity. The codon usage of B. cinerea genes strongly deviates from that of commonly used GFP-encoding genes and reveals a lower GC content than other fungi. In this study, we report the development and use of a codon-optimized version of the B. cinerea enhanced GFP (eGFP)-encoding gene (Bcgfp) for improved expression in B. cinerea. Both the codon optimization and, to a smaller extent, the insertion of an intron resulted in higher mRNA levels and increased fluorescence. Bcgfp was used for localization of nuclei in germinating spores and for visualizing host penetration. We further demonstrate the use of promoter-Bcgfp fusions for quantitative evaluation of various toxic compounds as inducers of the atrB gene encoding an ABC-type drug efflux transporter of B. cinerea. In addition, a codon-optimized mCherry-encoding gene was constructed which yielded bright red fluorescence in B. cinerea.

Meta-omic signatures of microbial metal and nitrogen cycling in marine oxygen minimum zones

PubMed Central

Glass, Jennifer B.; Kretz, Cecilia B.; Ganesh, Sangita; Ranjan, Piyush; Seston, Sherry L.; Buck, Kristen N.; Landing, William M.; Morton, Peter L.; Moffett, James W.; Giovannoni, Stephen J.; Vergin, Kevin L.; Stewart, Frank J.

2015-01-01

Iron (Fe) and copper (Cu) are essential cofactors for microbial metalloenzymes, but little is known about the metalloenyzme inventory of anaerobic marine microbial communities despite their importance to the nitrogen cycle. We compared dissolved O2, NO3−, NO2−, Fe and Cu concentrations with nucleic acid sequences encoding Fe and Cu-binding proteins in 21 metagenomes and 9 metatranscriptomes from Eastern Tropical North and South Pacific oxygen minimum zones and 7 metagenomes from the Bermuda Atlantic Time-series Station. Dissolved Fe concentrations increased sharply at upper oxic-anoxic transition zones, with the highest Fe:Cu molar ratio (1.8) occurring at the anoxic core of the Eastern Tropical North Pacific oxygen minimum zone and matching the predicted maximum ratio based on data from diverse ocean sites. The relative abundance of genes encoding Fe-binding proteins was negatively correlated with O2, driven by significant increases in genes encoding Fe-proteins involved in dissimilatory nitrogen metabolisms under anoxia. Transcripts encoding cytochrome c oxidase, the Fe- and Cu-containing terminal reductase in aerobic respiration, were positively correlated with O2 content. A comparison of the taxonomy of genes encoding Fe- and Cu-binding vs. bulk proteins in OMZs revealed that Planctomycetes represented a higher percentage of Fe genes while Thaumarchaeota represented a higher percentage of Cu genes, particularly at oxyclines. These results are broadly consistent with higher relative abundance of genes encoding Fe-proteins in the genome of a marine planctomycete vs. higher relative abundance of genes encoding Cu-proteins in the genome of a marine thaumarchaeote. These findings highlight the importance of metalloenzymes for microbial processes in oxygen minimum zones and suggest preferential Cu use in oxic habitats with Cu > Fe vs. preferential Fe use in anoxic niches with Fe > Cu. PMID:26441925

Meta-omic signatures of microbial metal and nitrogen cycling in marine oxygen minimum zones.

PubMed

Glass, Jennifer B; Kretz, Cecilia B; Ganesh, Sangita; Ranjan, Piyush; Seston, Sherry L; Buck, Kristen N; Landing, William M; Morton, Peter L; Moffett, James W; Giovannoni, Stephen J; Vergin, Kevin L; Stewart, Frank J

2015-01-01

Iron (Fe) and copper (Cu) are essential cofactors for microbial metalloenzymes, but little is known about the metalloenyzme inventory of anaerobic marine microbial communities despite their importance to the nitrogen cycle. We compared dissolved O2, NO[Formula: see text], NO[Formula: see text], Fe and Cu concentrations with nucleic acid sequences encoding Fe and Cu-binding proteins in 21 metagenomes and 9 metatranscriptomes from Eastern Tropical North and South Pacific oxygen minimum zones and 7 metagenomes from the Bermuda Atlantic Time-series Station. Dissolved Fe concentrations increased sharply at upper oxic-anoxic transition zones, with the highest Fe:Cu molar ratio (1.8) occurring at the anoxic core of the Eastern Tropical North Pacific oxygen minimum zone and matching the predicted maximum ratio based on data from diverse ocean sites. The relative abundance of genes encoding Fe-binding proteins was negatively correlated with O2, driven by significant increases in genes encoding Fe-proteins involved in dissimilatory nitrogen metabolisms under anoxia. Transcripts encoding cytochrome c oxidase, the Fe- and Cu-containing terminal reductase in aerobic respiration, were positively correlated with O2 content. A comparison of the taxonomy of genes encoding Fe- and Cu-binding vs. bulk proteins in OMZs revealed that Planctomycetes represented a higher percentage of Fe genes while Thaumarchaeota represented a higher percentage of Cu genes, particularly at oxyclines. These results are broadly consistent with higher relative abundance of genes encoding Fe-proteins in the genome of a marine planctomycete vs. higher relative abundance of genes encoding Cu-proteins in the genome of a marine thaumarchaeote. These findings highlight the importance of metalloenzymes for microbial processes in oxygen minimum zones and suggest preferential Cu use in oxic habitats with Cu > Fe vs. preferential Fe use in anoxic niches with Fe > Cu.

The Yersinia pestis gcvB gene encodes two small regulatory RNA molecules

PubMed Central

McArthur, Sarah D; Pulvermacher, Sarah C; Stauffer, George V

2006-01-01

Background In recent years it has become clear that small non-coding RNAs function as regulatory elements in bacterial virulence and bacterial stress responses. We tested for the presence of the small non-coding GcvB RNAs in Y. pestis as possible regulators of gene expression in this organism. Results In this study, we report that the Yersinia pestis KIM6 gcvB gene encodes two small RNAs. Transcription of gcvB is activated by the GcvA protein and repressed by the GcvR protein. The gcvB-encoded RNAs are required for repression of the Y. pestis dppA gene, encoding the periplasmic-binding protein component of the dipeptide transport system, showing that the GcvB RNAs have regulatory activity. A deletion of the gcvB gene from the Y. pestis KIM6 chromosome results in a decrease in the generation time of the organism as well as a change in colony morphology. Conclusion The results of this study indicate that the Y. pestis gcvB gene encodes two small non-coding regulatory RNAs that repress dppA expression. A gcvB deletion is pleiotropic, suggesting that the sRNAs are likely involved in controlling genes in addition to dppA. PMID:16768793

Overexpression of GhSARP1 encoding a E3 ligase from cotton reduce the tolerance to salt in transgenic Arabidopsis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Yongchang; Zhang, Xinyu; Zhu, Shouhong

Ubiquitination plays a very important role in the response to abiotic stresses of plant. To identify key regulators of salt stress, a gene GhSARP1(Salt-Associated Ring finger Protein)encoding C3H2C3-type E3 ligase, was cloned from cotton. Transcription level of GhSARP1 was high in leaf, flower and fiber of 24,27 and 27DPA (Days Post-Anthesis), but low in root and stem. Except PEG6000 treatment, the expression of GhSARP1 was down-regulated by NaCl, cold and ABA after being treated for 1 h. GhSARP1-GFP fusion protein located on the plasma membrane, which was dependent on trans-membrane motif. In vitro ubiquitination assay showed that GhSARP1 had E3 ligase activity.more » Heterogeneous overexpression of GhSARP1reduced salt tolerance of transgenic Arabidopsis in germination and post-germination stage. Our results suggested that the GhSARP1 might negatively regulate the response to salt stress mediated by the ubiquitination in cotton. - Highlights: • GhSARP1 expression was regulated by various abiotic stresses. • GhSARP1 have E3 ligase activity in vitro and locate on the plasma membrane. • Overexpression of GhSARP1 in Arabidopsis reduced the salt tolerance.« less

Screening for ATM Mutations in an African-American Population to Identify a Predictor of Breast Cancer Susceptibility

DTIC Science & Technology

2006-07-01

ATM genetic variant identified affects radiosensitivity and levels of the protein encoded by the ATM gene for each mutation examined. 15. SUBJECT...women without breast cancer. An additional objective is to determine the functional impact upon the protein encoded by the ATM gene for each mutation ...each ATM variant identified affects radiosensitivity and levels of the protein encoded by the ATM gene for mutations identified. Body STATEMENT

Isolation of a gene encoding a novel spectinomycin phosphotransferase from Legionella pneumophila.

PubMed

Suter, T M; Viswanathan, V K; Cianciotto, N P

1997-06-01

A gene capable of conferring spectinomycin resistance was isolated from Legionella pneumophila, the agent of Legionnaires' disease. The gene (aph) encoded a 36-kDa protein which has similarity to aminoglycoside phosphotransferases. Biochemical analysis confirmed that aph encodes a phosphotransferase which modifies spectinomycin but not hygromycin, kanamycin, or streptomycin. The strain that was the source of aph demonstrated resistance to spectinomycin, and Southern hybridizations determined that aph also exists in other legionellae.

Isolation of a gene encoding a novel spectinomycin phosphotransferase from Legionella pneumophila.

PubMed Central

Suter, T M; Viswanathan, V K; Cianciotto, N P

1997-01-01

A gene capable of conferring spectinomycin resistance was isolated from Legionella pneumophila, the agent of Legionnaires' disease. The gene (aph) encoded a 36-kDa protein which has similarity to aminoglycoside phosphotransferases. Biochemical analysis confirmed that aph encodes a phosphotransferase which modifies spectinomycin but not hygromycin, kanamycin, or streptomycin. The strain that was the source of aph demonstrated resistance to spectinomycin, and Southern hybridizations determined that aph also exists in other legionellae. PMID:9174205

Escherichia coli yjjPB genes encode a succinate transporter important for succinate production.

PubMed

Fukui, Keita; Nanatani, Kei; Hara, Yoshihiko; Yamakami, Suguru; Yahagi, Daiki; Chinen, Akito; Tokura, Mitsunori; Abe, Keietsu

2017-09-01

Under anaerobic conditions, Escherichia coli produces succinate from glucose via the reductive tricarboxylic acid cycle. To date, however, no genes encoding succinate exporters have been established in E. coli. Therefore, we attempted to identify genes encoding succinate exporters by screening an E. coli MG1655 genome library. We identified the yjjPB genes as candidates encoding a succinate transporter, which enhanced succinate production in Pantoea ananatis under aerobic conditions. A complementation assay conducted in Corynebacterium glutamicum strain AJ110655ΔsucE1 demonstrated that both YjjP and YjjB are required for the restoration of succinate production. Furthermore, deletion of yjjPB decreased succinate production in E. coli by 70% under anaerobic conditions. Taken together, these results suggest that YjjPB constitutes a succinate transporter in E. coli and that the products of both genes are required for succinate export.

Neurological toxicity after phenytoin infusion in a pediatric patient with epilepsy: influence of CYP2C9, CYP2C19 and ABCB1 genetic polymorphisms.

PubMed

Dorado, P; López-Torres, E; Peñas-Lledó, E M; Martínez-Antón, J; Llerena, A

2013-08-01

Pharmacogenetic studies have shown that genetic defects in drug-metabolizing enzymes encoded by CYP2C9, CYP2C19 genes and by the transporter ABCB1 gene can influence phenytoin (PTH) plasma levels and toxicity. The patient reported here is a 2-year-old girl with a medical history of cryptogenic (probably symptomatic) epilepsy, who had her first focal seizure with secondary generalization at 13 months of age. She initially received oral valproate treatment and three months later, she was prescribed an oral oxcarbazepine treatment. At 20 months of age, she was admitted to the Emergency Department because of generalized convulsive Status Epilepticus needing to be immediately treated with rectal diazepam (0.5 mg kg(-1)), intravenous diazepam (0.3 mg kg(-1)), and intravenous phenytoin with an initial-loading dose of 15 mg kg(-1). However, two hours after the initial-loading dose of PTH, the patient developed dizziness, nystagmus, ataxia and excessive sedation. Other potential causes of PTH toxicity were excluded such as drug interactions, decreased albumin or lab error. Therefore, to explain the neurological toxicity, PTH plasma levels and CYP2C9, CYP2C19 and ABCB1 genetic polymorphisms were analyzed. Initial plasma PTH levels were higher than expected (69 mg l(-1); normal range: 10-20 mg l(-1)), and the patient was homozygous for the CYP2C9*2 allele, heterozygous for the CYP2C19*4 allele and homozygous for the 3435C and 1236C ABCB1 alleles. Present findings support the previously established relationship between CYP2C9 and CYP2C19 genetic polymorphisms and the increased risk to develop PTH toxicity owing to high plasma concentrations. Nevertheless, although the association of these genes with PTH-induced adverse effects has been well-documented in adult populations, this is the first report examining the influence of these genetic polymorphisms on PTH plasma levels and toxicity in a pediatric patient.

Molecular cloning and expression of the gene encoding the kinetoplast-associated type II DNA topoisomerase of Crithidia fasciculata.

PubMed

Pasion, S G; Hines, J C; Aebersold, R; Ray, D S

1992-01-01

A type II DNA topoisomerase, topoIImt, was shown previously to be associated with the kinetoplast DNA of the trypanosomatid Crithidia fasciculata. The gene encoding this kinetoplast-associated topoisomerase has been cloned by immunological screening of a Crithidia genomic expression library with monoclonal antibodies raised against the purified enzyme. The gene CfaTOP2 is a single copy gene and is expressed as a 4.8-kb polyadenylated transcript. The nucleotide sequence of CfaTOP2 has been determined and encodes a predicted polypeptide of 1239 amino acids with a molecular mass of 138,445. The identification of the cloned gene is supported by immunoblot analysis of the beta-galactosidase-CfaTOP2 fusion protein expressed in Escherichia coli and by analysis of tryptic peptide sequences derived from purified topoIImt. CfaTOP2 shares significant homology with nuclear type II DNA topoisomerases of other eukaryotes suggesting that in Crithidia both nuclear and mitochondrial forms of topoisomerase II are encoded by the same gene.

Two pheromone precursor genes are transcriptionally expressed in the homothallic ascomycete Sordaria macrospora.

PubMed

Pöggeler, S

2000-06-01

In order to analyze the involvement of pheromones in cell recognition and mating in a homothallic fungus, two putative pheromone precursor genes, named ppg1 and ppg2, were isolated from a genomic library of Sordaria macrospora. The ppg1 gene is predicted to encode a precursor pheromone that is processed by a Kex2-like protease to yield a pheromone that is structurally similar to the alpha-factor of the yeast Saccharomyces cerevisiae. The ppg2 gene encodes a 24-amino-acid polypeptide that contains a putative farnesylated and carboxy methylated C-terminal cysteine residue. The sequences of the predicted pheromones display strong structural similarity to those encoded by putative pheromones of heterothallic filamentous ascomycetes. Both genes are expressed during the life cycle of S. macrospora. This is the first description of pheromone precursor genes encoded by a homothallic fungus. Southern-hybridization experiments indicated that ppg1 and ppg2 homologues are also present in other homothallic ascomycetes.

Arxula adeninivorans (Blastobotrys adeninivorans) — A Dimorphic Yeast of Great Biotechnological Potential

NASA Astrophysics Data System (ADS)

Böer, Erik; Steinborn, Gerhard; Florschütz, Kristina; Körner, Martina; Gellissen, Gerd; Kunze, Gotthard

The dimorphic ascomycetous yeast Arxula adeninivorans exhibits some unusual properties. Being a thermo- and halotolerant species it is able to assimilate and ferment many compounds as sole carbon and/or nitrogen source. It utilises n-alkanes and is capable of degrading starch. Due to these unusual biochemical properties A. adeninivorans can be exploited as a gene donor for the production of enzymes with attractive biotechnological characteristics. Examples of A. adeninivorans-derived genes that are overexpressed include the ALIP1 gene encoding a secretory lipase, the AINV encoding invertase, the AXDH encoding xylitol dehydrogenase and the APHY encoding a secretory phosphatase with phytase activity.

Structure of the Elastin-Contractile Units in the Thoracic Aorta and How Genes That Cause Thoracic Aortic Aneurysms and Dissections Disrupt This Structure.

PubMed

Karimi, Ashkan; Milewicz, Dianna M

2016-01-01

The medial layer of the aorta confers elasticity and strength to the aortic wall and is composed of alternating layers of smooth muscle cells (SMCs) and elastic fibres. The SMC elastin-contractile unit is a structural unit that links the elastin fibres to the SMCs and is characterized by the following: (1) layers of elastin fibres that are surrounded by microfibrils; (2) microfibrils that bind to the integrin receptors in focal adhesions on the cell surface of the SMCs; and (3) SMC contractile filaments that are linked to the focal adhesions on the inner side of the membrane. The genes that are altered to cause thoracic aortic aneurysms and aortic dissections encode proteins involved in the structure or function of the SMC elastin-contractile unit. Included in this gene list are the genes encoding protein that are structural components of elastin fibres and microfibrils, FBN1, MFAP5, ELN, and FBLN4. Also included are genes that encode structural proteins in the SMC contractile unit, including ACTA2, which encodes SMC-specific α-actin and MYH11, which encodes SMC-specific myosin heavy chain, along with MYLK and PRKG1, which encode kinases that control SMC contraction. Finally, mutations in the gene encoding the protein linking integrin receptors to the contractile filaments, FLNA, also predispose to thoracic aortic disease. Thus, these data suggest that functional SMC elastin-contractile units are important for maintaining the structural integrity of the aorta. Copyright © 2016 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.

The Brown-Vialetto-Van Laere and Fazio Londe syndrome revisited: natural history, genetics, treatment and future perspectives.

PubMed

Bosch, Annet M; Stroek, Kevin; Abeling, Nico G; Waterham, Hans R; Ijlst, Lodewijk; Wanders, Ronald J A

2012-10-29

The Brown-Vialetto-Van Laere syndrome is a rare neurological disorder which may present at all ages with sensorineural deafness, bulbar palsy and respiratory compromise. Fazio-Londe syndrome is considered to be the same disease entity. Recently it was demonstrated that in some patients the disease is caused by mutations in the SLC52A3 gene which encodes the intestinal (hRFT2) riboflavin transporter. In these patients riboflavin deficiency is the cause of the BVVL/FL syndrome and supplementation of riboflavin proved a life saving treatment. Mutations in the SLC52A2 gene and the SLC52A1 (GPR172B) gene, coding for human riboflavin transporters hRFT3 and hRFT1 have been associated with the BVVL syndrome as well. We performed a review of the literature, with emphasis on the natural history and the effects of treatment in these patients. A total of 35 publications were traced reporting on the clinical presentation of 74 patients who presented before age 18. The most prevalent symptoms were bulbar palsy, hearing loss, facial weakness and respiratory compromise. Death was reported in 28 of the 61 untreated patients, with a very low survival in patients presenting before age 4. All 13 patients who were treated with riboflavin survived, with a strong clinical improvement after days to months of treatment in eight patients. Three patients demonstrated a stable clinical course and treatment was stopped early in two patients. Abnormalities in plasma flavin levels and/or plasma acylcarnitine profiles were observed in some but not in all patients, and also patients with normal plasma flavin levels and acylcarnitine profiles demonstrated a striking clinical improvement on riboflavin supplementation. It is now clear that proper diagnosis requires mutation analysis of all three transporter genes and treatment should be started immediately without first awaiting results of molecular analysis. Clinical improvement may be rapid or gradual over a period of more than 12 months.

The Brown-Vialetto-Van Laere and Fazio Londe syndrome revisited: natural history, genetics, treatment and future perspectives

PubMed Central

2012-01-01

The Brown-Vialetto-Van Laere syndrome is a rare neurological disorder which may present at all ages with sensorineural deafness, bulbar palsy and respiratory compromise. Fazio-Londe syndrome is considered to be the same disease entity. Recently it was demonstrated that in some patients the disease is caused by mutations in the SLC52A3 gene which encodes the intestinal (hRFT2) riboflavin transporter. In these patients riboflavin deficiency is the cause of the BVVL/FL syndrome and supplementation of riboflavin proved a life saving treatment. Mutations in the SLC52A2 gene and the SLC52A1 (GPR172B) gene, coding for human riboflavin transporters hRFT3 and hRFT1 have been associated with the BVVL syndrome as well. We performed a review of the literature, with emphasis on the natural history and the effects of treatment in these patients. A total of 35 publications were traced reporting on the clinical presentation of 74 patients who presented before age 18. The most prevalent symptoms were bulbar palsy, hearing loss, facial weakness and respiratory compromise. Death was reported in 28 of the 61 untreated patients, with a very low survival in patients presenting before age 4. All 13 patients who were treated with riboflavin survived, with a strong clinical improvement after days to months of treatment in eight patients. Three patients demonstrated a stable clinical course and treatment was stopped early in two patients. Abnormalities in plasma flavin levels and/or plasma acylcarnitine profiles were observed in some but not in all patients, and also patients with normal plasma flavin levels and acylcarnitine profiles demonstrated a striking clinical improvement on riboflavin supplementation. It is now clear that proper diagnosis requires mutation analysis of all three transporter genes and treatment should be started immediately without first awaiting results of molecular analysis. Clinical improvement may be rapid or gradual over a period of more than 12 months. PMID:23107375

Comparative molecular biological analysis of membrane transport genes in organisms

PubMed Central

Nagata, Toshifumi; Iizumi, Shigemi; Satoh, Kouji

2008-01-01

Comparative analyses of membrane transport genes revealed many differences in the features of transport homeostasis in eight diverse organisms, ranging from bacteria to animals and plants. In bacteria, membrane-transport systems depend mainly on single genes encoding proteins involved in an ATP-dependent pump and secondary transport proteins that use H+ as a co-transport molecule. Animals are especially divergent in their channel genes, and plants have larger numbers of P-type ATPase and secondary active transporters than do other organisms. The secondary transporter genes have diverged evolutionarily in both animals and plants for different co-transporter molecules. Animals use Na+ ions for the formation of concentration gradients across plasma membranes, dependent on secondary active transporters and on membrane voltages that in turn are dependent on ion transport regulation systems. Plants use H+ ions pooled in vacuoles and the apoplast to transport various substances; these proton gradients are also dependent on secondary active transporters. We also compared the numbers of membrane transporter genes in Arabidopsis and rice. Although many transporter genes are similar in these plants, Arabidopsis has a more diverse array of genes for multi-efflux transport and for response to stress signals, and rice has more secondary transporter genes for carbohydrate and nutrient transport. Electronic supplementary material The online version of this article (doi:10.1007/s11103-007-9287-z) contains supplementary material, which is available to authorized users. PMID:18293089

V-ATPase-dependent luminal acidification is required for endocytic recycling of a yeast cell wall stress sensor, Wsc1p

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ueno, Kazuma; Saito, Mayu; Nagashima, Makiko

Highlights: •A targeted genome screen identified 5 gene groups affecting Wsc1p recycling. •V-ATPase-dependent luminal acidification is required for Wsc1p recycling. •Activity of V-ATPase might be required for cargo recognition by the retromer complex. -- Abstract: Wsc1p is a major cell wall sensor protein localized at the polarized cell surface. The localization of Wsc1p is maintained by endocytosis and recycling from endosomes back to the cell surface, but changes to the vacuole when cells are subjected to heat stress. Exploiting this unique property of Wsc1p, we screened for yeast single-gene deletion mutants exhibiting defects in Wsc1p trafficking. By expressing 3GFP-tagged Wsc1pmore » in mutants with deleted genes whose function is related to intracellular trafficking, we identified 5 gene groups affecting Wsc1p trafficking, impaired respectively in endocytic internalization, multivesicular body sorting, the GARP complex, endosomal maturation/vacuolar fusion, and V-ATPase. Interestingly, deletion of the VPH1 gene, encoding the V{sub o} subunit of vacuolar-type H{sup +}-ATPase (V-ATPase), led to mis-localization of Wsc1p from the plasma membrane to the vacuole. In addition, disruption of other V-ATPase subunits (vma mutants) also caused defects of Wsc1p trafficking and vacuolar acidification similar to those seen in the vph1Δ mutant. Moreover, we found that deletion of the VPS26 gene, encoding a subunit of the retromer complex, also caused a defect in Wsc1p recycling and mis-localization of Wsc1p to the vacuole. These findings clarified the previously unidentified Wsc1p recycling pathway and requirement of V-ATPase-dependent luminal acidification for Wsc1p recycling.« less

Medicago truncatula contains a second gene encoding a plastid located glutamine synthetase exclusively expressed in developing seeds.

PubMed

Seabra, Ana R; Vieira, Cristina P; Cullimore, Julie V; Carvalho, Helena G

2010-08-19

Nitrogen is a crucial nutrient that is both essential and rate limiting for plant growth and seed production. Glutamine synthetase (GS), occupies a central position in nitrogen assimilation and recycling, justifying the extensive number of studies that have been dedicated to this enzyme from several plant sources. All plants species studied to date have been reported as containing a single, nuclear gene encoding a plastid located GS isoenzyme per haploid genome. This study reports the existence of a second nuclear gene encoding a plastid located GS in Medicago truncatula. This study characterizes a new, second gene encoding a plastid located glutamine synthetase (GS2) in M. truncatula. The gene encodes a functional GS isoenzyme with unique kinetic properties, which is exclusively expressed in developing seeds. Based on molecular data and the assumption of a molecular clock, it is estimated that the gene arose from a duplication event that occurred about 10 My ago, after legume speciation and that duplicated sequences are also present in closely related species of the Vicioide subclade. Expression analysis by RT-PCR and western blot indicate that the gene is exclusively expressed in developing seeds and its expression is related to seed filling, suggesting a specific function of the enzyme associated to legume seed metabolism. Interestingly, the gene was found to be subjected to alternative splicing over the first intron, leading to the formation of two transcripts with similar open reading frames but varying 5' UTR lengths, due to retention of the first intron. To our knowledge, this is the first report of alternative splicing on a plant GS gene. This study shows that Medicago truncatula contains an additional GS gene encoding a plastid located isoenzyme, which is functional and exclusively expressed during seed development. Legumes produce protein-rich seeds requiring high amounts of nitrogen, we postulate that this gene duplication represents a functional innovation of plastid located GS related to storage protein accumulation exclusive to legume seed metabolism.

Molecular cloning and expression of heteromeric ACCase subunit genes from Jatropha curcas.

PubMed

Gu, Keyu; Chiam, Huihui; Tian, Dongsheng; Yin, Zhongchao

2011-04-01

Acetyl-CoA carboxylase (ACCase) catalyzes the biotin-dependent carboxylation of acetyl-CoA to produce malonyl-CoA, which is the essential first step in the biosynthesis of long-chain fatty acids. ACCase exists as a multi-subunit enzyme in most prokaryotes and the chloroplasts of most plants and algae, while it is present as a multi-domain enzyme in the endoplasmic reticulum of most eukaryotes. The heteromeric ACCase of higher plants consists of four subunits: an α-subunit of carboxyltransferase (α-CT, encoded by accA gene), a biotin carboxyl carrier protein (BCCP, encoded by accB gene), a biotin carboxylase (BC, encoded by accC gene) and a β-subunit of carboxyltransferase (β-CT, encoded by accD gene). In this study, we cloned and characterized the genes accA, accB1, accC and accD that encode the subunits of heteromeric ACCase in Jatropha (Jatropha curcas), a potential biofuel plant. The full-length cDNAs of the four subunit genes were isolated from a Jatropha cDNA library and by using 5' RACE, whereas the genomic clones were obtained from a Jatropha BAC library. They encode a 771 amino acid (aa) α-CT, a 286-aa BCCP1, a 537-aa BC and a 494-aa β-CT, respectively. The single-copy accA, accB1 and accC genes are nuclear genes, while the accD gene is located in chloroplast genome. Jatropha α-CT, BCCP1, BC and β-CT show high identity to their homologues in other higher plants at amino acid level and contain all conserved domains for ACCase activity. The accA, accB1, accC and accD genes are temporally and spatially expressed in the leaves and endosperm of Jatropha plants, which are regulated by plant development and environmental factors. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Gene Cluster Encoding Cholate Catabolism in Rhodococcus spp.

PubMed Central

Wilbrink, Maarten H.; Casabon, Israël; Stewart, Gordon R.; Liu, Jie; van der Geize, Robert; Eltis, Lindsay D.

2012-01-01

Bile acids are highly abundant steroids with important functions in vertebrate digestion. Their catabolism by bacteria is an important component of the carbon cycle, contributes to gut ecology, and has potential commercial applications. We found that Rhodococcus jostii RHA1 grows well on cholate, as well as on its conjugates, taurocholate and glycocholate. The transcriptome of RHA1 growing on cholate revealed 39 genes upregulated on cholate, occurring in a single gene cluster. Reverse transcriptase quantitative PCR confirmed that selected genes in the cluster were upregulated 10-fold on cholate versus on cholesterol. One of these genes, kshA3, encoding a putative 3-ketosteroid-9α-hydroxylase, was deleted and found essential for growth on cholate. Two coenzyme A (CoA) synthetases encoded in the cluster, CasG and CasI, were heterologously expressed. CasG was shown to transform cholate to cholyl-CoA, thus initiating side chain degradation. CasI was shown to form CoA derivatives of steroids with isopropanoyl side chains, likely occurring as degradation intermediates. Orthologous gene clusters were identified in all available Rhodococcus genomes, as well as that of Thermomonospora curvata. Moreover, Rhodococcus equi 103S, Rhodococcus ruber Chol-4 and Rhodococcus erythropolis SQ1 each grew on cholate. In contrast, several mycolic acid bacteria lacking the gene cluster were unable to grow on cholate. Our results demonstrate that the above-mentioned gene cluster encodes cholate catabolism and is distinct from a more widely occurring gene cluster encoding cholesterol catabolism. PMID:23024343

Identification of a novel gene cluster in the upstream region of the S-layer gene sbpA involved in cell wall metabolism of Lysinibacillus sphaericus CCM 2177 and characterization of the recombinantly produced autolysin and pyruvyl transferase.

PubMed

Pleschberger, Magdalena; Hildner, Florian; Rünzler, Dominik; Gelbmann, Nicola; Mayer, Harald F; Sleytr, Uwe B; Egelseer, Eva M

2013-05-01

The S-layer protein SbpA of Lysinibacillus sphaericus CCM 2177 assembles into a square (p4) lattice structure and recognizes a pyruvylated secondary cell wall polymer (SCWP) as the proper anchoring structure to the rigid cell wall layer. Sequencing of 8,004 bp in the 5'-upstream region of the S-layer gene sbpA led to five ORFs-encoding proteins involved in cell wall metabolism. After cloning and heterologous expression of ORF1 and ORF5 in Escherichia coli, the recombinant autolysin rAbpA and the recombinant pyruvyl transferase rCsaB were isolated, purified, and correct folding was confirmed by circular dichroism. Although rAbpA encoded by ORF1 showed amidase activity, it could attack whole cells of Ly. sphaericus CCM 2177 only after complete extraction of the S-layer lattice. Despite the presence of three S-layer-homology motifs on the N-terminal part, rAbpA did not show detectable affinity to peptidoglycan-containing sacculi, nor to isolated SCWP. As the molecular mass of the autolysin lies above the molecular exclusion limit of the S-layer, AbpA is obviously trapped within the rigid cell wall layer by the isoporous protein lattice. Immunogold-labeling of ultrathin-sectioned whole cells of Ly. sphaericus CCM 2177 with a polyclonal rabbit antiserum raised against rCsaB encoded by ORF5, and cell fractionation experiments demonstrated that the pyruvyl transferase was located in the cytoplasm, but not associated with cell envelope components including the plasma membrane. In enzymatic assays, rCsaB clearly showed pyruvyl transferase activity. By using RT-PCR, specific transcripts for each ORF could be detected. Cotranscription could be confirmed for ORF2 and ORF3.

The prrF-Encoded Small Regulatory RNAs Are Required for Iron Homeostasis and Virulence of Pseudomonas aeruginosa

PubMed Central

Reinhart, Alexandria A.; Powell, Daniel A.; Nguyen, Angela T.; O'Neill, Maura; Djapgne, Louise; Wilks, Angela; Ernst, Robert K.

2014-01-01

Pseudomonas aeruginosa is an opportunistic pathogen that requires iron to cause infection, but it also must regulate the uptake of iron to avoid iron toxicity. The iron-responsive PrrF1 and PrrF2 small regulatory RNAs (sRNAs) are part of P. aeruginosa's iron regulatory network and affect the expression of at least 50 genes encoding iron-containing proteins. The genes encoding the PrrF1 and PrrF2 sRNAs are encoded in tandem in P. aeruginosa, allowing for the expression of a distinct, heme-responsive sRNA named PrrH that appears to regulate genes involved in heme metabolism. Using a combination of growth, mass spectrometry, and gene expression analysis, we showed that the ΔprrF1,2 mutant, which lacks expression of the PrrF and PrrH sRNAs, is defective for both iron and heme homeostasis. We also identified phuS, encoding a heme binding protein involved in heme acquisition, and vreR, encoding a previously identified regulator of P. aeruginosa virulence genes, as novel targets of prrF-mediated heme regulation. Finally, we showed that the prrF locus encoding the PrrF and PrrH sRNAs is required for P. aeruginosa virulence in a murine model of acute lung infection. Moreover, we showed that inoculation with a ΔprrF1,2 deletion mutant protects against future challenge with wild-type P. aeruginosa. Combined, these data demonstrate that the prrF-encoded sRNAs are critical regulators of P. aeruginosa virulence. PMID:25510881

Wide Tolerance to Amino Acids Substitutions In The OCTN1 Ergothioneine Transporter

PubMed Central

Frigeni, Marta; Iacobazzi, Francesco; Yin, Xue; Longo, Nicola

2016-01-01

Background Organic cation transporters transfer solutes with a positive charge across the plasma membrane. The novel organic cation transporter 1 (OCTN1) and 2 (OCTN2) transport ergothioneine and carnitine, respectively. Mutations in the SLC22A5 gene encoding OCTN2 cause primary carnitine deficiency, a recessive disorders resulting in low carnitine levels and defective fatty acid oxidation. Variations in the SLC22A4 gene encoding OCTN1 are associated with rheumatoid arthritis and Crohn disease. Methods Here we evaluate the functional properties of the OCTN1 transporter using chimeric transporters constructed by fusing different portion of the OCTN1 and OCTN2 cDNAs. Their relative abundance and subcellular distribution was evaluated through western blot analysis and confocal microscopy. Results Substitutions of the C-terminal portion of OCTN1 with the correspondent residues of OCTN2 generated chimeric OCTN transporters more active than wild-type OCTN1 in transporting ergothioneine. Additional single amino acid substitutions introduced in chimeric OCTN transporters further increased ergothioneine transport activity. Kinetic analysis indicated that increased transport activity was due to an increased Vmax, with modest changes in Km toward ergothioneine. Conclusions Our results indicate that the OCTN1 transporter is tolerant to extensive amino acid substitutions. This is in sharp contrast to the OCTN2 carnitine transporter that has been selected for high functional activity through evolution, with almost all substitutions reducing carnitine transport activity. General significance The widespread tolerance of OCTN1 to amino acid substitutions suggests that the corresponding SLC22A4 gene may have derived from a recent duplication of the SLC22A5 gene and might not yet have a defined physiological role. PMID:26994919

Non-hepatic tumors change the activity of genes encoding copper trafficking proteins in the liver

PubMed Central

Babich, Polina S.; Skvortsov, Alexey N; Rusconi, Paolo; Tsymbalenko, Nadezhda V.; Mutanen, Marja; Puchkova, Ludmila V.; Broggini, Massimo

2013-01-01

To assess the statistical relationship between tumor growth and copper metabolism, we performed a metaanalysis of studies in which patients with neoplasms were characterized according to any of the copper status indexes (atomic copper serum concentration, serum oxidase activity, ceruloplasmin protein content). Our metaanalysis shows that in the majority of cases (more than 3100 patients), tumor growth positively correlates with the copper status indexes. Nude athymic CD-1 nu/nu mice with subcutaneous tumors of human origin, C57Bl/6J mice with murine melanoma and ApcMin mice with spontaneously developing adenomas throughout the intestinal tract were studied to experimentally determine the relationship between tumor progression, liver copper metabolism, and copper status indexes. We showed that the copper status indexes increased significantly during tumor growth. In the liver tissue of tumor-bearing mice, ceruloplasmin gene expression, as well as the expression of genes related to ceruloplasmin metallation (CTR1 and ATP7B), increased significantly. Moreover, the presence of an mRNA splice variant encoding a form of ceruloplasmin anchored to the plasma membrane by glycosylphosphatidyl inositol, which is atypical for hepatocytes, was also detected. The ATP7A copper transporter gene, which is normally expressed in the liver only during embryonic copper metabolism, was also activated. Depletion of holo-ceruloplasmin resulted in retardation of human HCT116 colon carcinoma cell growth in nude mice and induced DNA fragmentation in tumor cells. In addition, the concentration of cytochrome c increased significantly in the cytosol, while decreasing in the mitochondria. We discuss a possible trans-effect of developing tumors on copper metabolism in the liver. PMID:23792645

CIP1 polypeptides and their uses

DOEpatents

Foreman, Pamela [Los Altos, CA; Van Solingen, Pieter [Naaldwijk, NL; Goedegebuur, Frits [Vlaardingen, NL; Ward, Michael [San Francisco, CA

2011-04-12

Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

Paralogous ALT1 and ALT2 Retention and Diversification Have Generated Catalytically Active and Inactive Aminotransferases in Saccharomyces cerevisiae

PubMed Central

Peñalosa-Ruiz, Georgina; Aranda, Cristina; Ongay-Larios, Laura; Colon, Maritrini; Quezada, Hector; Gonzalez, Alicia

2012-01-01

Background Gene duplication and the subsequent divergence of paralogous pairs play a central role in the evolution of novel gene functions. S. cerevisiae possesses two paralogous genes (ALT1/ALT2) which presumably encode alanine aminotransferases. It has been previously shown that Alt1 encodes an alanine aminotransferase, involved in alanine metabolism; however the physiological role of Alt2 is not known. Here we investigate whether ALT2 encodes an active alanine aminotransferase. Principal Findings Our results show that although ALT1 and ALT2 encode 65% identical proteins, only Alt1 displays alanine aminotransferase activity; in contrast ALT2 encodes a catalytically inert protein. ALT1 and ALT2 expression is modulated by Nrg1 and by the intracellular alanine pool. ALT1 is alanine-induced showing a regulatory profile of a gene encoding an enzyme involved in amino acid catabolism, in agreement with the fact that Alt1 is the sole pathway for alanine catabolism present in S. cerevisiae. Conversely, ALT2 expression is alanine-repressed, indicating a role in alanine biosynthesis, although the encoded-protein has no alanine aminotransferase enzymatic activity. In the ancestral-like yeast L. kluyveri, the alanine aminotransferase activity was higher in the presence of alanine than in the presence of ammonium, suggesting that as for ALT1, LkALT1 expression could be alanine-induced. ALT2 retention poses the questions of whether the encoded protein plays a particular function, and if this function was present in the ancestral gene. It could be hypotesized that ALT2 diverged after duplication, through neo-functionalization or that ALT2 function was present in the ancestral gene, with a yet undiscovered function. Conclusions ALT1 and ALT2 divergence has resulted in delegation of alanine aminotransferase activity to Alt1. These genes display opposed regulatory profiles: ALT1 is alanine-induced, while ALT2 is alanine repressed. Both genes are negatively regulated by the Nrg1 repressor. Presented results indicate that alanine could act as ALT2 Nrg1-co-repressor. PMID:23049841

Relating genes to function: identifying enriched transcription factors using the ENCODE ChIP-Seq significance tool.

PubMed

Auerbach, Raymond K; Chen, Bin; Butte, Atul J

2013-08-01

Biological analysis has shifted from identifying genes and transcripts to mapping these genes and transcripts to biological functions. The ENCODE Project has generated hundreds of ChIP-Seq experiments spanning multiple transcription factors and cell lines for public use, but tools for a biomedical scientist to analyze these data are either non-existent or tailored to narrow biological questions. We present the ENCODE ChIP-Seq Significance Tool, a flexible web application leveraging public ENCODE data to identify enriched transcription factors in a gene or transcript list for comparative analyses. The ENCODE ChIP-Seq Significance Tool is written in JavaScript on the client side and has been tested on Google Chrome, Apple Safari and Mozilla Firefox browsers. Server-side scripts are written in PHP and leverage R and a MySQL database. The tool is available at http://encodeqt.stanford.edu. abutte@stanford.edu Supplementary material is available at Bioinformatics online.

Rudimentary expression of RYamide in Drosophila melanogaster relative to other Drosophila species points to a functional decline of this neuropeptide gene.

PubMed

Veenstra, Jan A; Khammassi, Hela

2017-04-01

RYamides are arthropod neuropeptides with unknown function. In 2011 two RYamides were isolated from D. melanogaster as the ligands for the G-protein coupled receptor CG5811. The D. melanogaster gene encoding these neuropeptides is highly unusual, as there are four RYamide encoding exons in the current genome assembly, but an exon encoding a signal peptide is absent. Comparing the D. melanogaster gene structure with those from other species, including D. virilis, suggests that the gene is degenerating. RNAseq data from 1634 short sequence read archives at NCBI containing more than 34 billion spots yielded numerous individual spots that correspond to the RYamide encoding exons, of which a large number include the intron-exon boundary at the start of this exon. Although 72 different sequences have been spliced onto this RYamide encoding exon, none codes for the signal peptide of this gene. Thus, the RNAseq data for this gene reveal only noise and no signal. The very small quantities of peptide recovered during isolation and the absence of credible RNAseq data, indicates that the gene is very little expressed, while the RYamide gene structure in D. melanogaster suggests that it might be evolving into a pseudogene. Yet, the identification of the peptides it encodes clearly shows it is still functional. Using region specific antisera, we could localize numerous neurons and enteroendocrine cells in D. willistoni, D. virilis and D. pseudoobscura, but only two adult abdominal neurons in D. melanogaster. Those two neurons project to and innervate the rectal papillae, suggesting that RYamides may be involved in the regulation of water homeostasis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Analysis and Manipulation of Aspartate Pathway Genes for l-Lysine Overproduction from Methanol by Bacillus methanolicus▿

PubMed Central

Nærdal, Ingemar; Netzer, Roman; Ellingsen, Trond E.; Brautaset, Trygve

2011-01-01

We investigated the regulation and roles of six aspartate pathway genes in l-lysine overproduction in Bacillus methanolicus: dapG, encoding aspartokinase I (AKI); lysC, encoding AKII; yclM, encoding AKIII; asd, encoding aspartate semialdehyde dehydrogenase; dapA, encoding dihydrodipicolinate synthase; and lysA, encoding meso-diaminopimelate decarboxylase. Analysis of the wild-type strain revealed that in vivo lysC transcription was repressed 5-fold by l-lysine and induced 2-fold by dl-methionine added to the growth medium. Surprisingly, yclM transcription was repressed 5-fold by dl-methionine, while the dapG, asd, dapA, and lysA genes were not significantly repressed by any of the aspartate pathway amino acids. We show that the l-lysine-overproducing classical B. methanolicus mutant NOA2#13A52-8A66 has—in addition to a hom-1 mutation—chromosomal mutations in the dapG coding region and in the lysA promoter region. No mutations were found in its dapA, lysC, asd, and yclM genes. The mutant dapG gene product had abolished feedback inhibition by meso-diaminopimelate in vitro, and the lysA mutation was accompanied by an elevated (6-fold) lysA transcription level in vivo. Moreover, yclM transcription was increased 16-fold in mutant strain NOA2#13A52-8A66 compared to the wild-type strain. Overexpression of wild-type and mutant aspartate pathway genes demonstrated that all six genes are important for l-lysine overproduction as tested in shake flasks, and the effects were dependent on the genetic background tested. Coupled overexpression of up to three genes resulted in additive (above 80-fold) increased l-lysine production levels. PMID:21724876

Analysis and manipulation of aspartate pathway genes for L-lysine overproduction from methanol by Bacillus methanolicus.

PubMed

Nærdal, Ingemar; Netzer, Roman; Ellingsen, Trond E; Brautaset, Trygve

2011-09-01

We investigated the regulation and roles of six aspartate pathway genes in L-lysine overproduction in Bacillus methanolicus: dapG, encoding aspartokinase I (AKI); lysC, encoding AKII; yclM, encoding AKIII; asd, encoding aspartate semialdehyde dehydrogenase; dapA, encoding dihydrodipicolinate synthase; and lysA, encoding meso-diaminopimelate decarboxylase. Analysis of the wild-type strain revealed that in vivo lysC transcription was repressed 5-fold by L-lysine and induced 2-fold by dl-methionine added to the growth medium. Surprisingly, yclM transcription was repressed 5-fold by dl-methionine, while the dapG, asd, dapA, and lysA genes were not significantly repressed by any of the aspartate pathway amino acids. We show that the L-lysine-overproducing classical B. methanolicus mutant NOA2#13A52-8A66 has-in addition to a hom-1 mutation-chromosomal mutations in the dapG coding region and in the lysA promoter region. No mutations were found in its dapA, lysC, asd, and yclM genes. The mutant dapG gene product had abolished feedback inhibition by meso-diaminopimelate in vitro, and the lysA mutation was accompanied by an elevated (6-fold) lysA transcription level in vivo. Moreover, yclM transcription was increased 16-fold in mutant strain NOA2#13A52-8A66 compared to the wild-type strain. Overexpression of wild-type and mutant aspartate pathway genes demonstrated that all six genes are important for L-lysine overproduction as tested in shake flasks, and the effects were dependent on the genetic background tested. Coupled overexpression of up to three genes resulted in additive (above 80-fold) increased L-lysine production levels.

“Guilt by Association” Is the Exception Rather Than the Rule in Gene Networks

PubMed Central

Gillis, Jesse; Pavlidis, Paul

2012-01-01

Gene networks are commonly interpreted as encoding functional information in their connections. An extensively validated principle called guilt by association states that genes which are associated or interacting are more likely to share function. Guilt by association provides the central top-down principle for analyzing gene networks in functional terms or assessing their quality in encoding functional information. In this work, we show that functional information within gene networks is typically concentrated in only a very few interactions whose properties cannot be reliably related to the rest of the network. In effect, the apparent encoding of function within networks has been largely driven by outliers whose behaviour cannot even be generalized to individual genes, let alone to the network at large. While experimentalist-driven analysis of interactions may use prior expert knowledge to focus on the small fraction of critically important data, large-scale computational analyses have typically assumed that high-performance cross-validation in a network is due to a generalizable encoding of function. Because we find that gene function is not systemically encoded in networks, but dependent on specific and critical interactions, we conclude it is necessary to focus on the details of how networks encode function and what information computational analyses use to extract functional meaning. We explore a number of consequences of this and find that network structure itself provides clues as to which connections are critical and that systemic properties, such as scale-free-like behaviour, do not map onto the functional connectivity within networks. PMID:22479173

Mobile genetic element-encoded cytolysin connects virulence to methicillin resistance in MRSA.

PubMed

Queck, Shu Y; Khan, Burhan A; Wang, Rong; Bach, Thanh-Huy L; Kretschmer, Dorothee; Chen, Liang; Kreiswirth, Barry N; Peschel, Andreas; Deleo, Frank R; Otto, Michael

2009-07-01

Bacterial virulence and antibiotic resistance have a significant influence on disease severity and treatment options during bacterial infections. Frequently, the underlying genetic determinants are encoded on mobile genetic elements (MGEs). In the leading human pathogen Staphylococcus aureus, MGEs that contain antibiotic resistance genes commonly do not contain genes for virulence determinants. The phenol-soluble modulins (PSMs) are staphylococcal cytolytic toxins with a crucial role in immune evasion. While all known PSMs are core genome-encoded, we here describe a previously unidentified psm gene, psm-mec, within the staphylococcal methicillin resistance-encoding MGE SCCmec. PSM-mec was strongly expressed in many strains and showed the physico-chemical, pro-inflammatory, and cytolytic characteristics typical of PSMs. Notably, in an S. aureus strain with low production of core genome-encoded PSMs, expression of PSM-mec had a significant impact on immune evasion and disease. In addition to providing high-level resistance to methicillin, acquisition of SCCmec elements encoding PSM-mec by horizontal gene transfer may therefore contribute to staphylococcal virulence by substituting for the lack of expression of core genome-encoded PSMs. Thus, our study reveals a previously unknown role of methicillin resistance clusters in staphylococcal pathogenesis and shows that important virulence and antibiotic resistance determinants may be combined in staphylococcal MGEs.

Hypokalemic paralysis in a middle-aged female with classic Bartter syndrome.

PubMed

Chiang, Wen-Fang; Lin, Shih-Hung; Chan, Jenq-Shyong; Lin, Shih-Hua

2014-02-01

Inherited classic Bartter syndrome (cBS) is an autosomal recessive renal tubular disorder resulting from inactivating mutations in the asolateral chloride channel (C1C-Kb) and usually presents in early infancy or childhood with mild to moderate hypokalemia. Profound hypokalemic paralysis in patients with cBS is extremely rare, especially in middle age. A 45-year-old Chinese female patient was referred for evaluation of chronic severe hypokalemia despite regular K+ supplementation (1 mmol/kg/d). She had had two episodes of muscle paralysis due to severe hypokalemia (K+ 1.9 - 2.1 mmol/l) in the past 3 years. She denied vomiting, diarrhea, or the use of laxatives or diuretics. Her blood pressure was normal. Biochemical studies showed hypokalemia (K+ 2.5 mmol/l) with renal potassium wasting, metabolic alkalosis (HCO3- 32 mmol/l), normomagnesemia (Mg2+ 0.8 mmol/l), hypercalciuria (calcium to creatinine ratio 0.5 mmol/mmol; normal < 0.22 mmol/mol), high plasma renin activity, but normal plasma aldosterone concentration. Abdominal sonography revealed neither renal stones nor nephrocalcinosis. Acquired causes of cBS such as autoimmune disease and drugs were all excluded. Molecular analysis of the CLCNKB gene, encoding ClC-Kb, and SLC12A3, encoding the thiazide-sensitive sodium chloride cotransporter (NCC), revealed compound heterozygous mutations in CLCNKB (L335P and G470E) inherited from her parents; her SLC12A3 was normal. These two mutations were not identified in 100 healthy subjects. Her plasma K+ concentration rose to 3 - 3.5 mmol/l after the addition of spironolactone. Inherited cBS may present with hypokalemic paralysis and should be considered in adult patients with hypokalemia and metabolic alkalosis.

Novel Thrombotic Function of a Human SNP in STXBP5 Revealed by CRISPR/Cas9 Gene Editing in Mice.

PubMed

Zhu, Qiuyu Martin; Ko, Kyung Ae; Ture, Sara; Mastrangelo, Michael A; Chen, Ming-Huei; Johnson, Andrew D; O'Donnell, Christopher J; Morrell, Craig N; Miano, Joseph M; Lowenstein, Charles J

2017-02-01

To identify and characterize the effect of a SNP (single-nucleotide polymorphism) in the STXBP5 locus that is associated with altered thrombosis in humans. GWAS (genome-wide association studies) have identified numerous SNPs associated with human thrombotic phenotypes, but determining the functional significance of an individual candidate SNP can be challenging, particularly when in vivo modeling is required. Recent GWAS led to the discovery of STXBP5 as a regulator of platelet secretion in humans. Further clinical studies have identified genetic variants of STXBP5 that are linked to altered plasma von Willebrand factor levels and thrombosis in humans, but the functional significance of these variants in STXBP5 is not understood. We used CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated 9) techniques to produce a precise mouse model carrying a human coding SNP rs1039084 (encoding human p. N436S) in the STXBP5 locus associated with decreased thrombosis. Mice carrying the orthologous human mutation (encoding p. N437S in mouse STXBP5) have lower plasma von Willebrand factor levels, decreased thrombosis, and decreased platelet secretion compared with wild-type mice. This thrombosis phenotype recapitulates the phenotype of humans carrying the minor allele of rs1039084. Decreased plasma von Willebrand factor and platelet activation may partially explain the decreased thrombotic phenotype in mutant mice. Using precise mammalian genome editing, we have identified a human nonsynonymous SNP rs1039084 in the STXBP5 locus as a causal variant for a decreased thrombotic phenotype. CRISPR/Cas9 genetic editing facilitates the rapid and efficient generation of animals to study the function of human genetic variation in vascular diseases. © 2016 American Heart Association, Inc.

Staphylococcus aureus nasal carriage in Ukraine: antibacterial resistance and virulence factor encoding genes.

PubMed

Netsvyetayeva, Irina; Fraczek, Mariusz; Piskorska, Katarzyna; Golas, Marlena; Sikora, Magdalena; Mlynarczyk, Andrzej; Swoboda-Kopec, Ewa; Marusza, Wojciech; Palmieri, Beniamino; Iannitti, Tommaso

2014-03-05

The number of studies regarding the incidence of multidrug resistant strains and distribution of genes encoding virulence factors, which have colonized the post-Soviet states, is considerably limited. The aim of the study was (1) to assess the Staphylococcus (S.) aureus nasal carriage rate, including Methicillin Resistant S. aureus (MRSA) strains in adult Ukrainian population, (2) to determine antibiotic resistant pattern and (3) the occurrence of Panton Valentine Leukocidine (PVL)-, Fibronectin-Binding Protein A (FnBPA)- and Exfoliative Toxin (ET)-encoding genes. Nasal samples for S. aureus culture were obtained from 245 adults. The susceptibility pattern for several classes of antibiotics was determined by disk diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The virulence factor encoding genes, mecA, lukS-lukF, eta, etb, etd, fnbA, were detected by Polymerase Chain Reaction (PCR). The S. aureus nasal carriage rate was 40%. The prevalence of nasal MRSA carriage in adults was 3.7%. LukS-lukF genes were detected in over 58% of the strains. ET-encoding genes were detected in over 39% of the strains and the most prevalent was etd. The fnbA gene was detected in over 59% of the strains. All MRSA isolates tested were positive for the mecA gene. LukS-lukF genes and the etd gene were commonly co-present in MRSA, while lukS-lukF genes and the fnbA gene were commonly co-present in Methicillin Sensitive S. aureus (MSSA) isolates. No significant difference was detected between the occurrence of lukS-lukF genes (P > 0.05) and the etd gene (P > 0.05) when comparing MRSA and MSSA. The occurrence of the fnbA gene was significantly more frequent in MSSA strains (P < 0.05). In Ukraine, S. aureus is a common cause of infection. The prevalence of S. aureus nasal carriage in our cohort of patients from Ukraine was 40.4%. We found that 9.1% of the strains were classified as MRSA and all MRSA isolates tested positive for the mecA gene. We also observed a high prevalence of PVL- and ET- encoding genes among S. aureus nasal carriage strains. A systematic surveillance system can help prevent transmission and spread of drug resistant toxin producing S. aureus strains.

The Ebola virus matrix protein penetrates into the plasma membrane: a key step in viral protein 40 (VP40) oligomerization and viral egress.

PubMed

Adu-Gyamfi, Emmanuel; Soni, Smita P; Xue, Yi; Digman, Michelle A; Gratton, Enrico; Stahelin, Robert V

2013-02-22

Ebola, a fatal virus in humans and non-human primates, has no Food and Drug Administration-approved vaccines or therapeutics. The virus from the Filoviridae family causes hemorrhagic fever, which rapidly progresses and in some cases has a fatality rate near 90%. The Ebola genome encodes seven genes, the most abundantly expressed of which is viral protein 40 (VP40), the major Ebola matrix protein that regulates assembly and egress of the virus. It is well established that VP40 assembles on the inner leaflet of the plasma membrane; however, the mechanistic details of plasma membrane association by VP40 are not well understood. In this study, we used an array of biophysical experiments and cellular assays along with mutagenesis of VP40 to investigate the role of membrane penetration in VP40 assembly and egress. Here we demonstrate that VP40 is able to penetrate specifically into the plasma membrane through an interface enriched in hydrophobic residues in its C-terminal domain. Mutagenesis of this hydrophobic region consisting of Leu(213), Ile(293), Leu(295), and Val(298) demonstrated that membrane penetration is critical to plasma membrane localization, VP40 oligomerization, and viral particle egress. Taken together, VP40 membrane penetration is an important step in the plasma membrane localization of the matrix protein where oligomerization and budding are defective in the absence of key hydrophobic interactions with the membrane.

Carbohydrate metabolism genes and pathways in insects: insights from the honey bee genome

PubMed Central

Kunieda, T; Fujiyuki, T; Kucharski, R; Foret, S; Ament, S A; Toth, A L; Ohashi, K; Takeuchi, H; Kamikouchi, A; Kage, E; Morioka, M; Beye, M; Kubo, T; Robinson, G E; Maleszka, R

2006-01-01

Carbohydrate-metabolizing enzymes may have particularly interesting roles in the honey bee, Apis mellifera, because this social insect has an extremely carbohydrate-rich diet, and nutrition plays important roles in caste determination and socially mediated behavioural plasticity. We annotated a total of 174 genes encoding carbohydrate-metabolizing enzymes and 28 genes encoding lipid-metabolizing enzymes, based on orthology to their counterparts in the fly, Drosophila melanogaster, and the mosquito, Anopheles gambiae. We found that the number of genes for carbohydrate metabolism appears to be more evolutionarily labile than for lipid metabolism. In particular, we identified striking changes in gene number or genomic organization for genes encoding glycolytic enzymes, cellulase, glucose oxidase and glucose dehydrogenases, glucose-methanol-choline (GMC) oxidoreductases, fucosyltransferases, and lysozymes. PMID:17069632

Novel Type V Staphylococcal Cassette Chromosome mec Driven by a Novel Cassette Chromosome Recombinase, ccrC

PubMed Central

Ito, Teruyo; Ma, Xiao Xue; Takeuchi, Fumihiko; Okuma, Keiko; Yuzawa, Harumi; Hiramatsu, Keiichi

2004-01-01

Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic element composed of the mec gene complex, which encodes methicillin resistance, and the ccr gene complex, which encodes the recombinases responsible for its mobility. The mec gene complex has been classified into four classes, and the ccr gene complex has been classified into three allotypes. Different combinations of mec gene complex classes and ccr gene complex types have so far defined four types of SCCmec elements. Now we introduce the fifth allotype of SCCmec, which was found on the chromosome of a community-acquired methicillin-resistant Staphylococcus aureus strain (strain WIS [WBG8318]) isolated in Australia. The element shared the same chromosomal integration site with the four extant types of SCCmec and the characteristic nucleotide sequences at the chromosome-SCCmec junction regions. The novel SCCmec carried mecA bracketed by IS431 (IS431-mecA-ΔmecR1-IS431), which is designated the class C2 mec gene complex; and instead of ccrA and ccrB genes, it carried a single copy of a gene homologue that encoded cassette chromosome recombinase. Since the open reading frame (ORF) was found to encode an enzyme which catalyzes the precise excision as well as site- and orientation-specific integration of the element, we designated the ORF cassette chromosome recombinase C (ccrC), and we designated the element type V SCCmec. Type V SCCmec is a small SCCmec element (28 kb) and does not carry any antibiotic resistance genes besides mecA. Unlike the extant SCCmec types, it carries a set of foreign genes encoding a restriction-modification system that might play a role in the stabilization of the element on the chromosome. PMID:15215121

Complete genome sequence of Nitrosospira multiformis, an ammonia-oxidizing bacterium from the soil environment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Norton, Jeanette M.; Klotz, Martin G; Stein, Lisa Y

2008-01-01

The complete genome of the ammonia-oxidizing bacterium, Nitrosospira multiformis (ATCC 25196T), consists of a circular chromosome and three small plasmids totaling 3,234,309 bp and encoding 2827 putative proteins. Of these, 2026 proteins have predicted functions and 801 are without conserved functional domains, yet 747 of these have similarity to other predicted proteins in databases. Gene homologs from Nitrosomonas europaea and N. eutropha were the best match for 42% of the predicted genes in N. multiformis. The genome contains three nearly identical copies of amo and hao gene clusters as large repeats. Distinguishing features compared to N. europaea include: the presencemore » of gene clusters encoding urease and hydrogenase, a RuBisCO-encoding operon of distinctive structure and phylogeny, and a relatively small complement of genes related to Fe acquisition. Systems for synthesis of a pyoverdine-like siderophore and for acyl-homoserine lactone were unique to N. multiformis among the sequenced AOB genomes. Gene clusters encoding proteins associated with outer membrane and cell envelope functions including transporters, porins, exopolysaccharide synthesis, capsule formation and protein sorting/export were abundant. Numerous sensory transduction and response regulator gene systems directed towards sensing of the extracellular environment are described. Gene clusters for glycogen, polyphosphate and cyanophycin storage and utilization were identified providing mechanisms for meeting energy requirements under substrate-limited conditions. The genome of N. multiformis encodes the core pathways for chemolithoautotrophy along with adaptations for surface growth and survival in soil environments.« less

Transcriptional Profiling of Caulobacter crescentus during Growth on Complex and Minimal Media

PubMed Central

Hottes, Alison K.; Meewan, Maliwan; Yang, Desiree; Arana, Naomi; Romero, Pedro; McAdams, Harley H.; Stephens, Craig

2004-01-01

Microarray analysis was used to examine gene expression in the freshwater oligotrophic bacterium Caulobacter crescentus during growth on three standard laboratory media, including peptone-yeast extract medium (PYE) and minimal salts medium with glucose or xylose as the carbon source. Nearly 400 genes (approximately 10% of the genome) varied significantly in expression between at least two of these media. The differentially expressed genes included many encoding transport systems, most notably diverse TonB-dependent outer membrane channels of unknown substrate specificity. Amino acid degradation pathways constituted the largest class of genes induced in PYE. In contrast, many of the genes upregulated in minimal media encoded enzymes for synthesis of amino acids, including incorporation of ammonia and sulfate into glutamate and cysteine. Glucose availability induced expression of genes encoding enzymes of the Entner-Doudoroff pathway, which was demonstrated here through mutational analysis to be essential in C. crescentus for growth on glucose. Xylose induced expression of genes encoding several hydrolytic exoenzymes as well as an operon that may encode a novel pathway for xylose catabolism. A conserved DNA motif upstream of many xylose-induced genes was identified and shown to confer xylose-specific expression. Xylose is an abundant component of xylan in plant cell walls, and the microarray data suggest that in addition to serving as a carbon source for growth of C. crescentus, this pentose may be interpreted as a signal to produce enzymes associated with plant polymer degradation. PMID:14973021

Pathogen recognition in the innate immune response.

PubMed

Kumar, Himanshu; Kawai, Taro; Akira, Shizuo

2009-04-28

Immunity against microbial pathogens primarily depends on the recognition of pathogen components by innate receptors expressed on immune and non-immune cells. Innate receptors are evolutionarily conserved germ-line-encoded proteins and include TLRs (Toll-like receptors), RLRs [RIG-I (retinoic acid-inducible gene-I)-like receptors] and NLRs (Nod-like receptors). These receptors recognize pathogens or pathogen-derived products in different cellular compartments, such as the plasma membrane, the endosomes or the cytoplasm, and induce the expression of cytokines, chemokines and co-stimulatory molecules to eliminate pathogens and instruct pathogen-specific adaptive immune responses. In the present review, we will discuss the recent progress in the study of pathogen recognition by TLRs, RLRs and NLRs and their signalling pathways.

Molecular cloning and characterization of alpha - galactosidase gene from Glaciozyma antarctica

NASA Astrophysics Data System (ADS)

Moheer, Reyad Qaed Al; Bakar, Farah Diba Abu; Murad, Abdul Munir Abdul

2015-09-01

Psychrophilic enzymes are proteins produced by psychrophilic organisms which recently are the limelight for industrial applications. A gene encoding α-galactosidase from a psychrophilic yeast, Glaciozyma antarctica PI12 which belongs to glycoside hydrolase family 27, was isolated and analyzed using several bioinformatic tools. The cDNA of the gene with the size of 1,404-bp encodes a protein with 467 amino acid residues. Predicted molecular weight of protein was 48.59 kDa and hence we name the gene encoding α-galactosidase as GAL48. We found that the predicted protein sequences possessed signal peptide sequence and are highly conserved among other fungal α-galactosidase.

Comprehensive analysis of gene expression patterns in Friedreich's ataxia fibroblasts by RNA sequencing reveals altered levels of protein synthesis factors and solute carriers

PubMed Central

Li, Yanjie; Lu, Yue; Lin, Kevin; Hauser, Lauren A.; Lynch, David R.

2017-01-01

ABSTRACT Friedreich's ataxia (FRDA) is an autosomal recessive neurodegenerative disease usually caused by large homozygous expansions of GAA repeat sequences in intron 1 of the frataxin (FXN) gene. FRDA patients homozygous for GAA expansions have low FXN mRNA and protein levels when compared with heterozygous carriers or healthy controls. Frataxin is a mitochondrial protein involved in iron–sulfur cluster synthesis, and many FRDA phenotypes result from deficiencies in cellular metabolism due to lowered expression of FXN. Presently, there is no effective treatment for FRDA, and biomarkers to measure therapeutic trial outcomes and/or to gauge disease progression are lacking. Peripheral tissues, including blood cells, buccal cells and skin fibroblasts, can readily be isolated from FRDA patients and used to define molecular hallmarks of disease pathogenesis. For instance, FXN mRNA and protein levels as well as FXN GAA-repeat tract lengths are routinely determined using all of these cell types. However, because these tissues are not directly involved in disease pathogenesis, their relevance as models of the molecular aspects of the disease is yet to be decided. Herein, we conducted unbiased RNA sequencing to profile the transcriptomes of fibroblast cell lines derived from 18 FRDA patients and 17 unaffected control individuals. Bioinformatic analyses revealed significantly upregulated expression of genes encoding plasma membrane solute carrier proteins in FRDA fibroblasts. Conversely, the expression of genes encoding accessory factors and enzymes involved in cytoplasmic and mitochondrial protein synthesis was consistently decreased in FRDA fibroblasts. Finally, comparison of genes differentially expressed in FRDA fibroblasts to three previously published gene expression signatures defined for FRDA blood cells showed substantial overlap between the independent datasets, including correspondingly deficient expression of antioxidant defense genes. Together, these results indicate that gene expression profiling of cells derived from peripheral tissues can, in fact, consistently reveal novel molecular pathways of the disease. When performed on statistically meaningful sample group sizes, unbiased global profiling analyses utilizing peripheral tissues are critical for the discovery and validation of FRDA disease biomarkers. PMID:29125828

Isolation of Nicotiana plumbaginifolia cDNAs encoding isoforms of serine acetyltransferase and O-acetylserine (thiol) lyase in a yeast two-hybrid system with Escherichia coli cysE and cysK genes as baits.

PubMed

Liszewska, Frantz; Gaganidze, Dali; Sirko, Agnieszka

2005-01-01

We applied the yeast two-hybrid system for screening of a cDNA library of Nicotiana plumbaginifolia for clones encoding plant proteins interacting with two proteins of Escherichia coli: serine acetyltransferase (SAT, the product of cysE gene) and O-acetylserine (thiol)lyase A, also termed cysteine synthase (OASTL-A, the product of cysK gene). Two plant cDNA clones were identified when using the cysE gene as a bait. These clones encode a probable cytosolic isoform of OASTL and an organellar isoform of SAT, respectively, as indicated by evolutionary trees. The second clone, encoding SAT, was identified independently also as a "prey" when using cysK as a bait. Our results reveal the possibility of applying the two-hybrid system for cloning of plant cDNAs encoding enzymes of the cysteine synthase complex in the two-hybrid system. Additionally, using genome walking sequences located upstream of the sat1 cDNA were identified. Subsequently, in silico analyses were performed aiming towards identification of the potential signal peptide and possible location of the deduced mature protein encoded by sat1.

Genetic and functional properties of uncultivated thermophilic crenarchaeotes from a subsurface gold mine as revealed by analysis of genome fragments.

PubMed

Nunoura, Takuro; Hirayama, Hisako; Takami, Hideto; Oida, Hanako; Nishi, Shinro; Shimamura, Shigeru; Suzuki, Yohey; Inagaki, Fumio; Takai, Ken; Nealson, Kenneth H; Horikoshi, Koki

2005-12-01

Within a phylum Crenarchaeota, only some members of the hyperthermophilic class Thermoprotei, have been cultivated and characterized. In this study, we have constructed a metagenomic library from a microbial mat formation in a subsurface hot water stream of the Hishikari gold mine, Japan, and sequenced genome fragments of two different phylogroups of uncultivated thermophilic Crenarchaeota: (i) hot water crenarchaeotic group (HWCG) I (41.2 kb), and (ii) HWCG III (49.3 kb). The genome fragment of HWCG I contained a 16S rRNA gene, two tRNA genes and 35 genes encoding proteins but no 23S rRNA gene. Among the genes encoding proteins, several genes for putative aerobic-type carbon monoxide dehydrogenase represented a potential clue with regard to the yet unknown metabolism of HWCG I Archaea. The genome fragment of HWCG III contained a 16S/23S rRNA operon and 44 genes encoding proteins. In the 23S rRNA gene, we detected a homing-endonuclease encoding a group I intron similar to those detected in hyperthermophilic Crenarchaeota and Bacteria, as well as eukaryotic organelles. The reconstructed phylogenetic tree based on the 23S rRNA gene sequence reinforced the intermediate phylogenetic affiliation of HWCG III bridging the hyperthermophilic and non-thermophilic uncultivated Crenarchaeota.

A de novo KCNA1 Mutation in a Patient with Tetany and Hypomagnesemia.

PubMed

van der Wijst, Jenny; Konrad, Martin; Verkaart, Sjoerd A J; Tkaczyk, Marcin; Latta, Femke; Altmüller, Janine; Thiele, Holger; Beck, Bodo; Schlingmann, Karl Peter; de Baaij, Jeroen H F

2018-05-23

Mutations in the KCNA1 gene encoding the voltage-gated potassium (K+) channel Kv1.1 have been linked to rare neurological syndromes, episodic ataxia type 1 (EA1) and myokymia. In 2009, a KCNA1 mutation was identified in a large family with autosomal dominant hypomagnesemia. Despite efforts in establishing a genotype-phenotype correlation for the wide variety of symptoms in EA1, little is known on the serum magnesium (Mg2+) levels in these patients. In the present study, we describe a new de novo KCNA1 mutation in a Polish patient with tetany and hypomagnesemia. Electrophysiological and biochemical analyses were performed to determine the pathogenicity of the mutation. A female patient presented with low serum Mg2+ levels, renal Mg2+ wasting, muscle cramps, and tetanic episodes. Whole exome sequencing identified a p.Leu328Val mutation in KCNA1 encoding the Kv1.1 K+ channel. Electrophysiological examinations demonstrated that the p.Leu328Val mutation caused a dominant-negative loss of function of the encoded Kv1.1 channel. Cell surface biotinylation showed normal plasma membrane expression. Taken together, this is the second report linking KCNA1 with hypomagnesemia, thereby emphasizing the need for further evaluation of the clinical phenotypes observed in patients carrying KCNA1 mutations. © 2018 S. Karger AG, Basel.

The genomic structure of the human Charcot-Leyden crystal protein gene is analogous to those of the galectin genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dyer, K.D.; Handen, J.S.; Rosenberg, H.F.

The Charcot-Leyden crystal (CLC) protein, or eosinophil lysophospholipase, is a characteristic protein of human eosinophils and basophils; recent work has demonstrated that the CLC protein is both structurally and functionally related to the galectin family of {beta}-galactoside binding proteins. The galectins as a group share a number of features in common, including a linear ligand binding site encoded on a single exon. In this work, we demonstrate that the intron-exon structure of the gene encoding CLC is analogous to those encoding the galectins. The coding sequence of the CLC gene is divided into four exons, with the entire {beta}-galactoside bindingmore » site encoded by exon III. We have isolated CLC {beta}-galactoside binding sites from both orangutan (Pongo pygmaeus) and murine (Mus musculus) genomic DNAs, both encoded on single exons, and noted conservation of the amino acids shown to interact directly with the {beta}-galactoside ligand. The most likely interpretation of these results suggests the occurrence of one or more exon duplication and insertion events, resulting in the distribution of this lectin domain to CLC as well as to the multiple galectin genes. 35 refs., 3 figs.« less

Recombination and mutation of class II histocompatibility genes in wild mice.

PubMed

Wakeland, E K; Darby, B R

1983-12-01

We have compared the tryptic peptide fingerprints of the A alpha, A beta, E alpha, and E beta subunits encoded by four wild-derived H-2 complexes expressing A molecules closely related to Ak. The A molecules encoded by these Ak-related mice have A alpha and A beta subunits that differ from A alpha k and A beta k by less than 10% of their tryptic peptides. Comparisons among the four wild-derived A molecules suggested that these contemporary A alpha and A beta alleles arose by sequential mutational events from common ancestor A alpha and A beta alleles. These results suggest that A alpha and A beta may co-evolve as an A beta A alpha gene duplex in wild mice. Tryptic peptide fingerprint comparisons of the E beta gene linked to these Ak-related A beta A alpha gene duplexes indicate that two encode E beta d-like subunits, whereas another encodes an E beta s-like subunit. These results strongly suggest that the A beta A alpha duplex and E beta recombine in wild mouse populations. The significantly different evolutionary patterns exhibited by the class II genes encoding A vs E molecules are discussed.

Interleukin 28B gene polymorphisms and Epstein-Barr virus-associated lymphoproliferative diseases.

PubMed

Akay, Ela; Patel, Mauli; Conibear, Tim; Chaggar, Turren; Haque, Tanzina

2014-01-01

Single-nucleotide polymorphisms (SNPs) near the interleukin (IL) 28B gene encoding a type III interferon (IFN-λ) are the most important genetic predictors of treatment response to hepatitis C virus (HCV). This retrospective study was undertaken to determine any association between IL28B SNPs and the development of viraemia in Epstein-Barr virus (EBV)-driven acute infectious mononucleosis (IM) and post-transplant lymphoproliferative disease (PTLD). Genomic DNA extracted from plasma from 45 EBV seropositive controls and 46 acute IM, 23 non-PTLD (transplant) and 21 PTLD patients was tested by PCR for 2 SNPs within IL28B. EBV DNA levels were tested in IM and PTLD samples by a real-time quantitative PCR. No significant differences were seen in SNP frequencies at rs12979860 and rs8099917 in IM and PTLD patients compared to EBV seropositive controls and transplant patients. EBV DNA levels were lower in IM and PTLD patients with CC (a favourable genotype in HCV) at rs12979860 compared to non-CC genotypes (p = 0.055). Acute IM patients with CC had significantly lower levels of EBV DNA in plasma compared to those with non-CC genotypes (p = 0.011). Genotype CC may influence anti-viral responses of IFN-λ, thereby allowing better control of EBV viraemia during lymphoproliferation, particularly in IM.

A Common Variant in the Von Willebrand Factor Gene is Associated With Multiple Functional Consequences

PubMed Central

Vaidya, Dhananjay; Yanek, Lisa R.; Herrera-Galeano, J. Enrique; Mathias, Rasika A.; Moy, Taryn F.; Faraday, Nauder; Becker, Lewis C.; Becker, Diane M.

2010-01-01

Von Willebrand Factor (vWF) is a plasma protein involved in thrombosis and hemostasis [1]. We examined whether common single nucleotide polymorphisms (SNPs) in the vWF gene were associated with vWF levels and platelet aggregation-related functional consequences in1230 Whites and 837 African Americans in a cross-sectional family based genetic study of platelet function. From a high-density scan, 28 SNPs with a minor allele frequency > 5% in both races were tested for association using age and sex adjusted variance components analysis in MERLIN. SNP rs216321, with the strongest association with vWF levels in biracial metaanalysis (p=9.5×10−6, Whites–p=8.1×10−4, African Americans–p=3.6×10−3), encoding a R852Q substitution in the D’D3 protein domain, demonstrated negative association with plasma vWF. The R852Q variant was recessively associated with 15.5% lower collagen-induced platelet aggregation adjusting for dose-response relationship (p=0.010, vWF-level adjusted p=0.003). Each copy of the R852Q variant was additively associated with 31% higher FVIII levels (p=0.039, vWF-adjusted p=0.033). In conclusion, this common missense polymorphism appears to have pleiotropic functional consequences. PMID:20941784

Multi-functional acetyl-CoA carboxylase from Brassica napus is encoded by a multi-gene family: indication for plastidic localization of at least one isoform.

PubMed

Schulte, W; Töpfer, R; Stracke, R; Schell, J; Martini, N

1997-04-01

Three genes coding for different multifunctional acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) isoenzymes from Brassica napus were isolated and divided into two major classes according to structural features in their 5' regions: class I comprises two genes with an additional coding exon of approximately 300 bp at the 5' end, and class II is represented by one gene carrying an intron of 586 bp in its 5' untranslated region. Fusion of the peptide sequence encoded by the additional first exon of a class I ACCase gene to the jellyfish Aequorea victoria green fluorescent protein (GFP) and transient expression in tobacco protoplasts targeted GFP to the chloroplasts. In contrast to the deduced primary structure of the biotin carboxylase domain encoded by the class I gene, the corresponding amino acid sequence of the class II ACCase shows higher identity with that of the Arabidopsis ACCase, both lacking a transit peptide. The Arabidopsis ACCase has been proposed to be a cytosolic isoenzyme. These observations indicate that the two classes of ACCase genes encode plastidic and cytosolic isoforms of multi-functional, eukaryotic type, respectively, and that B. napus contains at least one multi-functional ACCase besides the multi-subunit, prokaryotic type located in plastids. Southern blot analysis of genomic DNA from B. napus, Brassica rapa, and Brassica oleracea, the ancestors of amphidiploid rapeseed, using a fragment of a multi-functional ACCase gene as a probe revealed that ACCase is encoded by a multi-gene family of at least five members.

Apolipoprotein E gene polymorphism and gender.

PubMed

Kolovou, Genovefa; Damaskos, Dimitris; Anagnostopoulou, Katherine; Cokkinos, Dennis V

2009-01-01

Many studies have shown that the prevalence and onset of coronary heart disease (CHD) is sex-dependent. CHD prevalence is lower in women than in men at all ages. Furthermore, women's age of CHD onset seems to be 10 yr later. This is widely attributed to the fact that men have less favorable CHD risk factors (eg, plasma lipid profile) compared to women. Mean levels of protective high density lipoprotein cholesterol are lower, while triglyceride levels are higher in men than in women. It is possible that many of the genes involved in lipid metabolism, such as Apolipoprotein (Apo) E, as well as their polymorphisms, may be expressed in a sexually dimorphic manner. The human Apo E gene is polymorphic, encoding one of 3 common epsilon (epsilon) alleles (epsilon 2, epsilon 3, epsilon 4), with the epsilon 3 allele occurring most frequently (78%) in the Caucasian population. Association studies have shown a protective effect on CHD in epsilon 2 carriers and a harmful effect in epsilon 4 carriers. However, there are conflicting results regarding such allelic effects in respect to gender. This review is focused on the gender-related influence of Apo E polymorphism in respect to plasma lipid levels and the risk of CHD. Additionally, an effort is made to determine if this relation exists and if it can be satisfactorily explained. The studies cited here demonstrate a complex, multifactorial association between these factors, in need of further corroboration in greater population samples.

CXCL14 is a candidate biomarker for Hedgehog signalling in idiopathic pulmonary fibrosis.

PubMed

Jia, Guiquan; Chandriani, Sanjay; Abbas, Alexander R; DePianto, Daryle J; N'Diaye, Elsa N; Yaylaoglu, Murat B; Moore, Heather M; Peng, Ivan; DeVoss, Jason; Collard, Harold R; Wolters, Paul J; Egen, Jackson G; Arron, Joseph R

2017-09-01

Idiopathic pulmonary fibrosis (IPF) is associated with aberrant expression of developmental pathways, including Hedgehog (Hh). As Hh signalling contributes to multiple pro-fibrotic processes, Hh inhibition may represent a therapeutic option for IPF. However, no non-invasive biomarkers are available to monitor lung Hh activity. We assessed gene and protein expression in IPF and control lung biopsies, mouse lung, fibroblasts stimulated in vitro with sonic hedgehog (SHh), and plasma in IPF patients versus controls, and cancer patients before and after treatment with vismodegib, a Hh inhibitor. Lung tissue from IPF patients exhibited significantly greater expression of Hh-related genes versus controls. The gene most significantly upregulated in both IPF lung biopsies and fibroblasts stimulated in vitro with SHh was CXCL14 , which encodes a soluble secreted chemokine whose expression is inhibited in vitro by the addition of vismodegib. CXCL14 expression was induced by SHh overexpression in mouse lung. Circulating CXCL14 protein levels were significantly higher in plasma from IPF patients than controls. In cancer patients, circulating CXCL14 levels were significantly reduced upon vismodegib treatment. CXCL14 is a systemic biomarker that could be used to identify IPF patients with increased Hh pathway activity and monitor the pharmacodynamic effects of Hh antagonist therapy in IPF. Post-results, NCT00968981. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Characterization of GM-CSF-inhibitory factor and Uracil DNA glycosylase encoding genes from camel pseudocowpoxvirus.

PubMed

Nagarajan, G; Swami, Shelesh Kumar; Dahiya, Shyam Singh; Narnaware, S D; Mehta, S C; Singh, P K; Singh, Raghvendar; Tuteja, F C; Patil, N V

2015-06-01

The present study describes the PCR amplification of GM-CSF-inhibitory factor (GIF) and Uracil DNA glycosylase (UDG) encoding genes of pseudocowpoxvirus (PCPV) from the Indian Dromedaries (Camelus dromedarius) infected with contagious ecthyma using the primers based on the corresponding gene sequences of human PCPV and reindeer PCPV, respectively. The length of GIF gene of PCPV obtained from camel is 795 bp and due to the addition of one cytosine residue at position 374 and one adenine residue at position 516, the open reading frame (ORF) got altered, resulting in the production of truncated polypeptide. The ORF of UDG encoding gene of camel PCPV is 696 bp encoding a polypeptide of 26.0 kDa. Comparison of amino acid sequence homologies of GIF and UDG of camel PCPV revealed that the camel PCPV is closer to ORFV and PCPV (reference stains of both human and reindeer), respectively. Copyright © 2015 Elsevier Ltd. All rights reserved.

A mutation in a new gene bglJ, activates the bgl operon in Escherichia coli K-12

DOE Office of Scientific and Technical Information (OSTI.GOV)

Giel, M.; Desnoyer, M.; Lopilato, J.

1996-06-01

A new mutation , bglJ4, has been characterized that results in the expression of the silent bgl operon. The bgl operon encodes proteins necessary for the transport and utilization of the aromatic {beta}-glucosides arbutin and salicin. A variety of mutations activate the operon and result in a Bgl{sup +} phenotype. Activating mutations are located upstream of the bgl promoter and in genes located elsewhere on the chromosome. Mutations outside of the bgl operon occur in the genes encoding DNA gyrase and in the gene encoding the nucleoid associated protein H-NS. The mutation described here, bglJ4, has been mapped to amore » new locus at min 99 on the Escherichia coli K-12 genetic map. The putative protein encoded by the bglJ gene has homology to a family of transcriptional activators. Evidence is presented that increased expression of the bglJ product is needed for activation of the bgl operon. 56 refs., 3 figs., 3 tabs.« less

Two Closely Related Genes of Arabidopsis Encode Plastidial Cytidinediphosphate Diacylglycerol Synthases Essential for Photoautotrophic Growth1[C

PubMed Central

Haselier, André; Akbari, Hana; Weth, Agnes; Baumgartner, Werner; Frentzen, Margrit

2010-01-01

Cytidinediphosphate diacylglycerol synthase (CDS) catalyzes the formation of cytidinediphosphate diacylglycerol, an essential precursor of anionic phosphoglycerolipids like phosphatidylglycerol or -inositol. In plant cells, CDS isozymes are located in plastids, mitochondria, and microsomes. Here, we show that these isozymes are encoded by five genes in Arabidopsis (Arabidopsis thaliana). Alternative translation initiation or alternative splicing of CDS2 and CDS4 transcripts can result in up to 10 isoforms. Most of the cDNAs encoding the various plant isoforms were functionally expressed in yeast and rescued the nonviable phenotype of the mutant strain lacking CDS activity. The closely related genes CDS4 and CDS5 were found to encode plastidial isozymes with similar catalytic properties. Inactivation of both genes was required to obtain Arabidopsis mutant lines with a visible phenotype, suggesting that the genes have redundant functions. Analysis of these Arabidopsis mutants provided further independent evidence for the importance of plastidial phosphatidylglycerol for structure and function of thylakoid membranes and, hence, for photoautotrophic growth. PMID:20442275

The rgg0182 gene encodes a transcriptional regulator required for the full Streptococcus thermophilus LMG18311 thermal adaptation.

PubMed

Henry, Romain; Bruneau, Emmanuelle; Gardan, Rozenn; Bertin, Stéphane; Fleuchot, Betty; Decaris, Bernard; Leblond-Bourget, Nathalie

2011-10-07

Streptococcus thermophilus is an important starter strain for the production of yogurt and cheeses. The analysis of sequenced genomes of four strains of S. thermophilus indicates that they contain several genes of the rgg familly potentially encoding transcriptional regulators. Some of the Rgg proteins are known to be involved in bacterial stress adaptation. In this study, we demonstrated that Streptococcus thermophilus thermal stress adaptation required the rgg0182 gene which transcription depends on the culture medium and the growth temperature. This gene encoded a protein showing similarity with members of the Rgg family transcriptional regulator. Our data confirmed that Rgg0182 is a transcriptional regulator controlling the expression of its neighboring genes as well as chaperones and proteases encoding genes. Therefore, analysis of a Δrgg0182 mutant revealed that this protein played a role in the heat shock adaptation of Streptococcus thermophilus LMG18311. These data showed the importance of the Rgg0182 transcriptional regulator on the survival of S. thermophilus during dairy processes and more specifically during changes in temperature.

Identification of an opd (organophosphate degradation) gene in an Agrobacterium isolate.

PubMed

Horne, Irene; Sutherland, Tara D; Harcourt, Rebecca L; Russell, Robyn J; Oakeshott, John G

2002-07-01

We isolated a bacterial strain, Agrobacterium radiobacter P230, which can hydrolyze a wide range of organophosphate (OP) insecticides. A gene encoding a protein involved in OP hydrolysis was cloned from A. radiobacter P230 and sequenced. This gene (called opdA) had sequence similarity to opd, a gene previously shown to encode an OP-hydrolyzing enzyme in Flavobacterium sp. strain ATCC 27551 and Brevundimonas diminuta MG. Insertional mutation of the opdA gene produced a strain lacking the ability to hydrolyze OPs, suggesting that this is the only gene encoding an OP-hydrolyzing enzyme in A. radiobacter P230. The OPH and OpdA proteins, encoded by opd and opdA, respectively, were overexpressed and purified as maltose-binding proteins, and the maltose-binding protein moiety was cleaved and removed. Neither protein was able to hydrolyze the aliphatic OP malathion. The kinetics of the two proteins for diethyl OPs were comparable. For dimethyl OPs, OpdA had a higher k(cat) than OPH. It was also capable of hydrolyzing the dimethyl OPs phosmet and fenthion, which were not hydrolyzed at detectable levels by OPH.

Detection of β-lactamase encoding genes in feces, soil and water from a Brazilian pig farm.

PubMed

Furlan, João Pedro Rueda; Stehling, Eliana Guedes

2018-01-10

β-lactam antibiotics are widely used for the treatment of different types of infections worldwide and the resistance to these antibiotics has grown sharply, which is of great concern. Resistance to β-lactams in gram-negative bacteria is mainly due to the production of β-lactamases, which are classified according to their functional activities. The aim of this study was to verify the presence of β-lactamases encoding genes in feces, soil, and water from a Brazilian pig farm. Different β-lactamases encoding genes were found, including bla CTX-M-Gp1 , bla CTX-M-Gp9 , bla SHV , bla OXA-1-like , bla GES , and bla VEB . The bla SHV and bla CTX-M-Gp1 genes have been detected in all types of samples, indicating the spread of β-lactam resistant bacteria among farm pigs and the environment around them. These results indicate that β-lactamase encoding genes belonging to the cloxacillinase, ESBL, and carbapenemase and they have high potential to spread in different sources, due to the fact that genes are closely related to mobile genetic elements, especially plasmids.

Major bleeding caused by warfarin in a genetically susceptible patient.

PubMed

Bloch, Aharon; Ben-Chetrit, Eldad; Muszkat, Mordechai; Caraco, Yoseph

2002-01-01

A 90-year-old woman was hospitalized for gastrointestinal bleeding. Although she had been receiving only warfarin 5 mg/day, her international normalized ratio (INR) was 66. Warfarin was discontinued, and her INR fell to 3.7 after transfusion of fresh-frozen plasma. However, it rose again spontaneously to 7.5. Eleven days after the last dose of warfarin had been administered, it was still detectable in the patient's plasma, indicating that impaired warfarin clearance may have caused an enhanced anticoagulation effect. Genetic analysis of the cytochrome P450 (CYP) isoenzyme 2C9, which mediates the major deactivating pathway of S-warfarin, revealed that the patient was a compound heterozygote carrying two variant alleles: CYP2C9*2 and CYP2C9*3. The patient's enhanced sensitivity to warfarin 5 mg/day can be ascribed to decreased clearance of S-warfarin secondary to genetic alteration of the gene encoding CYP2C9, resulting in a life-threatening complication.

S-adenosylhomocysteine hydrolase deficiency in a 26-year-old man.

PubMed

Buist, N R M; Glenn, B; Vugrek, O; Wagner, C; Stabler, S; Allen, R H; Pogribny, I; Schulze, A; Zeisel, S H; Barić, I; Mudd, S H

2006-08-01

This paper reports the third proven human case of deficient S-adenosylhomocysteine (AdoHcy) hydrolase activity. The patient is similar to the only two previously reported cases with this disorder in having severe myopathy, developmental delay, elevated serum creatine kinase (CK) concentrations, and hypermethioninaemia. Although he has been followed from infancy, the basic enzyme deficiency was established only at age 26 years. The diagnosis was based on markedly elevated plasma concentrations of both AdoHcy and S-adenosylmethionine, some 20% of the mean control activity of AdoHcy hydrolase activity in haemolysates of his red-blood cells, and two missense mutations in his gene encoding AdoHcy hydrolase. He had low values of erythrocyte phosphatidylcholine and plasma free choline and marginally elevated excretion of guanidinoacetate, suggesting that the elevated AdoHcy may have been inhibiting methylation of phosphatidylethanolamine and guanidinoacetate. His leukocyte DNA was globally more methylated than the DNA's of his parents or the mean extent of methylation measured in age-matched control subjects.

Deletion of the Mouse Slc30a8 Gene Encoding Zinc Transporter-8 Results in Impaired Insulin Secretion

PubMed Central

Pound, Lynley D.; Sarkar, Suparna; Benninger, Richard K. P.; Wang, Yingda; Suwanichkul, Adisak; Shadoan, Melanie K.; Printz, Richard L.; Oeser, James K.; Lee, Catherine E.; Piston, David W.; McGuinness, Owen P.; Hutton, John C.; Powell, David R.; O’Brien, Richard M.

2010-01-01

Synopsis The Slc30a8 gene encodes the islet-specific zinc transporter ZnT-8, which provides zinc for insulin-hexamer formation. Polymorphic variants in amino acid 325 of human ZnT-8 are associated with altered susceptibility to type 2 diabetes and ZnT-8 autoantibody epitope specificity changes in type 1 diabetes. To assess the physiological importance of ZnT-8, mice carrying a Slc30a8 exon 3 deletion were analyzed histologically and phenotyped for energy metabolism and pancreatic hormone secretion. No gross anatomical or behavioral changes or differences in body weight were observed between wild type and ZnT-8 −/− mice and ZnT-8 −/− mouse islets were indistinguishable from wild type in terms of their numbers, size and cellular composition. However, total zinc content was markedly reduced in ZnT-8 −/− mouse islets, as evaluated both by Timm’s histochemical staining of pancreatic sections and direct measurements in isolated islets. Blood glucose levels were unchanged in 16 week old, 6 hr fasted animals of either gender, however, plasma insulin concentrations were reduced in both female (~31%) and male (~47%) ZnT-8 −/− mice. Intraperitoneal glucose tolerance tests demonstrated no impairment in glucose clearance in male ZnT-8 −/− mice but glucose-stimulated insulin secretion from isolated islets was reduced ~33% relative to wild type littermates. In summary, Slc30a8 gene deletion is accompanied by a modest impairment in insulin secretion without major alterations in glucose metabolism. PMID:19450229

The molecular genetics of Usher syndrome.

PubMed

Ahmed, Z M; Riazuddin, S; Riazuddin, S; Wilcox, E R

2003-06-01

Association of sensorineural deafness and progressive retinitis pigmentosa with and without a vestibular abnormality is the hallmark of Usher syndrome and involves at least 12 loci among three different clinical subtypes. Genes identified for the more commonly inherited loci are USH2A (encoding usherin), MYO7A (encoding myosin VIIa), CDH23 (encoding cadherin 23), PCDH15 (encoding protocadherin 15), USH1C (encoding harmonin), USH3A (encoding clarin 1), and USH1G (encoding SANS). Transcripts from all these genes are found in many tissues/cell types other than the inner ear and retina, but all are uniquely critical for retinal and cochlear cell function. Many of these protein products have been demonstrated to have direct interactions with each other and perform an essential role in stereocilia homeostasis.

Cloning of the Pichia anomala SEC61 gene and its expression in a Saccharomyces cerevisiae sec61 mutant.

PubMed

Ruíz, Teresa; De la Rosa, José M; Domínguez, Angel; Rodríguez, Luis

2003-05-01

In several organisms, including Saccharomyces cerevisiae and other yeast species, the product encoded by the SEC61 gene is considered to be the core element of the translocation apparatus within the endoplasmic reticulum membrane through which translocation of secretory and membrane proteins occurs. In this study, we have cloned and characterized the homolog of the SEC61 gene from the yeast Pichia anomala. The cloned gene includes an ORF, interrupted after the first ten nucleotides by an intron of 131 bp, encoding a 479-amino acid putative polypeptide exhibiting homology to the products encoded by different eukaryotic SEC61 genes, particularly to those from other yeast species. We show that the P. anomala SEC61 gene is correctly processed (intron splicing) when expressed in S. cerevisiae and that it is able to complement the thermosensitive phenotype associated with a mutation in the S. cerevisiae SEC61 gene.

Identification of the gene for fly non-muscle myosin heavy chain: Drosophila myosin heavy chains are encoded by a gene family.

PubMed Central

Kiehart, D P; Lutz, M S; Chan, D; Ketchum, A S; Laymon, R A; Nguyen, B; Goldstein, L S

1989-01-01

In contrast to vertebrate species Drosophila has a single myosin heavy chain gene that apparently encodes all sarcomeric heavy chain polypeptides. Flies also contain a cytoplasmic myosin heavy chain polypeptide that by immunological and peptide mapping criteria is clearly different from the major thoracic muscle isoform. Here, we identify the gene that encodes this cytoplasmic isoform and demonstrate that it is distinct from the muscle myosin heavy chain gene. Thus, fly myosin heavy chains are the products of a gene family. Our data suggest that the contractile function required to power myosin based movement in non-muscle cells requires myosin diversity beyond that available in a single heavy chain gene. In addition, we show, that accumulation of cytoplasmic myosin transcripts is regulated in a developmental stage specific fashion, consistent with a key role for this protein in the movements of early embryogenesis. Images PMID:2498088

A novel chlorophyll a/b binding (Cab) protein gene from petunia which encodes the lower molecular weight Cab precursor protein.

PubMed

Stayton, M M; Black, M; Bedbrook, J; Dunsmuir, P

1986-12-22

The 16 petunia Cab genes which have been characterized are all closely related at the nucleotide sequence level and they encode Cab precursor polypeptides which are similar in sequence and length. Here we describe a novel petunia Cab gene which encodes a unique Cab precursor protein. This protein is a member of the smallest class of Cab precursor proteins for which no gene has previously been assigned in petunia or any other species. The features of this Cab precursor protein are that it is shorter by 2-3 amino acids than the formerly characterized Cab precursors, its transit peptide sequence is unrelated, and the mature polypeptide is significantly diverged at the functionally important N terminus from other petunia Cab proteins. Gene structure also discriminates this gene which is the only intron containing Cab gene in petunia genomic DNA.

Identification of chitinolytic bacteria isolated from shrimp pond sediment and characterization of their chitinase encoding gene

NASA Astrophysics Data System (ADS)

Triwijayani, A. U.; Puspita, I. D.; Murwantoko; Ustadi

2018-03-01

Chitinolytic bacteria are a group of bacteria owning enzymes that able to hydrolyze chitin. Previously, we isolated chitinolytic bacteria from shrimp pond sediment in Bantul, Yogyakarta, and obtained five isolates showing high chitinolytic index named as isolate PT1, PT2, PT5, PT6 and PB2. The aims of this study were to identify chitinolytic bacteria isolated from shrimp pond sediment and to characterize the chitinase encoding gene from each isolate. The molecular technique was performed by amplification of 16S rDNA, amplification of chitinase encoding gene and sequence analysis. Two chitinolytic bacteria of PT1 and PT2 were similar to Aeromonas bivalvium strain D15, PT5 to Pseudomonas stutzeri strain BD-2.2.1, PT6 to Serratia marcescens strain FZSF02 and PB2 to Streptomyces misionensis strain OsiRt-1. The comparison of chitinase encoding gene between three isolates with those in Gen Bank shows that PT1 had similar sequences with the chi1 gene in Aeromonas sp. 17m, PT2 with chi1 gene in A. caviae (CB101) and PT6 with chiB gene in S. Marcescens (BJL200).

Characterization of EhaJ, a New Autotransporter Protein from Enterohemorrhagic and Enteropathogenic Escherichia coli

PubMed Central

Easton, Donna M.; Totsika, Makrina; Allsopp, Luke P.; Phan, Minh-Duy; Idris, Adi; Wurpel, Daniël J.; Sherlock, Orla; Zhang, Bing; Venturini, Carola; Beatson, Scott A.; Mahony, Timothy J.; Cobbold, Rowland N.; Schembri, Mark A.

2011-01-01

Enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC) are diarrheagenic pathotypes of E. coli that cause gastrointestinal disease with the potential for life-threatening sequelae. While certain EHEC and EPEC virulence mechanisms have been extensively studied, the factors that mediate host colonization remain to be properly defined. Previously, we identified four genes (ehaA, ehaB, ehaC, and ehaD) from the prototypic EHEC strain EDL933 that encode for proteins that belong to the autotransporter (AT) family. Here we have examined the prevalence of these genes, as well as several other AT-encoding genes, in a collection of EHEC and EPEC strains. We show that the complement of AT-encoding genes in EHEC and EPEC strains is variable, with some AT-encoding genes being highly prevalent. One previously uncharacterized AT-encoding gene, which we have termed ehaJ, was identified in 12/44 (27%) of EHEC and 2/20 (10%) of EPEC strains. The ehaJ gene lies immediately adjacent to a gene encoding a putative glycosyltransferase (referred to as egtA). Western blot analysis using an EhaJ-specific antibody indicated that EhaJ is glycosylated by EgtA. Expression of EhaJ in a recombinant E. coli strain, revealed EhaJ is located at the cell surface and in the presence of the egtA glycosyltransferase gene mediates strong biofilm formation in microtiter plate and flow cell assays. EhaJ also mediated adherence to a range of extracellular matrix proteins, however this occurred independent of glycosylation. We also demonstrate that EhaJ is expressed in a wild-type EPEC strain following in vitro growth. However, deletion of ehaJ did not significantly alter its adherence or biofilm properties. In summary, EhaJ is a new glycosylated AT protein from EPEC and EHEC. Further studies are required to elucidate the function of EhaJ in colonization and virulence. PMID:21687429

Targeted next-generation sequencing helps to decipher the genetic and phenotypic heterogeneity of hypertrophic cardiomyopathy

PubMed Central

Cecconi, Massimiliano; Parodi, Maria I.; Formisano, Francesco; Spirito, Paolo; Autore, Camillo; Musumeci, Maria B.; Favale, Stefano; Forleo, Cinzia; Rapezzi, Claudio; Biagini, Elena; Davì, Sabrina; Canepa, Elisabetta; Pennese, Loredana; Castagnetta, Mauro; Degiorgio, Dario; Coviello, Domenico A.

2016-01-01

Hypertrophic cardiomyopathy (HCM) is mainly associated with myosin, heavy chain 7 (MYH7) and myosin binding protein C, cardiac (MYBPC3) mutations. In order to better explain the clinical and genetic heterogeneity in HCM patients, in this study, we implemented a target-next generation sequencing (NGS) assay. An Ion AmpliSeq™ Custom Panel for the enrichment of 19 genes, of which 9 of these did not encode thick/intermediate and thin myofilament (TTm) proteins and, among them, 3 responsible of HCM phenocopy, was created. Ninety-two DNA samples were analyzed by the Ion Personal Genome Machine: 73 DNA samples (training set), previously genotyped in some of the genes by Sanger sequencing, were used to optimize the NGS strategy, whereas 19 DNA samples (discovery set) allowed the evaluation of NGS performance. In the training set, we identified 72 out of 73 expected mutations and 15 additional mutations: the molecular diagnosis was achieved in one patient with a previously wild-type status and the pre-excitation syndrome was explained in another. In the discovery set, we identified 20 mutations, 5 of which were in genes encoding non-TTm proteins, increasing the diagnostic yield by approximately 20%: a single mutation in genes encoding non-TTm proteins was identified in 2 out of 3 borderline HCM patients, whereas co-occuring mutations in genes encoding TTm and galactosidase alpha (GLA) altered proteins were characterized in a male with HCM and multiorgan dysfunction. Our combined targeted NGS-Sanger sequencing-based strategy allowed the molecular diagnosis of HCM with greater efficiency than using the conventional (Sanger) sequencing alone. Mutant alleles encoding non-TTm proteins may aid in the complete understanding of the genetic and phenotypic heterogeneity of HCM: co-occuring mutations of genes encoding TTm and non-TTm proteins could explain the wide variability of the HCM phenotype, whereas mutations in genes encoding only the non-TTm proteins are identifiable in patients with a milder HCM status. PMID:27600940

Characterization of two genes encoding the Mycobacterium tuberculosis ribonucleotide reductase small subunit.

PubMed Central

Yang, F; Curran, S C; Li, L S; Avarbock, D; Graf, J D; Chua, M M; Lu, G; Salem, J; Rubin, H

1997-01-01

Two nrdF genes, nrdF1 and nrdF2, encoding the small subunit (R2) of ribonucleotide reductase (RR) from Mycobacterium tuberculosis have 71% identity at the amino acid level and are both highly homologous with Salmonella typhimurium R2F. The calculated molecular masses of R2-1 and R2-2 are 36,588 (322 amino acids [aa]) and 36,957 (324 aa) Da, respectively. Western blot analysis of crude M. tuberculosis extracts indicates that both R2s are expressed in vivo. Recombinant R2-2 is enzymatically active when assayed with pure recombinant M. tuberculosis R1 subunit. Both ATP and dATP are activators for CDP reduction up to 2 and 1 mM, respectively. The gene encoding M. tuberculosis R2-1, nrdF1, is not linked to nrdF2, nor is either gene linked to the gene encoding the large subunit, M. tuberculosis nrdE. The gene encoding MTP64 was found downstream from nrdF1, and the gene encoding alcohol dehydrogenase was found downstream from nrdF2. A nrdA(Ts) strain of E. coli (E101) could be complemented by simultaneous transformation with M. tuberculosis nrdE and nrdF2. An M. tuberculosis nrdF2 variant in which the codon for the catalytically necessary tyrosine was replaced by the phenylalanine codon did not complement E101 when cotransformed with M. tuberculosis nrdE. Similarly, M. tuberculosis nrdF1 and nrdE did not complement E101. Activity of recombinant M. tuberculosis RR was inhibited by incubating the enzyme with a peptide corresponding to the 7 C-terminal amino acid residues of the R2-2 subunit. M. tuberculosis is a species in which a nrdEF system appears to encode the biologically active species of RR and also the only bacterial species identified so far in which class I RR subunits are not arranged on an operon. PMID:9335290

Cloning and sequence analysis of the LEU2 homologue gene from Pichia anomala.

PubMed

De la Rosa, J M; Pérez, J A; Gutiérrez, F; González, J M; Ruiz, T; Rodríguez, L

2001-11-01

The Pichia anomala LEU2 gene (PaLEU2) was isolated by complementation of a leu2 Saccharomyces cerevisiae mutant. The cloned gene also allowed growth of a Escherichia coli leuB mutant in leucine-lacking medium, indicating that it encodes a product able to complement the beta-isopropylmalate dehydrogenase deficiency of the mutants. The sequenced DNA fragment contains a complete ORF of 1092 bp, and the deduced polypeptide shares significant homologies with the products of the LEU2 genes from S. cerevisiae (84% identity) and other yeast species. A sequence resembling the GC-rich palindrome motif identified in the 5' region of S. cerevisiae LEU2 gene as the binding site for the transcription activating factor encoded by the LEU3 gene was found at the promoter region. In addition, upstream of the PaLEU2 the 3'-terminal half of a gene of the same orientation, encoding a homologue of the S. cerevisiae NFS1/SPL1 gene that encodes a mitochondrial cysteine desulphurase involved in both tRNA processing and mitochondrial metabolism, was found. The genomic organization of the PaNFS1-PaLEU2 gene pair is similar to that found in several other yeast species, including S. cerevisiae and Candida albicans, except that in some of them the LEU2 gene appears in the reverse orientation. Copyright 2001 John Wiley & Sons, Ltd.

Asparaginase treatment side-effects may be due to genes with homopolymeric Asn codons (Review-Hypothesis)

PubMed Central

BANERJI, JULIAN

2015-01-01

The present treatment of childhood T-cell leukemias involves the systemic administration of prokary-otic L-asparaginase (ASNase), which depletes plasma Asparagine (Asn) and inhibits protein synthesis. The mechanism of therapeutic action of ASNase is poorly understood, as are the etiologies of the side-effects incurred by treatment. Protein expression from genes bearing Asn homopolymeric coding regions (N-hCR) may be particularly susceptible to Asn level fluctuation. In mammals, N-hCR are rare, short and conserved. In humans, misfunctions of genes encoding N-hCR are associated with a cluster of disorders that mimic ASNase therapy side-effects which include impaired glycemic control, dislipidemia, pancreatitis, compromised vascular integrity, and neurological dysfunction. This paper proposes that dysregulation of Asn homeostasis, potentially even by ASNase produced by the microbiome, may contribute to several clinically important syndromes by altering expression of N-hCR bearing genes. By altering amino acid abundance and modulating ribosome translocation rates at codon repeats, the microbiomic environment may contribute to genome decoding and to shaping the proteome. We suggest that impaired translation at poly Asn codons elevates diabetes risk and severity. PMID:26178806

Asparaginase treatment side-effects may be due to genes with homopolymeric Asn codons (Review-Hypothesis).

PubMed

Banerji, Julian

2015-09-01

The present treatment of childhood T-cell leukemias involves the systemic administration of prokaryotic L-asparaginase (ASNase), which depletes plasma Asparagine (Asn) and inhibits protein synthesis. The mechanism of therapeutic action of ASNase is poorly understood, as are the etiologies of the side-effects incurred by treatment. Protein expression from genes bearing Asn homopolymeric coding regions (N-hCR) may be particularly susceptible to Asn level fluctuation. In mammals, N-hCR are rare, short and conserved. In humans, misfunctions of genes encoding N-hCR are associated with a cluster of disorders that mimic ASNase therapy side-effects which include impaired glycemic control, dislipidemia, pancreatitis, compromised vascular integrity, and neurological dysfunction. This paper proposes that dysregulation of Asn homeostasis, potentially even by ASNase produced by the microbiome, may contribute to several clinically important syndromes by altering expression of N-hCR bearing genes. By altering amino acid abundance and modulating ribosome translocation rates at codon repeats, the microbiomic environment may contribute to genome decoding and to shaping the proteome. We suggest that impaired translation at poly Asn codons elevates diabetes risk and severity.

Treatment of Diabetes and Long-Term Survival After Insulin and Glucokinase Gene Therapy

PubMed Central

Callejas, David; Mann, Christopher J.; Ayuso, Eduard; Lage, Ricardo; Grifoll, Iris; Roca, Carles; Andaluz, Anna; Ruiz-de Gopegui, Rafael; Montané, Joel; Muñoz, Sergio; Ferre, Tura; Haurigot, Virginia; Zhou, Shangzhen; Ruberte, Jesús; Mingozzi, Federico; High, Katherine A.; Garcia, Felix; Bosch, Fatima

2013-01-01

Diabetes is associated with severe secondary complications, largely caused by poor glycemic control. Treatment with exogenous insulin fails to prevent these complications completely, leading to significant morbidity and mortality. We previously demonstrated that it is possible to generate a “glucose sensor” in skeletal muscle through coexpression of glucokinase and insulin, increasing glucose uptake and correcting hyperglycemia in diabetic mice. Here, we demonstrate long-term efficacy of this approach in a large animal model of diabetes. A one-time intramuscular administration of adeno-associated viral vectors of serotype 1 encoding for glucokinase and insulin in diabetic dogs resulted in normalization of fasting glycemia, accelerated disposal of glucose after oral challenge, and no episodes of hypoglycemia during exercise for >4 years after gene transfer. This was associated with recovery of body weight, reduced glycosylated plasma proteins levels, and long-term survival without secondary complications. Conversely, exogenous insulin or gene transfer for insulin or glucokinase alone failed to achieve complete correction of diabetes, indicating that the synergistic action of insulin and glucokinase is needed for full therapeutic effect. This study provides the first proof-of-concept in a large animal model for a gene transfer approach to treat diabetes. PMID:23378612

Genetic variants in a haplotype block spanning IDE are significantly associated with plasma Abeta42 levels and risk for Alzheimer disease.

PubMed

Ertekin-Taner, Nilüfer; Allen, Mariet; Fadale, Daniel; Scanlin, Leah; Younkin, Linda; Petersen, Ronald C; Graff-Radford, Neill; Younkin, Steven G

2004-04-01

Risk for late onset Alzheimer disease (LOAD) and plasma amyloid beta levels (Abeta42; encoded by APP), an intermediate phenotype for LOAD, show linkage to chromosome 10q. Several strong candidate genes (VR22, PLAU, IDE) lie within the 1-lod support interval for linkage. Others have independently identified haplotypes in the chromosome 10q region harboring IDE that show highly significant association with intermediate AD phenotypes and with risk for AD. To pursue these associations, we analyzed the same haplotypes for association with plasma Abeta42 in 24 extended LOAD families and for association with LOAD in two independent case-control series. One series (MCR, 188 age-matched case-control pairs) did not show association (p=0.64) with the six haplotypes in the 276-kb region spanning three genes (IDE, KNSL1, and HHEX) previously shown to associate with LOAD. The other series (MCJ, 109 age-matched case-control pairs) showed significant (p=0.003) association with these haplotypes. In the MCJ series, the H4 (odds ratio [OR]=5.1, p=0.003) and H2(H7) haplotypes (OR=0.60, p=0.04) had the same effects previously reported. In this series, the H8 haplotype (OR=2.7, p=0.098) also had an effect similar as in one previous case control series but not in others. In the extended families, the H8 haplotype was associated with significantly elevated plasma Abeta42 (p=0.02). In addition, the H5(H10) haplotype, which is associated with reduced risk for AD in the other study is associated with reduced plasma Abeta42 (p=0.007) in our family series. These results provide strong evidence for pathogenic variant(s) in the 276-kb region harboring IDE that influence intermediate AD phenotypes and risk for AD. Copyright 2004 Wiley-Liss, Inc.

Constitutive activation of a plasma membrane H(+)-ATPase prevents abscisic acid-mediated stomatal closure.

PubMed

Merlot, Sylvain; Leonhardt, Nathalie; Fenzi, Francesca; Valon, Christiane; Costa, Miguel; Piette, Laurie; Vavasseur, Alain; Genty, Bernard; Boivin, Karine; Müller, Axel; Giraudat, Jérôme; Leung, Jeffrey

2007-07-11

Light activates proton (H(+))-ATPases in guard cells, to drive hyperpolarization of the plasma membrane to initiate stomatal opening, allowing diffusion of ambient CO(2) to photosynthetic tissues. Light to darkness transition, high CO(2) levels and the stress hormone abscisic acid (ABA) promote stomatal closing. The overall H(+)-ATPase activity is diminished by ABA treatments, but the significance of this phenomenon in relationship to stomatal closure is still debated. We report two dominant mutations in the OPEN STOMATA2 (OST2) locus of Arabidopsis that completely abolish stomatal response to ABA, but importantly, to a much lesser extent the responses to CO(2) and darkness. The OST2 gene encodes the major plasma membrane H(+)-ATPase AHA1, and both mutations cause constitutive activity of this pump, leading to necrotic lesions. H(+)-ATPases have been traditionally assumed to be general endpoints of all signaling pathways affecting membrane polarization and transport. Our results provide evidence that AHA1 is a distinct component of an ABA-directed signaling pathway, and that dynamic downregulation of this pump during drought is an essential step in membrane depolarization to initiate stomatal closure.

Exogenous fatty acids and niacin on acute prostaglandin D2 production in human myeloid cells.

PubMed

Montserrat-de la Paz, Sergio; Bermudez, Beatriz; Lopez, Sergio; Naranjo, Maria C; Romero, Yolanda; Bando-Hidalgo, Maria J; Abia, Rocio; Muriana, Francisco J G

2017-01-01

Niacin activates HCA 2 receptor that results in the release of PGD 2 . However, little is known on PGD 2 -producing cells and the role of fatty acids. Notably M-CSF macrophages exhibited a timely dependent PGD 2 production upon niacin challenge. Short pretreatment of M-CSF macrophages with autologous postprandial TRLs induced the down-regulation of HCA 2 gene and up-regulation of genes encoding COX1 and COX2 enzymes in a fatty acid-dependent manner. These effects were paralleled by a higher PGD 2 production with postprandial TRL-SFAs. The niacin-mediated transcriptional activity of all genes involved in PGD 2 biosynthesis was desensitized in a time-dependent manner by postprandial TRLs, leading to a decreased PGD 2 release. In vivo, the net excursions of PGD 2 in plasma followed similar fatty acid-dependent patterns as those found for PGD 2 release in vitro. The predominant fatty acid class in the diet acutely modulates PGD 2 biosynthetic pathway both in vitro and in vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

A Different Microbiome Gene Repertoire in the Airways of Cystic Fibrosis Patients with Severe Lung Disease

PubMed Central

Bacci, Giovanni; Fiscarelli, Ersilia; Taccetti, Giovanni; Dolce, Daniela; Paganin, Patrizia; Morelli, Patrizia; Tuccio, Vanessa; De Alessandri, Alessandra; Lucidi, Vincenzina

2017-01-01

In recent years, next-generation sequencing (NGS) was employed to decipher the structure and composition of the microbiota of the airways in cystic fibrosis (CF) patients. However, little is still known about the overall gene functions harbored by the resident microbial populations and which specific genes are associated with various stages of CF lung disease. In the present study, we aimed to identify the microbial gene repertoire of CF microbiota in twelve patients with severe and normal/mild lung disease by performing sputum shotgun metagenome sequencing. The abundance of metabolic pathways encoded by microbes inhabiting CF airways was reconstructed from the metagenome. We identified a set of metabolic pathways differently distributed in patients with different pulmonary function; namely, pathways related to bacterial chemotaxis and flagellar assembly, as well as genes encoding efflux-mediated antibiotic resistance mechanisms and virulence-related genes. The results indicated that the microbiome of CF patients with low pulmonary function is enriched in virulence-related genes and in genes encoding efflux-mediated antibiotic resistance mechanisms. Overall, the microbiome of severely affected adults with CF seems to encode different mechanisms for the facilitation of microbial colonization and persistence in the lung, consistent with the characteristics of multidrug-resistant microbial communities that are commonly observed in patients with severe lung disease. PMID:28758937

Activation of Pathogenesis-related Genes by the Rhizobacterium, Bacillus sp. JS, Which Induces Systemic Resistance in Tobacco Plants.

PubMed

Kim, Ji-Seong; Lee, Jeongeun; Lee, Chan-Hui; Woo, Su Young; Kang, Hoduck; Seo, Sang-Gyu; Kim, Sun-Hyung

2015-06-01

Plant growth promoting rhizobacteria (PGPR) are known to confer disease resistance to plants. Bacillus sp. JS demonstrated antifungal activities against five fungal pathogens in in vitro assays. To verify whether the volatiles of Bacillus sp. JS confer disease resistance, tobacco leaves pre-treated with the volatiles were damaged by the fungal pathogen, Rhizoctonia solani and oomycete Phytophthora nicotianae. Pre-treated tobacco leaves had smaller lesion than the control plant leaves. In pathogenesis-related (PR) gene expression analysis, volatiles of Bacillus sp. JS caused the up-regulation of PR-2 encoding β-1,3-glucanase and acidic PR-3 encoding chitinase. Expression of acidic PR-4 encoding chitinase and acidic PR-9 encoding peroxidase increased gradually after exposure of the volatiles to Bacillus sp. JS. Basic PR-14 encoding lipid transfer protein was also increased. However, PR-1 genes, as markers of salicylic acid (SA) induced resistance, were not expressed. These results suggested that the volatiles of Bacillus sp. JS confer disease resistance against fungal and oomycete pathogens through PR genes expression.

Fusagene vectors: a novel strategy for the expression of multiple genes from a single cistron.

PubMed

Gäken, J; Jiang, J; Daniel, K; van Berkel, E; Hughes, C; Kuiper, M; Darling, D; Tavassoli, M; Galea-Lauri, J; Ford, K; Kemeny, M; Russell, S; Farzaneh, F

2000-12-01

Transduction of cells with multiple genes, allowing their stable and co-ordinated expression, is difficult with the available methodologies. A method has been developed for expression of multiple gene products, as fusion proteins, from a single cistron. The encoded proteins are post-synthetically cleaved and processed into each of their constituent proteins as individual, biologically active factors. Specifically, linkers encoding cleavage sites for the Golgi expressed endoprotease, furin, have been incorporated between in-frame cDNA sequences encoding different secreted or membrane bound proteins. With this strategy we have developed expression vectors encoding multiple proteins (IL-2 and B7.1, IL-4 and B7.1, IL-4 and IL-2, IL-12 p40 and p35, and IL-12 p40, p35 and IL-2 ). Transduction and analysis of over 100 individual clones, derived from murine and human tumour cell lines, demonstrate the efficient expression and biological activity of each of the encoded proteins. Fusagene vectors enable the co-ordinated expression of multiple gene products from a single, monocistronic, expression cassette.

Structure, Expression, Chromosomal Location and Product of the Gene Encoding Adh2 in Petunia

PubMed Central

Gregerson, R. G.; Cameron, L.; McLean, M.; Dennis, P.; Strommer, J.

1993-01-01

In most higher plants the genes encoding alcohol dehydrogenase comprise a small gene family, usually with two members. The Adh1 gene of Petunia has been cloned and analyzed, but a second identifiable gene was not recovered from any of three genomic libraries. We have therefore employed the polymerase chain reaction to obtain the major portion of a second Adh gene. From sequence, mapping and northern data we conclude this gene encodes ADH2, the major anaerobically inducible Adh gene of Petunia. The availability of both Adh1 and Adh2 from Petunia has permitted us to compare their structures and patterns of expression to those of the well-studied Adh genes of maize, of which one is highly expressed developmentally, while both are induced in response to hypoxia. Despite their evolutionary distance, evidenced by deduced amino acid sequence as well as taxonomic classification, the pairs of genes are regulated in strikingly similar ways in maize and Petunia. Our findings suggest a significant biological basis for the regulatory strategy employed by these distant species for differential expression of multiple Adh genes. PMID:8096485

Regulation of Bacteriocin Production in Streptococcus mutans by the Quorum-Sensing System Required for Development of Genetic Competence

PubMed Central

van der Ploeg, Jan R.

2005-01-01

In Streptococcus mutans, competence for genetic transformation and biofilm formation are dependent on the two-component signal transduction system ComDE together with the inducer peptide pheromone competence-stimulating peptide (CSP) (encoded by comC). Here, it is shown that the same system is also required for expression of the nlmAB genes, which encode a two-peptide nonlantibiotic bacteriocin. Expression from a transcriptional nlmAB′-lacZ fusion was highest at high cell density and was increased up to 60-fold following addition of CSP, but it was abolished when the comDE genes were interrupted. Two more genes, encoding another putative bacteriocin and a putative bacteriocin immunity protein, were also regulated by this system. The regions upstream of these genes and of two further putative bacteriocin-encoding genes and a gene encoding a putative bacteriocin immunity protein contained a conserved 9-bp repeat element just upstream of the transcription start, which suggests that expression of these genes is also dependent on the ComCDE regulatory system. Mutations in the repeat element of the nlmAB promoter region led to a decrease in CSP-dependent expression of nlmAB′-lacZ. In agreement with these results, a comDE mutant and mutants unable to synthesize or export CSP did not produce bacteriocins. It is speculated that, at high cell density, bacteriocin production is induced to liberate DNA from competing streptococci. PMID:15937160

Functional characterization of an apple (Malus x domestica) LysM domain receptor encoding gene for its role in defense response

USDA-ARS?s Scientific Manuscript database

Apple gene MDP0000136494 was identified as the only LysM containing protein encoding gene which was specifically up-regulated in P. ultimum infected apple root by a previous transcriptome analysis. In current study, the functional identity of MDP0000136494 was investigated using combined genomic, tr...

Cloning and sequencing the genes encoding goldfish and carp ependymin.

PubMed

Adams, D S; Shashoua, V E

1994-04-20

Ependymins (EPNs) are brain glycoproteins thought to function in optic nerve regeneration and long-term memory consolidation. To date, epn genes have been characterized in two orders of teleost fish. In this study, polymerase chain reactions (PCR) were used to amplify the complete 1.6-kb epn genes, gf-I and cc-I, from genomic DNA of Cypriniformes, goldfish and carp, respectively. Amplified bands were cloned and sequenced. Each gene consists of six exons and five introns. The exon portion of gf-I encodes a predicted 215-amino-acid (aa) protein previously characterized as GF-I, while cc-I encodes a predicted 215-aa protein 95% homologous to GF-I.

The complete mitochondrial genome sequence of Eimeria innocua (Eimeriidae, Coccidia, Apicomplexa).

PubMed

Hafeez, Mian Abdul; Vrba, Vladimir; Barta, John Robert

2016-07-01

The complete mitochondrial genome of Eimeria innocua KR strain (Eimeriidae, Coccidia, Apicomplexa) was sequenced. This coccidium infects turkeys (Meleagris gallopavo), Bobwhite quails (Colinus virginianus), and Grey partridges (Perdix perdix). Genome organization and gene contents were comparable with other Eimeria spp. infecting galliform birds. The circular-mapping mt genome of E. innocua is 6247 bp in length with three protein-coding genes (cox1, cox3, and cytb), 19 gene fragments encoding large subunit (LSU) rRNA and 14 gene fragments encoding small subunit (SSU) rRNA. Like other Apicomplexa, no tRNA was encoded. The mitochondrial genome of E. innocua confirms its close phylogenetic affinities to Eimeria dispersa.

Genes encoding novel lipid transporters and their use to increase oil production in vegetative tissues of plants

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Changcheng; Fan, Jilian; Yan, Chengshi

The present invention discloses a novel gene encoding a transporter protein trigalactosyldiacylglycerol-5 (TGD5), mutations thereof and their use to enhance TAG production and retention in plant vegetative tissue.

Isolation and characterization of polygalacturonase genes (pecA and pecB) from Aspergillus flavus.

PubMed Central

Whitehead, M P; Shieh, M T; Cleveland, T E; Cary, J W; Dean, R A

1995-01-01

Two genes, pecA and pecB, encoding endopolyglacturonases were cloned from a highly aggressive strain of Aspergillus flavus. The pecA gene consisted of 1,228 bp encoding a protein of 363 amino acids with a predicted molecular mass of 37.6 kDa, interrupted by two introns of 58 and 81 bp in length. Accumulation of pecA mRNA in both pectin- or glucose-grown mycelia in the highly aggressive strain matched the activity profile of a pectinase previously identified as P2c. Transformants of a weakly aggressive strain containing a functional copy of the pecA gene produced P2c in vitro, confirming that pecA encodes P2c. The coding region of pecB was determined to be 1,217 bp in length interrupted by two introns of 65 and 54 bp in length. The predicted protein of 366 amino acids had an estimated molecular mass of 38 kDa. Transcripts of this gene accumulated in mycelia grown in medium containing pectin alone, never in mycelia grown in glucose-containing medium, for both highly and weakly aggressive strains. Thus, pecB encodes the activity previously identified as P1 or P3. pecA and pecB share a high degree of sequence identity with polygalacturonase genes from Aspergillus parasiticus and Aspergillus oryzae, further establishing the close relationships between members of the A. flavus group. Conservation of intron positions in these genes also indicates that they share a common ancestor with genes encoding endopolyglacturonases of Aspergillus niger. PMID:7574642

The association of eight potentially functional polymorphisms in five adrenergic receptor-encoding genes with myocardial infarction risk in Han Chinese.

PubMed

Xia, Kun; Ding, Rongjing; Zhang, Zhiyong; Li, Weiming; Shang, Xiaoming; Yang, Xinchun; Wang, Lefeng; Zhang, Qi

2017-08-15

Adrenergic receptors play a key role in activating the sympathetic nervous system, which often accompanies with the development of myocardial infarction (MI). Here, we aimed to test the association of eight potentially functional polymorphisms in five adrenergic receptor-encoding genes with MI risk. Genotypes were available for 717 MI patients and 612 controls. There were no detectable deviations from the Hardy-Weinberg equilibrium for all study polymorphisms. Allele frequencies differed remarkably for ADRA2B D/I (P<0.001), ADRB1 Ser49Gly (P=0.002), ADRB2 Gln27Glu (P=0.005), and ADRB3 Trp64Arg (P<0.001) polymorphisms, even after the Bonferroni correction. Systolic blood pressure was significantly lower in ADRA2B II genotype carriers than in the DD genotype carriers (P=0.006), while plasma high-density lipoprotein cholesterol was significantly higher in patients carrying ADRA2B I allele and ADRB1 49Ser allele than in patients with the DD genotype and 49Gly/49Gly genotype, respectively (P=0.018 and 0.033). Overall best interaction model consisted of ADRA2B D/I, ADRB1 Ser49Gly, dyslipidemia and hypertension, with the highest testing accuracy of 0.627 and the maximal 10-fold cross-validation consistency (P=0.017). Finally, a nomogram was depicted based on four significant polymorphisms and metabolic risk factors, and it had a better predictive utility and was internally validated with a discrimination C-index of 0.723 (P<0.001). Altogether, we identified two polymorphisms, ADRA2B D/I and ADRB1 Ser49Arg, which not only altered genetic susceptibility to MI, but also impacted on blood pressure and plasma lipid changes, and their combination with metabolic risk factors constituted the overall best interaction model. Copyright © 2017. Published by Elsevier B.V.

The Arabidopsis polyamine transporter LHR1/PUT3 modulates heat responsive gene expression by enhancing mRNA stability.

PubMed

Shen, Yun; Ruan, Qingxia; Chai, Haoxi; Yuan, Yongze; Yang, Wannian; Chen, Junping; Xin, Zhanguo; Shi, Huazhong

2016-12-01

Polyamines involve in gene regulation by interacting with and modulating the functions of various anionic macromolecules such as DNA, RNA and proteins. In this study, we identified an important function of the polyamine transporter LHR1 (LOWER EXPRESSION OF HEAT RESPONSIVE GENE1) in heat-inducible gene expression in Arabidopsis thaliana. The lhr1 mutant was isolated through a forward genetic screening for altered expression of the luciferase reporter gene driven by the promoter from the heat-inducible gene AtHSP18.2. The lhr1 mutant showed reduced induction of the luciferase gene in response to heat stress and was more sensitive to high temperature than the wild type. Map-based cloning identified that the LHR1 gene encodes the polyamine transporter PUT3 (POLYAMINE UPTAKE TRANSPORTER 3) localized in the plasma membrane. The LHR1/PUT3 is required for the uptake of extracellular polyamines and plays an important role in stabilizing the mRNAs of several crucial heat stress responsive genes under high temperature. Genome-wide gene expression analysis using RNA-seq identified an array of differentially expressed genes, among which the transcript levels of some of the heat shock protein genes significantly reduced in response to prolonged heat stress in the lhr1 mutant. Our findings revealed an important heat stress response and tolerance mechanism involving polyamine influx which modulates mRNA stability of heat-inducible genes under heat stress conditions. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Draft Genome Sequence of Ezakiella peruensis Strain M6.X2, a Human Gut Gram-Positive Anaerobic Coccus.

PubMed

Diop, Awa; Diop, Khoudia; Tomei, Enora; Raoult, Didier; Fenollar, Florence; Fournier, Pierre-Edouard

2018-03-01

We report here the draft genome sequence of Ezakiella peruensis strain M6.X2 T The draft genome is 1,672,788 bp long and harbors 1,589 predicted protein-encoding genes, including 26 antibiotic resistance genes with 1 gene encoding vancomycin resistance. The genome also exhibits 1 clustered regularly interspaced short palindromic repeat region and 333 genes acquired by horizontal gene transfer. Copyright © 2018 Diop et al.

Deregulation of Rab and Rab Effector Genes in Bladder Cancer

PubMed Central

Ho, Joel R.; Chapeaublanc, Elodie; Kirkwood, Lisa; Nicolle, Remy; Benhamou, Simone; Lebret, Thierry; Allory, Yves; Southgate, Jennifer; Radvanyi, François; Goud, Bruno

2012-01-01

Growing evidence indicates that Rab GTPases, key regulators of intracellular transport in eukaryotic cells, play an important role in cancer. We analysed the deregulation at the transcriptional level of the genes encoding Rab proteins and Rab-interacting proteins in bladder cancer pathogenesis, distinguishing between the two main progression pathways so far identified in bladder cancer: the Ta pathway characterized by a high frequency of FGFR3 mutation and the carcinoma in situ pathway where no or infrequent FGFR3 mutations have been identified. A systematic literature search identified 61 genes encoding Rab proteins and 223 genes encoding Rab-interacting proteins. Transcriptomic data were obtained for normal urothelium samples and for two independent bladder cancer data sets corresponding to 152 and 75 tumors. Gene deregulation was analysed with the SAM (significant analysis of microarray) test or the binomial test. Overall, 30 genes were down-regulated, and 13 were up-regulated in the tumor samples. Five of these deregulated genes (LEPRE1, MICAL2, RAB23, STXBP1, SYTL1) were specifically deregulated in FGFR3-non-mutated muscle-invasive tumors. No gene encoding a Rab or Rab-interacting protein was found to be specifically deregulated in FGFR3-mutated tumors. Cluster analysis showed that the RAB27 gene cluster (comprising the genes encoding RAB27 and its interacting partners) was deregulated and that this deregulation was associated with both pathways of bladder cancer pathogenesis. Finally, we found that the expression of KIF20A and ZWINT was associated with that of proliferation markers and that the expression of MLPH, MYO5B, RAB11A, RAB11FIP1, RAB20 and SYTL2 was associated with that of urothelial cell differentiation markers. This systematic analysis of Rab and Rab effector gene deregulation in bladder cancer, taking relevant tumor subgroups into account, provides insight into the possible roles of Rab proteins and their effectors in bladder cancer pathogenesis. This approach is applicable to other group of genes and types of cancer. PMID:22724020

Differences in virulence genes and genome patterns of mastitis-associated Staphylococcus aureus among goat, cow, and human isolates in Taiwan.

PubMed

Chu, Chishih; Wei, Yajiun; Chuang, Shih-Te; Yu, Changyou; Changchien, Chih-Hsuan; Su, Yaochi

2013-03-01

A total of 117 mastitis-associated Staphylococcus aureus isolates from cow, goat, and human patients were analyzed for differences in antibiotic susceptibility, virulence genes, and genotypes using accessory gene regulator (agr) typing, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Multidrug-resistant (MDR) S. aureus were commonly found in all sources, though they were predominantly found in human and goat isolates. Almost 70% of the isolates were resistant to ampicillin and penicillin. Host-associated virulence genes were identified as follows: tst, a gene encoding toxic shock syndrome toxin, was found in goat isolates; lukED and lukM, genes encoding leukocidin, found in cow isolates; lukPV, a gene encoding leukocidin, found in human isolates; and eta, a gene encoding for exfoliative toxin, found in both human and cow isolates. All four types of hemolysin, α, β, γ, and δ, were identified in human isolates, three types (α, γ, and δ), were identified in cow isolates, and two types (α and δ) were identified in goat isolates. Agr-typing determined agr1 to be the main subtype in human and cow isolates. PFGE and MLST analysis revealed the presence of diverse genotypes among the three sources. In conclusion, mastitis-associated, genetically diverse strains of MDR S. aureus differed in virulence genes among human, cow, and goat isolates.

Functions of the Magnaporthe oryzae Flb3p and Flb4p transcription factors in the regulation of conidiation.

PubMed

Matheis, S; Yemelin, A; Scheps, D; Andresen, K; Jacob, S; Thines, E; Foster, A J

2017-03-01

The Magnaporthe oryzae genes FLB3 and FLB4, orthologues of the Aspergillus nidulans regulators of conidiation FlbC and FlbD, were inactivated. These genes encode C2H2 zinc finger and Myb-like transcription factors, respectively, in A. nidulans. Analysis of the resultant mutants demonstrated that FLB4 is essential for spore formation and that strains lacking this gene are fluffy in their colony morphology due to an inability to complete conidiophore formation. Meanwhile, FLB3 is required for normal levels of aerial mycelium formation. We identified genes dependent on both transcription factors using microarray analysis. This analysis revealed that the transcription of several genes encoding proteins implicated in sporulation in Magnaporthe oryzae and other filamentous fungi are affected by FLB3 or FLB4 inactivation. Furthermore, the microarray analysis indicates that Flb3p may effectively reprogramme the cell metabolically by repressing transcription of genes encoding biosynthetic enzymes and inducing transcription of genes encoding catabolic enzymes. Additionally, qRT-PCR was employed and showed that FLB3 and FLB4 transcripts are enriched in synchronously sporulating cultures, as were the transcripts of other genes that are necessary for normal conidiation, consistent with a role for their gene products in this process. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

Ornithorhynchus anatinus (platypus) links the evolution of immunoglobulin genes in eutherian mammals and nonmammalian tetrapods.

PubMed

Zhao, Yaofeng; Cui, Huiting; Whittington, Camilla M; Wei, Zhiguo; Zhang, Xiaofeng; Zhang, Ziding; Yu, Li; Ren, Liming; Hu, Xiaoxiang; Zhang, Yaping; Hellman, Lars; Belov, Katherine; Li, Ning; Hammarström, Lennart

2009-09-01

The evolutionary origins of mammalian immunoglobulin H chain isotypes (IgM, IgD, IgG, IgE, and IgA) are still incompletely understood as these isotypes differ considerably in structure and number from their counterparts in nonmammalian tetrapods. We report in this study that the platypus (Ornithorhynchus anatinus) Ig H chain constant region gene locus contains eight Ig encoding genes, which are arranged in an mu-delta-omicron-gamma2-gamma1-alpha1-epsilon-alpha2 order, spanning a total of approximately 200 kb DNA, encoding six distinct isotypes. The omicron (omicron for Ornithorhynchus) gene encodes a novel Ig H chain isotype that consists of four constant region domains and a hinge, and is structurally different from any of the five known mammalian Ig classes. This gene is phylogenetically related to upsilon (epsilon) and gamma, and thus appears to be a structural intermediate between these two genes. The platypus delta gene encodes ten heavy chain constant region domains, lacks a hinge region and is similar to IgD in amphibians and fish, but strikingly different from that in eutherian mammals. The platypus Ig H chain isotype repertoire thus shows a unique combination of genes that share similarity both to those of nonmammalian tetrapods and eutherian animals and demonstrates how phylogenetically informative species can be used to reconstruct the evolutionary history of functionally important genes.

Development of Broadly Neutralizing Antibodies and Their Mapping by Monomeric gp120 in Human Immunodeficiency Virus Type 1-Infected Humans and Simian-Human Immunodeficiency Virus SHIVSF162P3N-Infected Macaques.

PubMed

Jia, Manxue; Lu, Hong; Markowitz, Martin; Cheng-Mayer, Cecilia; Wu, Xueling

2016-04-01

To improve our understanding of the similarities and differences between neutralizing antibodies elicited by simian-human immunodeficiency virus (SHIV)-infected rhesus macaques and human immunodeficiency virus type 1 (HIV-1)-infected humans, we examined the plasma of 13 viremic macaques infected with SHIVSF162P3Nand 85 HIV-1-infected humans with known times of infection. We identified 5 macaques (38%) from 1 to 2 years postinfection (p.i.) with broadly neutralizing antibodies (bnAbs) against tier 2 HIV-1. In comparison, only 2 out of 42 (5%) human plasma samples collected in a similar time frame of 1 to 3 years p.i. exhibited comparable neutralizing breadths and potencies, with the number increasing to 7 out of 21 (30%) after 3 years p.i. Plasma mapping with monomeric gp120 identified only 2 out of 9 humans and 2 out of 4 macaques that contained gp120-reactive neutralizing antibodies, indicating distinct specificities in these plasma samples, with most of them recognizing the envelope trimer (including gp41) rather than the gp120 monomer. Indeed, a total of 20 gp120-directed monoclonal antibodies (MAbs) isolated from a human subject (AD358) and a Chinese rhesus macaque (GB40) displayed no or limited neutralizing activity against tier 2 strains. These isolated MAbs, mapped to the CD4-binding site, the V3 loop, the inner domain, and the C5 region of gp120, revealed genetic similarity between the human and macaque immunoglobulin genes used to encode some V3-directed MAbs. These results also support the use of envelope trimer probes for efficient isolation of HIV-1 bnAbs. HIV-1 vaccine research can benefit from understanding the development of broadly neutralizing antibodies (bnAbs) in rhesus macaques, commonly used to assess vaccine immunogenicity and efficacy. Here, we examined 85 HIV-1-infected humans and 13 SHIVSF162P3N-infected macaques for bnAbs and found that, similar to HIV-1-infected humans, bnAbs in SHIV-infected macaques are also rare, but their development might have been faster in some of the studied macaques. Plasma mapping with monomeric gp120 indicated that most bnAbs bind to the envelope trimer rather than the gp120 monomer. In support of this, none of the isolated gp120-reactive monoclonal antibodies (MAbs) displayed the neutralization breadth observed in the corresponding plasma. However, the MAb sequences revealed similarity between human and macaque genes used to encode some V3-directed MAbs. Our study sheds light on the timing and development of bnAbs in SHIV-infected macaques in comparison to HIV-1-infected humans and highlights the use of envelope trimer probes for efficient recovery of bnAbs. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

The bglA Gene of Aspergillus kawachii Encodes Both Extracellular and Cell Wall-Bound β-Glucosidases

PubMed Central

Iwashita, Kazuhiro; Nagahara, Tatsuya; Kimura, Hitoshi; Takano, Makoto; Shimoi, Hitoshi; Ito, Kiyoshi

1999-01-01

We cloned the genomic DNA and cDNA of bglA, which encodes β-glucosidase in Aspergillus kawachii, based on a partial amino acid sequence of purified cell wall-bound β-glucosidase CB-1. The nucleotide sequence of the cloned bglA gene revealed a 2,933-bp open reading frame with six introns that encodes an 860-amino-acid protein. Based on the deduced amino acid sequence, we concluded that the bglA gene encodes cell wall-bound β-glucosidase CB-1. The amino acid sequence exhibited high levels of homology with the amino acid sequences of fungal β-glucosidases classified in subfamily B. We expressed the bglA cDNA in Saccharomyces cerevisiae and detected the recombinant β-glucosidase in the periplasm fraction of the recombinant yeast. A. kawachii can produce two extracellular β-glucosidases (EX-1 and EX-2) in addition to the cell wall-bound β-glucosidase. A. kawachii in which the bglA gene was disrupted produced none of the three β-glucosidases, as determined by enzyme assays and a Western blot analysis. Thus, we concluded that the bglA gene encodes both extracellular and cell wall-bound β-glucosidases in A. kawachii. PMID:10584016

Dopa-decarboxylase gene polymorphisms affect the motor response to L-dopa in Parkinson's disease.

PubMed

Devos, David; Lejeune, Stéphanie; Cormier-Dequaire, Florence; Tahiri, Khadija; Charbonnier-Beaupel, Fanny; Rouaix, Nathalie; Duhamel, Alain; Sablonnière, Bernard; Bonnet, Anne-Marie; Bonnet, Cecilia; Zahr, Noel; Costentin, Jean; Vidailhet, Marie; Corvol, Jean-Christophe

2014-02-01

In Parkinson's disease (PD), the response to L-dopa is highly variable and unpredictable. The major pathway for dopamine synthesis from L-dopa is decarboxylation by aromatic L-amino acid decarboxylase (AAAD, encoded by the DDC gene). To determine the motor response to L-dopa in PD patients as a function of the DDC gene promoter polymorphisms (rs921451 T > C polymorphism (DDC(T/C)) and rs3837091 AGAG del (DDC(AGAG/-))). Thirty-three Caucasian PD patients underwent an acute l-dopa challenge together with the peripheral AAAD inhibitor benserazide and were genotyped for rs921451 and rs3837091. The primary efficacy criterion was the motor response to L-dopa, as estimated by the area under the curve for the change in the Unified Parkinson's Disease Rating Scale part III (UPDRS) score relative to baseline (AUCΔUPDRS) in the 4 h following L-dopa administration. Secondary endpoints were pharmacokinetic parameters for plasma levels of L-dopa and dopamine. Investigators and patients were blinded to genotypes data throughout the study. When adjusted for the L-dopa dose, the AUCΔUPDRS was significantly lower in DDC(CC/CT) patients (n = 14) than in DDC(TT) patients (n = 19) and significantly lower in DDC(-/- or AGAG/-) patients (n = 8) than in DDC(AGAG/AGAG) patients (n = 25). There were no significant intergroup differences in plasma pharmacokinetic parameters for L-dopa and dopamine. The rs921451 and rs3837091 polymorphisms of the DDC gene promoter influence the motor response to L-dopa but do not significantly change peripheral pharmacokinetic parameters for L-dopa and dopamine. Our results suggest that DDC may be a genetic modifier of the l-dopa response in Parkinson's disease. Copyright © 2013 Elsevier Ltd. All rights reserved.

Expression of the gene encoding the ghrelin receptor in rats selected for differential alcohol preference.

PubMed

Landgren, Sara; Engel, Jörgen A; Hyytiä, Petri; Zetterberg, Henrik; Blennow, Kaj; Jerlhag, Elisabet

2011-08-01

The mechanisms involved in alcohol use disorder, a chronic relapsing brain disorder, are complex and involve various signalling systems in the brain. Recently, the orexigenic peptide ghrelin was shown to be required for alcohol-induced reward, an effect mediated via ghrelin receptors, GHS-R1A, at the level of the cholinergic-dopaminergic reward link. Moreover, ghrelin increases and GHR-R1A antagonists reduce moderate alcohol consumption in mice, and a single nucleotide polymorphism in the GHS-R1A gene has been associated with high alcohol consumption in humans. Therefore, GHS-R1A gene expression and alcohol intake were investigated in high, AA (Alko, Alcohol), versus low, ANA (Alko, Non-Alcohol), alcohol consuming rats as well as in Wistar rats. In the AA and ANA rats plasma ghrelin levels were also measured. GHS-R1A gene expression was increased in AA compared to ANA rats in nucleus accumbens, ventral tegmental area, amygdala, prefrontal cortex and hippocampus. A similar trend was observed in the ventral tegmental area of Wistar rats consuming high amounts of alcohol. Furthermore, the AA rats had significantly smaller reduction of plasma ghrelin levels over time, after several weeks of alcohol exposure, than had the ANA rats. The present study provides further evidence for that the ghrelin signalling system, in particular at the level of the mesocortocolimbic dopamine system, is involved in alcohol consumption, and thus possibly contributes to alcohol use disorder. Therefore the GHS-R1A may constitute a novel candidate for development of new treatment strategies for alcohol dependence. Copyright © 2011 Elsevier B.V. All rights reserved.

Microarray analysis of toxicogenomic effects of Ortho-phenylphenol in Staphylococcus aureus

PubMed Central

Jang, Hyeung-Jin; Nde, Chantal; Toghrol, Freshteh; Bentley, William E

2008-01-01

Background Staphylococcus aureus (S. aureus), is responsible for many infectious diseases, ranging from benign skin infections to life-threatening endocarditis and toxic shock syndrome. Ortho-phenylphenol (OPP) is an antimicrobial agent and an active ingredient of EPA-registered disinfectants with wide human exposure in various agricultural, hospital and veterinary disinfectant products. Despite many uses, an understanding of a cellular response to OPP and it's mechanism of action, targeted genes, and the connectivity between targeted genes and the rest of cell metabolism remains obscure. Results Herein, we performed a genome-wide transcriptome analysis of the cellular responses of S. aureus when exposed to 0.82 mM of OPP for 20 and 60 min. Our data indicated that OPP downregulated the biosynthesis of many amino acids, which are required for protein synthesis. In particular, the genes encoding the enzymes of the diaminopimelate (DAP) pathway which results in lysine biosynthesis were significantly downregualted. Intriguingly, we revealed that the transcription of genes encoding ribosomal proteins was upregulated by OPP and at the same time, the genes encoding iron acquisition and transport were downregulated. The genes encoding virulence factors were upregulated and genes encoding phospholipids were downregulated upon 20 min exposure to OPP. Conclusion By using microarray analysis that enables us to simultaneously and globally examine the complete transcriptome during cellular responses, we have revealed novel information regarding the mode of action of OPP on Staphylococcus: OPP inhibits anabolism of many amino acids and highly downregulates the genes that encode the enzymes involved in the DAP pathway. Lysine and DAP are essential for building up the peptidoglycan cell wall. It was concluded that the mode of action of OPP is similar to the mechanism of action of some antibiotics. The discovery of this phenomenon provides useful information that will benefit further antimicrobial research on S. aureus. PMID:18793396

Degradation of triglycerides by a pseudomonad isolated from milk: molecular analysis of a lipase-encoding gene and its expression in Escherichia coli.

PubMed Central

Johnson, L A; Beacham, I R; MacRae, I C; Free, M L

1992-01-01

Psychrotrophic lipolytic bacteria represent a significant problem in the storage of refrigerated dairy products. A lipase-encoding gene has been cloned and characterized from a highly lipolytic strain of Pseudomonas. The nucleotide sequence of the gene predicts a polypeptide of M(r) 49,905, which was identified when the gene was expressed in Escherichia coli. Images PMID:1622251

Genes encoding giant danio and golden shiner ependymin.

PubMed

Adams, D S; Kiyokawa, M; Getman, M E; Shashoua, V E

1996-03-01

Ependymin (EPN) is a brain glycoprotein that functions as a neurotrophic factor in optic nerve regeneration and long-term memory consolidation in goldfish. To date, true epn genes have been characterized in one order of teleost fish, Cypriniformes. In the study presented here, polymerase chain reactions were used to analyze the complete epn genes, gd (1480 bp), and sh (2071 bp), from Cypriniformes giant danio and shiner, respectively. Southern hybridizations demonstrated the existence of one copy of each gene per corresponding haploid genome. Each gene was found to contain six exons and five introns. Gene gd encodes a predicted 218-amino acid (aa) protein GD 93 percent conserved to goldfish EPN, while sh encodes a predicted 214-aa protein SH 91 percent homologous to goldfish. Evidence is presented classifying proteins previously termed "EPNs" into two major categories: true EPNs and non-EPN cerebrospinal fluid glycoproteins. Proteins GD and SH contain all the hallmark, features of true EPNs.

C2-domain abscisic acid-related proteins mediate the interaction of PYR/PYL/RCAR abscisic acid receptors with the plasma membrane and regulate abscisic acid sensitivity in Arabidopsis.

PubMed

Rodriguez, Lesia; Gonzalez-Guzman, Miguel; Diaz, Maira; Rodrigues, Americo; Izquierdo-Garcia, Ana C; Peirats-Llobet, Marta; Fernandez, Maria A; Antoni, Regina; Fernandez, Daniel; Marquez, Jose A; Mulet, Jose M; Albert, Armando; Rodriguez, Pedro L

2014-12-01

Membrane-delimited abscisic acid (ABA) signal transduction plays a critical role in early ABA signaling, but the molecular mechanisms linking core signaling components to the plasma membrane are unclear. We show that transient calcium-dependent interactions of PYR/PYL ABA receptors with membranes are mediated through a 10-member family of C2-domain ABA-related (CAR) proteins in Arabidopsis thaliana. Specifically, we found that PYL4 interacted in an ABA-independent manner with CAR1 in both the plasma membrane and nucleus of plant cells. CAR1 belongs to a plant-specific gene family encoding CAR1 to CAR10 proteins, and bimolecular fluorescence complementation and coimmunoprecipitation assays showed that PYL4-CAR1 as well as other PYR/PYL-CAR pairs interacted in plant cells. The crystal structure of CAR4 was solved, which revealed that, in addition to a classical calcium-dependent lipid binding C2 domain, a specific CAR signature is likely responsible for the interaction with PYR/PYL receptors and their recruitment to phospholipid vesicles. This interaction is relevant for PYR/PYL function and ABA signaling, since different car triple mutants affected in CAR1, CAR4, CAR5, and CAR9 genes showed reduced sensitivity to ABA in seedling establishment and root growth assays. In summary, we identified PYR/PYL-interacting partners that mediate a transient Ca(2+)-dependent interaction with phospholipid vesicles, which affects PYR/PYL subcellular localization and positively regulates ABA signaling. © 2014 American Society of Plant Biologists. All rights reserved.

C2-Domain Abscisic Acid-Related Proteins Mediate the Interaction of PYR/PYL/RCAR Abscisic Acid Receptors with the Plasma Membrane and Regulate Abscisic Acid Sensitivity in Arabidopsis[C][W

PubMed Central

Rodriguez, Lesia; Diaz, Maira; Rodrigues, Americo; Izquierdo-Garcia, Ana C.; Peirats-Llobet, Marta; Fernandez, Maria A.; Antoni, Regina; Fernandez, Daniel; Marquez, Jose A.; Mulet, Jose M.; Albert, Armando; Rodriguez, Pedro L.

2014-01-01

Membrane-delimited abscisic acid (ABA) signal transduction plays a critical role in early ABA signaling, but the molecular mechanisms linking core signaling components to the plasma membrane are unclear. We show that transient calcium-dependent interactions of PYR/PYL ABA receptors with membranes are mediated through a 10-member family of C2-domain ABA-related (CAR) proteins in Arabidopsis thaliana. Specifically, we found that PYL4 interacted in an ABA-independent manner with CAR1 in both the plasma membrane and nucleus of plant cells. CAR1 belongs to a plant-specific gene family encoding CAR1 to CAR10 proteins, and bimolecular fluorescence complementation and coimmunoprecipitation assays showed that PYL4-CAR1 as well as other PYR/PYL-CAR pairs interacted in plant cells. The crystal structure of CAR4 was solved, which revealed that, in addition to a classical calcium-dependent lipid binding C2 domain, a specific CAR signature is likely responsible for the interaction with PYR/PYL receptors and their recruitment to phospholipid vesicles. This interaction is relevant for PYR/PYL function and ABA signaling, since different car triple mutants affected in CAR1, CAR4, CAR5, and CAR9 genes showed reduced sensitivity to ABA in seedling establishment and root growth assays. In summary, we identified PYR/PYL-interacting partners that mediate a transient Ca2+-dependent interaction with phospholipid vesicles, which affects PYR/PYL subcellular localization and positively regulates ABA signaling. PMID:25465408

Ti plasmid-encoded genes responsible for catabolism of the crown gall opine mannopine by Agrobacterium tumefaciens are homologs of the T-region genes responsible for synthesis of this opine by the plant tumor.

PubMed

Kim, K S; Farrand, S K

1996-06-01

Agrobacterium tumefaciens NT1 harboring pSaB4, which contains the 14-kb BamHI fragment 4 from the octopine/mannityl opine-type Ti plasmid pTi15955, grew well with agropine (AGR) but slowly with mannopine (MOP) as the sole carbon source. When a second plasmid encoding a dedicated transport system for MOP was introduced, these cells grew well with both AGR and MOP. Transposon insertion mutagenesis and subcloning identified a 5.7-kb region of BamHI fragment 4 that encodes functions required for the degradation of MOP. DNA sequence analysis revealed seven putative genes in this region: mocD (moc for mannityl opine catabolism) and mocE, oriented from right to left, and mocRCBAS, oriented from left to right. Significant identities exist at the nucleotide and derived amino acid sequence levels between these moc genes and the mas genes that are responsible for opine biosynthesis in crown gall tumors. MocD is a homolog of Mas2, the anabolic conjugase encoded by mas2'. MocE and MocC are related to the amino half and the carboxyl half, respectively, of Mas1 (MOP reductase), the second enzyme for MOP biosynthesis. These results indicate that the moc and mas genes evolved from a common origin. MocR and MocS are related to each other and to a putative repressor for the AGR degradation system encoded by the rhizogenic plasmid pRiA4. MocB and MocA are homologs of 6-phosphogluconate dehydratase and glucose-6-phosphate dehydrogenase, respectively. Mutations in mocD and mocE, but not mocC, are suppressed by functions encoded by the chromosome or the 450-kb megaplasmid present in many Agrobacterium isolates. We propose that moc genes derived from genes located elsewhere in the bacterial genome and that the tumor-expressed mas genes evolved from the bacterial moc genes.

Ti plasmid-encoded genes responsible for catabolism of the crown gall opine mannopine by Agrobacterium tumefaciens are homologs of the T-region genes responsible for synthesis of this opine by the plant tumor.

PubMed Central

Kim, K S; Farrand, S K

1996-01-01

Agrobacterium tumefaciens NT1 harboring pSaB4, which contains the 14-kb BamHI fragment 4 from the octopine/mannityl opine-type Ti plasmid pTi15955, grew well with agropine (AGR) but slowly with mannopine (MOP) as the sole carbon source. When a second plasmid encoding a dedicated transport system for MOP was introduced, these cells grew well with both AGR and MOP. Transposon insertion mutagenesis and subcloning identified a 5.7-kb region of BamHI fragment 4 that encodes functions required for the degradation of MOP. DNA sequence analysis revealed seven putative genes in this region: mocD (moc for mannityl opine catabolism) and mocE, oriented from right to left, and mocRCBAS, oriented from left to right. Significant identities exist at the nucleotide and derived amino acid sequence levels between these moc genes and the mas genes that are responsible for opine biosynthesis in crown gall tumors. MocD is a homolog of Mas2, the anabolic conjugase encoded by mas2'. MocE and MocC are related to the amino half and the carboxyl half, respectively, of Mas1 (MOP reductase), the second enzyme for MOP biosynthesis. These results indicate that the moc and mas genes evolved from a common origin. MocR and MocS are related to each other and to a putative repressor for the AGR degradation system encoded by the rhizogenic plasmid pRiA4. MocB and MocA are homologs of 6-phosphogluconate dehydratase and glucose-6-phosphate dehydrogenase, respectively. Mutations in mocD and mocE, but not mocC, are suppressed by functions encoded by the chromosome or the 450-kb megaplasmid present in many Agrobacterium isolates. We propose that moc genes derived from genes located elsewhere in the bacterial genome and that the tumor-expressed mas genes evolved from the bacterial moc genes. PMID:8655509

Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks

PubMed Central

Damiani, Isabelle; Drain, Alice; Guichard, Marjorie; Balzergue, Sandrine; Boscari, Alexandre; Boyer, Jean-Christophe; Brunaud, Véronique; Cottaz, Sylvain; Rancurel, Corinne; Da Rocha, Martine; Fizames, Cécile; Fort, Sébastien; Gaillard, Isabelle; Maillol, Vincent; Danchin, Etienne G. J.; Rouached, Hatem; Samain, Eric; Su, Yan-Hua; Thouin, Julien; Touraine, Bruno; Puppo, Alain; Frachisse, Jean-Marie; Pauly, Nicolas; Sentenac, Hervé

2016-01-01

Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as “papilionoid legume-specific” were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This transcriptome provides a valuable resource to investigate root hair biology in legumes and the roles that these cells play in rhizobial symbiosis establishment. These results could also contribute to the long-term objective of transferring this symbiotic capacity to non-legume plants. PMID:27375649

Single proteins that serve linked functions in intracellular and extracellular microenvironments

DOE Office of Scientific and Technical Information (OSTI.GOV)

Radisky, Derek C.; Stallings-Mann, Melody; Hirai, Yohei

2009-06-03

Maintenance of organ homeostasis and control of appropriate response to environmental alterations requires intimate coordination of cellular function and tissue organization. An important component of this coordination may be provided by proteins that can serve distinct, but linked, functions on both sides of the plasma membrane. Here we present a novel hypothesis in which non-classical secretion can provide a mechanism through which single proteins can integrate complex tissue functions. Single genes can exert a complex, dynamic influence through a number of different processes that act to multiply the function of the gene product(s). Alternative splicing can create many different transcriptsmore » that encode proteins of diverse, even antagonistic, function from a single gene. Posttranslational modifications can alter the stability, activity, localization, and even basic function of proteins. A protein can exist in different subcellular localizations. More recently, it has become clear that single proteins can function both inside and outside the cell. These proteins often lack defined secretory signal sequences, and transit the plasma membrane by mechanisms separate from the classical ER/Golgi secretory process. When examples of such proteins are examined individually, the multifunctionality and lack of a signal sequence are puzzling - why should a protein with a well known function in one context function in such a distinct fashion in another? We propose that one reason for a single protein to perform intracellular and extracellular roles is to coordinate organization and maintenance of a global tissue function. Here, we describe in detail three specific examples of proteins that act in this fashion, outlining their specific functions in the extracellular space and in the intracellular space, and we discuss how these functions may be linked. We present epimorphin/syntaxin-2, which may coordinate morphogenesis of secretory organs (as epimorphin) with control of protein secretion (as syntaxin-2), amphoterin/high mobility group box-1 (HMGB1), which may link inflammation (as amphoterin) with regulation of gene expression (as HMGB1), and tissue transglutaminase, which affects delivery of and response to apoptotic signals by serving a related function on both sides of the plasma membrane. As it is notable that all three of these proteins have been reported to transit the plasma membrane through non-classical secretory mechanisms, we will also discuss why coordinated inside/outside functions may be found in some examples of proteins which transit the plasma membrane through non-classical mechanisms and how this relationship can be used to identify additional proteins that share these characteristics.« less

Transgenic tomatoes express an antigenic polypeptide containing epitopes of the diphtheria, pertussis and tetanus exotoxins, encoded by a synthetic gene.

PubMed

Soria-Guerra, Ruth Elena; Rosales-Mendoza, Sergio; Márquez-Mercado, Crisóforo; López-Revilla, Rubén; Castillo-Collazo, Rosalba; Alpuche-Solís, Angel Gabriel

2007-07-01

A current priority of vaccinology is the development of multicomponent vaccines that protect against several pathogens. The diphtheria-pertussis-tetanus (DPT) vaccine prevents the symptoms of three serious and often fatal diseases due to the exotoxins produced by Corynebacterium diphteriae, Bordetella pertussis and Clostridium tetani. We are attempting to develop an edible DPT multicomponent vaccine in plants, based on the fusion of protective exotoxin epitopes encoded by synthetic genes. By means of Agrobacterium mediated transformation we generated transgenic tomatoes with a plant-optimised synthetic gene encoding a novel polypeptide containing two adjuvant and six DPT immunoprotective exotoxin epitopes joined by peptide linkers. In transformed tomato plants, integration of the synthetic DPT (sDPT) gene detected by PCR was confirmed by Southern blot, and specific transcripts of the expected molecular size were detected by RT-PCR. Expression of the putative polypeptide encoded by the sDPT gene was detected by immunoassay with specific antibodies to the diphtheria, pertussis and tetanus exotoxins. The sDPT gene is therefore integrated, transcribed and translated as the expected recombinant sDPT multiepitope polypeptide in transgenic tomatoes that constitute a potential edible vaccine.

Ubiquitin--conserved protein or selfish gene?

PubMed

Catic, André; Ploegh, Hidde L

2005-11-01

The posttranslational modifier ubiquitin is encoded by a multigene family containing three primary members, which yield the precursor protein polyubiquitin and two ubiquitin moieties, Ub(L40) and Ub(S27), that are fused to the ribosomal proteins L40 and S27, respectively. The gene encoding polyubiquitin is highly conserved and, until now, those encoding Ub(L40) and Ub(S27) have been generally considered to be equally invariant. The evolution of the ribosomal ubiquitin moieties is, however, proving to be more dynamic. It seems that the genes encoding Ub(L40) and Ub(S27) are actively maintained by homologous recombination with the invariant polyubiquitin locus. Failure to recombine leads to deterioration of the sequence of the ribosomal ubiquitin moieties in several phyla, although this deterioration is evidently constrained by the structural requirements of the ubiquitin fold. Only a few amino acids in ubiquitin are vital for its function, and we propose that conservation of all three ubiquitin genes is driven not only by functional properties of the ubiquitin protein, but also by the propensity of the polyubiquitin locus to act as a 'selfish gene'.

Alternative intronic promoters in development and disease.

PubMed

Vacik, Tomas; Raska, Ivan

2017-05-01

Approximately 20,000 mammalian genes are estimated to encode between 250 thousand and 1 million different proteins. This enormous diversity of the mammalian proteome is caused by the ability of a single-gene locus to encode multiple protein isoforms. Protein isoforms encoded by one gene locus can be functionally distinct, and they can even have antagonistic functions. One of the mechanisms involved in creating this proteome complexity is alternative promoter usage. Alternative intronic promoters are located downstream from their canonical counterparts and drive the expression of alternative RNA isoforms that lack upstream exons. These upstream exons can encode some important functional domains, and proteins encoded by alternative mRNA isoforms can be thus functionally distinct from the full-length protein encoded by canonical mRNA isoforms. Since any misbalance of functionally distinct protein isoforms is likely to have detrimental consequences for the cell and the whole organism, their expression must be precisely regulated. Misregulation of alternative intronic promoters is frequently associated with various developmental defects and diseases including cancer, and it is becoming increasingly clear that this phenomenon deserves more attention.

Amplification of Chromosome 1q Genes Encoding the Phosphoinositide Signalling Enzymes PI4KB, AKT3, PIP5K1A and PI3KC2B in Breast Cancer

PubMed Central

Waugh, Mark G.

2014-01-01

Little is known about the possible oncogenic roles of genes encoding for the phosphatidylinositol 4-kinases, a family of enzymes that regulate an early step in phosphoinositide signalling. To address this issue, the mutational status of all four human phosphatidylinositol 4-kinases genes was analyzed across 852 breast cancer samples using the COSMIC data resource. Point mutations in the phosphatidylinositol 4-kinase genes were uncommon and appeared in less than 1% of the patient samples however, 62% of the tumours had increases in gene copy number for PI4KB which encodes the phosphatidylinositol 4-kinase IIIbeta isozyme. Extending this analysis to subsequent enzymes in the phosphoinositide signalling cascades revealed that the only PIP5K1A, PI3KC2B and AKT3 genes exhibited similar patterns of gene copy number variation. By comparison, gene copy number increases for established oncogenes such as EGFR and HER2/Neu were only evident in 20% of the samples. The PI4KB, PIP5K1A, PI3KC2B and AKT3 genes are related in that they all localize to chromosome 1q which is often structurally and numerically abnormal in breast cancer. These results demonstrate that a gene quartet encoding a potential phosphoinositide signalling pathway is amplified in a subset of breast cancers. PMID:25368680

Bioinformatics analysis and detection of gelatinase encoded gene in Lysinibacillussphaericus

NASA Astrophysics Data System (ADS)

Repin, Rul Aisyah Mat; Mutalib, Sahilah Abdul; Shahimi, Safiyyah; Khalid, Rozida Mohd.; Ayob, Mohd. Khan; Bakar, Mohd. Faizal Abu; Isa, Mohd Noor Mat

2016-11-01

In this study, we performed bioinformatics analysis toward genome sequence of Lysinibacillussphaericus (L. sphaericus) to determine gene encoded for gelatinase. L. sphaericus was isolated from soil and gelatinase species-specific bacterium to porcine and bovine gelatin. This bacterium offers the possibility of enzymes production which is specific to both species of meat, respectively. The main focus of this research is to identify the gelatinase encoded gene within the bacteria of L. Sphaericus using bioinformatics analysis of partially sequence genome. From the research study, three candidate gene were identified which was, gelatinase candidate gene 1 (P1), NODE_71_length_93919_cov_158.931839_21 which containing 1563 base pair (bp) in size with 520 amino acids sequence; Secondly, gelatinase candidate gene 2 (P2), NODE_23_length_52851_cov_190.061386_17 which containing 1776 bp in size with 591 amino acids sequence; and Thirdly, gelatinase candidate gene 3 (P3), NODE_106_length_32943_cov_169.147919_8 containing 1701 bp in size with 566 amino acids sequence. Three pairs of oligonucleotide primers were designed and namely as, F1, R1, F2, R2, F3 and R3 were targeted short sequences of cDNA by PCR. The amplicons were reliably results in 1563 bp in size for candidate gene P1 and 1701 bp in size for candidate gene P3. Therefore, the results of bioinformatics analysis of L. Sphaericus resulting in gene encoded gelatinase were identified.

Role and Regulation of the Flp/Tad Pilus in the Virulence of Pectobacterium atrosepticum SCRI1043 and Pectobacterium wasabiae SCC3193

PubMed Central

Nykyri, Johanna; Mattinen, Laura; Niemi, Outi; Adhikari, Satish; Kõiv, Viia; Somervuo, Panu; Fang, Xin; Auvinen, Petri; Mäe, Andres; Palva, E. Tapio; Pirhonen, Minna

2013-01-01

In this study, we characterized a putative Flp/Tad pilus-encoding gene cluster, and we examined its regulation at the transcriptional level and its role in the virulence of potato pathogenic enterobacteria of the genus Pectobacterium. The Flp/Tad pilus-encoding gene clusters in Pectobacterium atrosepticum, Pectobacterium wasabiae and Pectobacterium aroidearum were compared to previously characterized flp/tad gene clusters, including that of the well-studied Flp/Tad pilus model organism Aggregatibacter actinomycetemcomitans, in which this pilus is a major virulence determinant. Comparative analyses revealed substantial protein sequence similarity and open reading frame synteny between the previously characterized flp/tad gene clusters and the cluster in Pectobacterium, suggesting that the predicted flp/tad gene cluster in Pectobacterium encodes a Flp/Tad pilus-like structure. We detected genes for a novel two-component system adjacent to the flp/tad gene cluster in Pectobacterium, and mutant analysis demonstrated that this system has a positive effect on the transcription of selected Flp/Tad pilus biogenesis genes, suggesting that this response regulator regulate the flp/tad gene cluster. Mutagenesis of either the predicted regulator gene or selected Flp/Tad pilus biogenesis genes had a significant impact on the maceration ability of the bacterial strains in potato tubers, indicating that the Flp/Tad pilus-encoding gene cluster represents a novel virulence determinant in Pectobacterium. Soft-rot enterobacteria in the genera Pectobacterium and Dickeya are of great agricultural importance, and an investigation of the virulence of these pathogens could facilitate improvements in agricultural practices, thus benefiting farmers, the potato industry and consumers. PMID:24040039

Role and regulation of the Flp/Tad pilus in the virulence of Pectobacterium atrosepticum SCRI1043 and Pectobacterium wasabiae SCC3193.

PubMed

Nykyri, Johanna; Mattinen, Laura; Niemi, Outi; Adhikari, Satish; Kõiv, Viia; Somervuo, Panu; Fang, Xin; Auvinen, Petri; Mäe, Andres; Palva, E Tapio; Pirhonen, Minna

2013-01-01

In this study, we characterized a putative Flp/Tad pilus-encoding gene cluster, and we examined its regulation at the transcriptional level and its role in the virulence of potato pathogenic enterobacteria of the genus Pectobacterium. The Flp/Tad pilus-encoding gene clusters in Pectobacterium atrosepticum, Pectobacterium wasabiae and Pectobacterium aroidearum were compared to previously characterized flp/tad gene clusters, including that of the well-studied Flp/Tad pilus model organism Aggregatibacter actinomycetemcomitans, in which this pilus is a major virulence determinant. Comparative analyses revealed substantial protein sequence similarity and open reading frame synteny between the previously characterized flp/tad gene clusters and the cluster in Pectobacterium, suggesting that the predicted flp/tad gene cluster in Pectobacterium encodes a Flp/Tad pilus-like structure. We detected genes for a novel two-component system adjacent to the flp/tad gene cluster in Pectobacterium, and mutant analysis demonstrated that this system has a positive effect on the transcription of selected Flp/Tad pilus biogenesis genes, suggesting that this response regulator regulate the flp/tad gene cluster. Mutagenesis of either the predicted regulator gene or selected Flp/Tad pilus biogenesis genes had a significant impact on the maceration ability of the bacterial strains in potato tubers, indicating that the Flp/Tad pilus-encoding gene cluster represents a novel virulence determinant in Pectobacterium. Soft-rot enterobacteria in the genera Pectobacterium and Dickeya are of great agricultural importance, and an investigation of the virulence of these pathogens could facilitate improvements in agricultural practices, thus benefiting farmers, the potato industry and consumers.

Distribution and Evolution of Yersinia Leucine-Rich Repeat Proteins

PubMed Central

Hu, Yueming; Huang, He; Hui, Xinjie; Cheng, Xi; White, Aaron P.

2016-01-01

Leucine-rich repeat (LRR) proteins are widely distributed in bacteria, playing important roles in various protein-protein interaction processes. In Yersinia, the well-characterized type III secreted effector YopM also belongs to the LRR protein family and is encoded by virulence plasmids. However, little has been known about other LRR members encoded by Yersinia genomes or their evolution. In this study, the Yersinia LRR proteins were comprehensively screened, categorized, and compared. The LRR proteins encoded by chromosomes (LRR1 proteins) appeared to be more similar to each other and different from those encoded by plasmids (LRR2 proteins) with regard to repeat-unit length, amino acid composition profile, and gene expression regulation circuits. LRR1 proteins were also different from LRR2 proteins in that the LRR1 proteins contained an E3 ligase domain (NEL domain) in the C-terminal region or an NEL domain-encoding nucleotide relic in flanking genomic sequences. The LRR1 protein-encoding genes (LRR1 genes) varied dramatically and were categorized into 4 subgroups (a to d), with the LRR1a to -c genes evolving from the same ancestor and LRR1d genes evolving from another ancestor. The consensus and ancestor repeat-unit sequences were inferred for different LRR1 protein subgroups by use of a maximum parsimony modeling strategy. Structural modeling disclosed very similar repeat-unit structures between LRR1 and LRR2 proteins despite the different unit lengths and amino acid compositions. Structural constraints may serve as the driving force to explain the observed mutations in the LRR regions. This study suggests that there may be functional variation and lays the foundation for future experiments investigating the functions of the chromosomally encoded LRR proteins of Yersinia. PMID:27217422

Characterization of the Genes Encoding the Cytosolic and Plastidial Forms of ADP-Glucose Pyrophosphorylase in Wheat Endosperm1

PubMed Central

Burton, Rachel A.; Johnson, Philip E.; Beckles, Diane M.; Fincher, Geoffrey B.; Jenner, Helen L.; Naldrett, Mike J.; Denyer, Kay

2002-01-01

In most species, the synthesis of ADP-glucose (Glc) by the enzyme ADP-Glc pyrophosphorylase (AGPase) occurs entirely within the plastids in all tissues so far examined. However, in the endosperm of many, if not all grasses, a second form of AGPase synthesizes ADP-Glc outside the plastid, presumably in the cytosol. In this paper, we show that in the endosperm of wheat (Triticum aestivum), the cytosolic form accounts for most of the AGPase activity. Using a combination of molecular and biochemical approaches to identify the cytosolic and plastidial protein components of wheat endosperm AGPase we show that the large and small subunits of the cytosolic enzyme are encoded by genes previously thought to encode plastidial subunits, and that a gene, Ta.AGP.S.1, which encodes the small subunit of the cytosolic form of AGPase, also gives rise to a second transcript by the use of an alternate first exon. This second transcript encodes an AGPase small subunit with a transit peptide. However, we could not find a plastidial small subunit protein corresponding to this transcript. The protein sequence of the purified plastidial small subunit does not match precisely to that encoded by Ta.AGP.S.1 or to the predicted sequences of any other known gene from wheat or barley (Hordeum vulgare). Instead, the protein sequence is most similar to those of the plastidial small subunits from chickpea (Cicer arietinum) and maize (Zea mays) and rice (Oryza sativa) seeds. These data suggest that the gene encoding the major plastidial small subunit of AGPase in wheat endosperm has yet to be identified. PMID:12428011

Effects of SULT1A1 Copy Number Variation on Estrogen Concentration and Tamoxifen-Associated Adverse Drug Reactions in Premenopausal Thai Breast Cancer Patients: A Preliminary Study.

PubMed

Charoenchokthavee, Wanaporn; Ayudhya, Duangchit Panomvana Na; Sriuranpong, Virote; Areepium, Nutthada

2016-01-01

Tamoxifen is a pharmacological estrogen inhibitor that binds to the estrogen receptor (ER) in breast cells. However, it shows an estrogenic effect in other organs, which causes adverse drug reactions (ADRs). The sulfotransferase 1A1 (SULT1A1) enzyme encoded by the SULT1A1 gene is involved in estrogen metabolism. Previous research has suggested that the SULT1A1 copy number is linked with the plasma estradiol (E2) concentration. Here, a total of 34 premenopausal breast cancer patients, selected from the Thai Tamoxifen (TTAM) Project, were screened for their SULT1A1 copy number, plasma E2 concentration and ADRs. The mean age was 44.3±11.1 years, and they were subtyped as ER+/ progesterone receptor (PR) + (28 patients), ER+/ PR- (5 patients) and ER-/PR- (1 patient). Three patients reported ADRs, which were irregular menstruation (2 patients) and vaginal discharge (1 patient). Most (33) patients had two SULT1A1 copies, with one patient having three copies. The median plasma E2 concentration was 1,575.6 (IQR 865.4) pg/ml. Patients with ADRs had significantly higher plasma E2 concentrations than those patients without ADRs (p = 0.014). The plasma E2 concentration was numerically higher in the patient with three SULT1A1 copies, but this lacked statistical significance.

Structural and transcriptional characterization of a novel member of the soybean urease gene family.

PubMed

Wiebke-Strohm, Beatriz; Ligabue-Braun, Rodrigo; Rechenmacher, Ciliana; De Oliveira-Busatto, Luisa Abruzzi; Carlini, Célia Regina; Bodanese-Zanettini, Maria Helena

2016-04-01

In plants, ureases have been related to urea degradation, to defense against pathogenic fungi and phytophagous insects, and to the soybean-Bradyrhizobium japonicum symbiosis. Two urease isoforms have been described for soybean: the embryo-specific, encoded by Eu1 gene, and the ubiquitous urease, encoded by Eu4. A third urease-encoding locus exists in the completed soybean genome. The gene was designated Eu5 and the putative product of its ORF as SBU-III. Phylogenetic analysis shows that 41 plant, moss and algal ureases have diverged from a common ancestor protein, but ureases from monocots, eudicots and ancient species have evolved independently. Genomes of ancient organisms present a single urease-encoding gene and urease-encoding gene duplication has occurred independently along the evolution of some eudicot species. SBU-III has a shorter amino acid sequence, since many gaps are found when compared to other sequences. A mutation in a highly conserved amino acid residue suggests absence of ureolytic activity, but the overall protein architecture remains very similar to the other ureases. The expression profile of urease-encoding genes in different organs and developmental stages was determined by RT-qPCR. Eu5 transcripts were detected in seeds one day after dormancy break, roots of young plants and embryos of developing seeds. Eu1 and Eu4 transcripts were found in all analyzed organs, but Eu4 expression was more prominent in seeds one day after dormancy break whereas Eu1 predominated in developing seeds. The evidence suggests that SBU-III may not be involved in nitrogen availability to plants, but it could be involved in other biological role(s). Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Systems-level analysis of risk genes reveals the modular nature of schizophrenia.

PubMed

Liu, Jiewei; Li, Ming; Luo, Xiong-Jian; Su, Bing

2018-05-19

Schizophrenia (SCZ) is a complex mental disorder with high heritability. Genetic studies (especially recent genome-wide association studies) have identified many risk genes for schizophrenia. However, the physical interactions among the proteins encoded by schizophrenia risk genes remain elusive and it is not known whether the identified risk genes converge on common molecular networks or pathways. Here we systematically investigated the network characteristics of schizophrenia risk genes using the high-confidence protein-protein interactions (PPI) from the human interactome. We found that schizophrenia risk genes encode a densely interconnected PPI network (P = 4.15 × 10 -31 ). Compared with the background genes, the schizophrenia risk genes in the interactome have significantly higher degree (P = 5.39 × 10 -11 ), closeness centrality (P = 7.56 × 10 -11 ), betweeness centrality (P = 1.29 × 10 -11 ), clustering coefficient (P = 2.22 × 10 -2 ), and shorter average shortest path length (P = 7.56 × 10 -11 ). Based on the densely interconnected PPI network, we identified 48 hub genes and 4 modules formed by highly interconnected schizophrenia genes. We showed that the proteins encoded by schizophrenia hub genes have significantly more direct physical interactions. Gene ontology (GO) analysis revealed that cell adhesion, cell cycle, immune system response, and GABR-receptor complex categories were enriched in the modules formed by highly interconnected schizophrenia risk genes. Our study reveals that schizophrenia risk genes encode a densely interconnected molecular network and demonstrates the modular nature of schizophrenia. Copyright © 2018 Elsevier B.V. All rights reserved.

Allelic variations in the CYBA gene of NADPH oxidase and risk of kidney complications in patients with type 1 diabetes.

PubMed

Patente, Thiago A; Mohammedi, Kamel; Bellili-Muñoz, Naïma; Driss, Fathi; Sanchez, Manuel; Fumeron, Frédéric; Roussel, Ronan; Hadjadj, Samy; Corrêa-Giannella, Maria Lúcia; Marre, Michel; Velho, Gilberto

2015-09-01

Oxidative stress plays a pivotal role in the pathophysiology of diabetic nephropathy, and the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system is an important source of reactive oxygen species in hyperglycemic conditions in the kidney. Plasma concentration of advanced oxidation protein products (AOPP), a marker of oxidative stress, is increased in patients with diabetic nephropathy. We investigated associations of variants in the CYBA gene, encoding the regulatory subunit p22(phox) of NADPH oxidase, with diabetic nephropathy and plasma AOPP and myeloperoxidase (MPO) concentrations in type 1 diabetic patients. Seven SNPs in the CYBA region were analyzed in 1357 Caucasian subjects with type 1 diabetes from the SURGENE (n=340), GENEDIAB (n=444), and GENESIS (n=573) cohorts. Duration of follow-up was 10, 9, and 6 years, respectively. Cox proportional hazards and logistic regression analyses were used to estimate hazard ratios (HR) or odds ratios (OR) for incidence and prevalence of diabetic nephropathy. The major G-allele of rs9932581 was associated with the incidence of renal events defined as new cases of microalbuminuria or the progression to a more severe stage of nephropathy during follow-up (HR 1.59, 95% CI 1.17-2.18, P=0.003) in SURGENE. The same allele was associated with established/advanced nephropathy (OR 1.52, 95% CI 1.22-1.92, P=0.0001) and with the incidence of end-stage renal disease (ESRD) (HR 2.01, 95% CI 1.30-3.24, P=0.001) in GENEDIAB/GENESIS pooled studies. The risk allele was also associated with higher plasma AOPP concentration in subsets of SURGENE and GENEDIAB, with higher plasma MPO concentration in a subset of GENEDIAB, and with lower estimated glomerular filtration rate (eGFR) in the three cohorts. In conclusion, a functional variant in the promoter of the CYBA gene was associated with lower eGFR and with prevalence and incidence of diabetic nephropathy and ESRD in type 1 diabetic patients. These results are consistent with a role for NADPH oxidase in the pathophysiology of kidney complications of diabetes. Copyright © 2015 Elsevier Inc. All rights reserved.

Cloning and expression of prion protein encoding gene of flounder ( Paralichthys olivaceus)

NASA Astrophysics Data System (ADS)

Zhang, Zhiwen; Sun, Xiuqin; Zhang, Jinxing; Zan, Jindong

2008-02-01

The prion protein (PrP) encoding gene of flounder ( Paralichthys olivaceus) was cloned. It was not interrupted by an intron. This gene has two promoters in its 5' upstream, indicating that its transcription may be intensive, and should have an important function. It was expressed in all 14 tissues tested, demonstrating that it is a house-keeping gene. Its expression in digestion and reproduction systems implies that the possible prions of fish may transfer horizontally.

Physiological roles of pyruvate ferredoxin oxidoreductase and pyruvate formate-lyase in Thermoanaerobacterium saccharolyticum JW/SL-YS485

DOE PAGES

Zhou, Jilai; Olson, Daniel G.; Lanahan, Anthony A.; ...

2015-09-15

We report that Thermoanaerobacter saccharolyticum is a thermophilic microorganism that has been engineered to produce ethanol at high titer (30–70 g/L) and greater than 90 % theoretical yield. However, few genes involved in pyruvate to ethanol production pathway have been unambiguously identified. In T. saccharolyticum, the products of six putative pfor gene clusters and one pfl gene may be responsible for the conversion of pyruvate to acetyl-CoA. To gain insights into the physiological roles of PFOR and PFL, we studied the effect of deletions of several genes thought to encode these activities. We found that that pyruvate ferredoxin oxidoreductase enzymemore » (PFOR) is encoded by the pforA gene and plays a key role in pyruvate dissimilation. We further demonstrated that pyruvate formate-lyase activity (PFL) is encoded by the pfl gene. Although the pfl gene is normally expressed at low levels, it is crucial for biosynthesis in T. saccharolyticum. In pforA deletion strains, pfl expression increased and was able to partially compensate for the loss of PFOR activity. Deletion of both pforA and pfl resulted in a strain that required acetate and formate for growth and produced lactate as the primary fermentation product, achieving 88 % theoretical lactate yield. PFOR encoded by Tsac_0046 and PFL encoded by Tsac_0628 are only two routes for converting pyruvate to acetyl-CoA in T. saccharolyticum. The physiological role of PFOR is pyruvate dissimilation, whereas that of PFL is supplying C1 units for biosynthesis.« less

Identification and functional characterization of a solute carrier family 15, member 4 gene in Litopenaeus vannamei.

PubMed

Chen, Yong-Gui; Yuan, Kai; Zhang, Ze-Zhi; Yuan, Feng-Hua; Weng, Shao-Ping; Yue, Hai-Tao; He, Jian-Guo; Chen, Yi-Hong

2016-04-01

Innate immunity in shrimp is important in resisting bacterial infection. The NF-κB pathway is pivotal in such an immune response. This study cloned and functionally characterized the solute carrier family (SLC) 15 member A 4 (LvSLC15A4) gene in Litopenaeus vannamei. The open reading frame of LvSLC15A4 is 1, 902 bp long and encodes a putative 633-amino acid protein, which is localized in the plasma membrane and intracellular vesicular compartments. Results of the reporter gene assay showed that LvSLC15A4 upregulated NF-κB target genes, including the immediate-early gene 1 of white spot syndrome virus, as well as several antimicrobial peptide genes, such as pen4, CecA, AttA, and Mtk in S2 cells. Moreover, knocked-down expression of LvSLC15A4 reduced pen4 expression in L. vannamei. LvSLC15A4 down-regulation also increased the cumulative mortality of Vibrio parahemolyticus-infected L. vannamei. Furthermore, LvSLC15A4 expression was induced by unfolded protein response (UPR) in L. vannamei hematocytes. These results suggest that LvSLC15A4 participates in L. vannamei innate immunity via the NF-κB pathway and thus may be related to UPR. Copyright © 2015 Elsevier Ltd. All rights reserved.

Transcriptome Analysis of Spartina pectinata in Response to Freezing Stress

PubMed Central

Nah, Gyoungju; Lee, Moonsub; Kim, Do-Soon; Rayburn, A. Lane; Voigt, Thomas; Lee, D. K.

2016-01-01

Prairie cordgrass (Spartina pectinata), a perennial C4 grass native to the North American prairie, has several distinctive characteristics that potentially make it a model crop for production in stressful environments. However, little is known about the transcriptome dynamics of prairie cordgrass despite its unique freezing stress tolerance. Therefore, the purpose of this work was to explore the transcriptome dynamics of prairie cordgrass in response to freezing stress at -5°C for 5 min and 30 min. We used a RNA-sequencing method to assemble the S. pectinata leaf transcriptome and performed gene-expression profiling of the transcripts under freezing treatment. Six differentially expressed gene (DEG) groups were categorized from the profiling. In addition, two major consecutive orders of gene expression were observed in response to freezing; the first being the acute up-regulation of genes involved in plasma membrane modification, calcium-mediated signaling, proteasome-related proteins, and transcription regulators (e.g., MYB and WRKY). The follow-up and second response was of genes involved in encoding the putative anti-freezing protein and the previously known DNA and cell-damage-repair proteins. Moreover, we identified the genes involved in epigenetic regulation and circadian-clock expression. Our results indicate that freezing response in S. pectinata reflects dynamic changes in rapid-time duration, as well as in metabolic, transcriptional, post-translational, and epigenetic regulation. PMID:27032112

Identification and Characterization of Three Differentially Expressed Genes, Encoding S-Adenosylhomocysteine Hydrolase, Methionine Aminopeptidase, and a Histone-Like Protein, in the Toxic Dinoflagellate Alexandrium fundyense†

PubMed Central

Taroncher-Oldenburg, Gaspar; Anderson, Donald M.

2000-01-01

Genes showing differential expression related to the early G1 phase of the cell cycle during synchronized circadian growth of the toxic dinoflagellate Alexandrium fundyense were identified and characterized by differential display (DD). The determination in our previous work that toxin production in Alexandrium is relegated to a narrow time frame in early G1 led to the hypothesis that transcriptionally up- or downregulated genes during this subphase of the cell cycle might be related to toxin biosynthesis. Three genes, encoding S-adenosylhomocysteine hydrolase (Sahh), methionine aminopeptidase (Map), and a histone-like protein (HAf), were isolated. Sahh was downregulated, while Map and HAf were upregulated, during the early G1 phase of the cell cycle. Sahh and Map encoded amino acid sequences with about 90 and 70% similarity to those encoded by several eukaryotic and prokaryotic Sahh and Map genes, respectively. The partial Map sequence also contained three cobalt binding motifs characteristic of all Map genes. HAf encoded an amino acid sequence with 60% similarity to those of two histone-like proteins from the dinoflagellate Crypthecodinium cohnii Biecheler. This study documents the potential of applying DD to the identification of genes that are related to physiological processes or cell cycle events in phytoplankton under conditions where small sample volumes represent an experimental constraint. The identification of an additional 21 genes with various cell cycle-related DD patterns also provides evidence for the importance of pretranslational or transcriptional regulation in dinoflagellates, contrary to previous reports suggesting the possibility that translational mechanisms are the primary means of circadian regulation in this group of organisms. PMID:10788388

Identification of a Novel Dioxygenase Involved in Metabolism of o-Xylene, Toluene, and Ethylbenzene by Rhodococcus sp. Strain DK17

PubMed Central

Kim, Dockyu; Chae, Jong-Chan; Zylstra, Gerben J.; Kim, Young-Soo; Kim, Seong-Ki; Nam, Myung Hee; Kim, Young Min; Kim, Eungbin

2004-01-01

Rhodococcus sp. strain DK17 is able to grow on o-xylene, benzene, toluene, and ethylbenzene. DK17 harbors at least two megaplasmids, and the genes encoding the initial steps in alkylbenzene metabolism are present on the 330-kb pDK2. The genes encoding alkylbenzene degradation were cloned in a cosmid clone and sequenced completely to reveal 35 open reading frames (ORFs). Among the ORFs, we identified two nearly exact copies (one base difference) of genes encoding large and small subunits of an iron sulfur protein terminal oxygenase that are 6 kb apart from each other. Immediately downstream of one copy of the dioxygenase genes (akbA1a and akbA2a) is a gene encoding a dioxygenase ferredoxin component (akbA3), and downstream of the other copy (akbA1b and akbA2b) are genes putatively encoding a meta-cleavage pathway. RT-PCR experiments show that the two copies of the dioxygenase genes are operonic with the downstream putative catabolic genes and that both operons are induced by o-xylene. When expressed in Escherichia coli, AkbA1a-AkbA2a-AkbA3 transformed o-xylene into 2,3- and 3,4-dimethylphenol. These were apparently derived from an unstable o-xylene cis-3,4-dihydrodiol, which readily dehydrates. This indicates a single point of attack of the dioxygenase on the aromatic ring. In contrast, attack of AkbA1a-AkbA2a-AkbA3 on ethylbenzene resulted in the formation of two different cis-dihydrodiols resulting from an oxidation at the 2,3 and the 3,4 positions on the aromatic ring, respectively. PMID:15574904

Discovery of Herpes B Virus-Encoded MicroRNAs▿

PubMed Central

Besecker, Michael I.; Harden, Mallory E.; Li, Guanglin; Wang, Xiu-Jie; Griffiths, Anthony

2009-01-01

Herpes B virus (BV) naturally infects macaque monkeys and is a close relative of herpes simplex virus. BV can zoonotically infect humans to cause a rapidly ascending encephalitis with ∼80% mortality. Therefore, BV is a serious danger to those who come into contact with these monkeys or their tissues and cells. MicroRNAs are regulators of gene expression, and there have been reports of virus-encoded microRNAs. We hypothesize that BV-encoded microRNAs are important for the regulation of viral and cellular genes. Herein, we report the discovery of three herpes B virus-encoded microRNAs. PMID:19144716

Choline Transport Activity Regulates Phosphatidylcholine Synthesis through Choline Transporter Hnm1 Stability*

PubMed Central

Fernández-Murray, J. Pedro; Ngo, Michael H.; McMaster, Christopher R.

2013-01-01

Choline is a precursor for the synthesis of phosphatidylcholine through the CDP-choline pathway. Saccharomyces cerevisiae expresses a single high affinity choline transporter at the plasma membrane, encoded by the HNM1 gene. We show that exposing cells to increasing levels of choline results in two different regulatory mechanisms impacting Hnm1 activity. Initial exposure to choline results in a rapid decrease in Hnm1-mediated transport at the level of transporter activity, whereas chronic exposure results in Hnm1 degradation through an endocytic mechanism that depends on the ubiquitin ligase Rsp5 and the casein kinase 1 redundant pair Yck1/Yck2. We present details of how the choline transporter is a major regulator of phosphatidylcholine synthesis. PMID:24187140

A null mutation in human APOC3 confers a favorable plasma lipid profile and apparent cardioprotection.

PubMed

Pollin, Toni I; Damcott, Coleen M; Shen, Haiqing; Ott, Sandra H; Shelton, John; Horenstein, Richard B; Post, Wendy; McLenithan, John C; Bielak, Lawrence F; Peyser, Patricia A; Mitchell, Braxton D; Miller, Michael; O'Connell, Jeffrey R; Shuldiner, Alan R

2008-12-12

Apolipoprotein C-III (apoC-III) inhibits triglyceride hydrolysis and has been implicated in coronary artery disease. Through a genome-wide association study, we have found that about 5% of the Lancaster Amish are heterozygous carriers of a null mutation (R19X) in the gene encoding apoC-III (APOC3) and, as a result, express half the amount of apoC-III present in noncarriers. Mutation carriers compared with noncarriers had lower fasting and postprandial serum triglycerides, higher levels of HDL-cholesterol and lower levels of LDL-cholesterol. Subclinical atherosclerosis, as measured by coronary artery calcification, was less common in carriers than noncarriers, which suggests that lifelong deficiency of apoC-III has a cardioprotective effect.

Cytochrome P450 and ABCB1 genetics: association with quetiapine and norquetiapine plasma and cerebrospinal fluid concentrations and with clinical response in patients suffering from schizophrenia. A pilot study.

PubMed

Nikisch, Georg; Baumann, Pierre; Oneda, Beatrice; Kiessling, Bernhard; Weisser, Heike; Mathé, Aleksander A; Yoshitake, Takashi; Kehr, Jan; Wiedemann, Georg; Eap, Chin B

2011-07-01

Variability in response to atypical antipsychotic drugs is due to genetic and environmental factors. Cytochrome P450 (CYP) isoforms are implicated in the metabolism of drugs, while the P-glycoprotein transporter (P-gp), encoded by the ABCB1 gene, may influence both the blood and brain drug concentrations. This study aimed to identify the possible associations of CYP and ABCB1 genetic polymorphisms with quetiapine and norquetiapine plasma and cerebrospinal fluid (CSF) concentrations and with response to treatment. Twenty-two patients with schizophrenia receiving 600 mg of quetiapine daily were genotyped for four CYP isoforms and ABCB1 polymorphisms. Quetiapine and norquetiapine peak plasma and CSF concentrations were measured after 4 weeks of treatment. Stepwise multiple regression analysis revealed that ABCB1 3435C > T (rs1045642), 2677G > T (rs2032582) and 1236C > T (rs1128503) polymorphisms predicted plasma quetiapine concentrations, explaining 41% of the variability (p = 0.001). Furthermore, the ABCB1 polymorphisms predicted 48% (p = 0.024) of the variability of the Δ PANSS total score, with the non-carriers of the 3435TT showing higher changes in the score. These results suggest that ABCB1 genetic polymorphisms may be a predictive marker of quetiapine treatment in schizophrenia.

Gene encoding acetyl-coenzyme A carboxylase

DOEpatents

Roessler, Paul G.; Ohlrogge, John B.

1996-01-01

A DNA encoding an acetyl-coenzyme A carboxylase (ACCase) from a photosynthetic organism and functional derivatives thereof which are resistant to inhibition from certain herbicides. This gene can be placed in organisms to increase their fatty acid content or to render them resistant to certain herbicides.

Oxidative dearomatisation: the key step of sorbicillinoid biosynthesis† †Electronic supplementary information (ESI) available: Containing all experimental details. See DOI: 10.1039/c3sc52911h Click here for additional data file.

PubMed Central

Fahad, Ahmed al; Abood, Amira; Fisch, Katja M.; Osipow, Anna; Davison, Jack; Avramović, Marija; Butts, Craig P.; Piel, Jörn; Simpson, Thomas J.

2014-01-01

An FAD-dependent monooxygenase encoding gene (SorbC) was cloned from Penicillium chrysogenum E01-10/3 and expressed as a soluble protein in Escherichia coli. The enzyme efficiently performed the oxidative dearomatisation of sorbicillin and dihydrosorbicillin to give sorbicillinol and dihydrosorbicillinol respectively. Bioinformatic examination of the gene cluster surrounding SorbC indicated the presence of two polyketide synthase (PKS) encoding genes designated sorbA and sorbB. The gene sorbA-encodes a highly reducing iterative PKS while SorbB encodes a non-reducing iterative PKS which features a reductive release domain usually involved in the production of polyketide aldehydes. Using these observations and previously reported results from isotopic feeding experiments a new and simpler biosynthetic route to the sorbicillin class of secondary metabolites is proposed which is consistent with all reported experimental results. PMID:25580210

A gene encoding the plant-like alternative oxidase is present in Phytomonas but absent in Leishmania spp.

PubMed

Van Hellemond, J J; Simons, B; Millenaar, F F; Tielens, A G

1998-01-01

The constituents of the respiratory chain are believed to differ among the trypanosomatids; bloodstream stages of African trypanosomes and Phytomonas promastigotes oxidize ubiquinol by a ubiquinol:oxygen oxidoreductase, also known as alternative oxidase, whereas Leishmania spp. oxidize ubiquinol via a classic cytochrome-containing respiratory chain. The molecular basis for this elementary difference in ubiquinol oxidation by the mitochondrial electron-transport chain in distinct trypanosomatids was investigated. The presence of a gene encoding the plant-like alternative oxidase could be demonstrated in Phytomonas and Trypanosoma brucei, trypanosomatids that are known to contain alternative oxidase activity. Our results further demonstrated that Leishmania spp. lack a gene encoding the plant-like alternative oxidase, and therefore, all stages of Leishmania spp. will lack the alternative oxidase protein. The observed fundamental differences between the respiratory chains of distinct members of the trypanosomatid family are thus caused by the presence or absence of a gene encoding the plant-like alternative oxidase.

Cloning of gene-encoded stem bromelain on system coming from Pichia pastoris as therapeutic protein candidate

NASA Astrophysics Data System (ADS)

Yusuf, Y.; Hidayati, W.

2018-01-01

The process of identifying bacterial recombination using PCR, and restriction, and then sequencing process was done after identifying the bacteria. This research aimed to get a yeast cell of Pichia pastoris which has an encoder gene of stem bromelain enzyme. The production of recombinant stem bromelain enzymes using yeast cells of P. pastoris can produce pure bromelain rod enzymes and have the same conformation with the enzyme’s conformation in pineapple plants. This recombinant stem bromelain enzyme can be used as a therapeutic protein in inflammatory, cancer and degenerative diseases. This study was an early stage of a step series to obtain bromelain rod protein derived from pineapple made with genetic engineering techniques. This research was started by isolating the RNA of pineapple stem which was continued with constructing cDNA using reserve transcriptase-PCR technique (RT-PCR), doing the amplification of bromelain enzyme encoder gene with PCR technique using a specific premiere couple which was designed. The process was continued by cloning into bacterium cells of Escherichia coli. A vector which brought the encoder gene of stem bromelain enzyme was inserted into the yeast cell of P. pastoris and was continued by identifying the yeast cell of P. pastoris which brought the encoder gene of stem bromelain enzyme. The research has not found enzyme gene of stem bromelain in yeast cell of P. pastoris yet. The next step is repeating the process by buying new reagent; RNase inhibitor, and buying liquid nitrogen.

Cloning and sequencing of a gene encoding a novel extracellular neutral proteinase from Streptomyces sp. strain C5 and expression of the gene in Streptomyces lividans 1326.

PubMed Central

Lampel, J S; Aphale, J S; Lampel, K A; Strohl, W R

1992-01-01

The gene encoding a novel milk protein-hydrolyzing proteinase was cloned on a 6.56-kb SstI fragment from Streptomyces sp. strain C5 genomic DNA into Streptomyces lividans 1326 by using the plasmid vector pIJ702. The gene encoding the small neutral proteinase (snpA) was located within a 2.6-kb BamHI-SstI restriction fragment that was partially sequenced. The molecular mass of the deduced amino acid sequence of the mature protein was determined to be 15,740, which corresponds very closely with the relative molecular mass of the purified protein (15,500) determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The N-terminal amino acid sequence of the purified neutral proteinase was determined, and the DNA encoding this sequence was found to be located within the sequenced DNA. The deduced amino acid sequence contains a conserved zinc binding site, although secondary ligand binding and active sites typical of thermolysinlike metalloproteinases are absent. The combination of its small size, deduced amino acid sequence, and substrate and inhibition profile indicate that snpA encodes a novel neutral proteinase. Images PMID:1569011

Structure of a gene encoding a murine thymus leukemia antigen, and organization of Tla genes in the BALB/c mouse

PubMed Central

1985-01-01

We have determined the DNA sequence of a gene encoding a thymus leukemia (TL) antigen in the BALB/c mouse, and have more definitively mapped the cloned BALB/c Tla-region class I gene clusters. Analysis of the sequence shows that the Tla gene is less closely related to the H-2 genes than H-2 genes are to one another or to a Qa-2,3-region genes. The Tla gene, 17.3A, contains an apparent gene conversion. Comparison of the BALB/c Tla genes with those from C57BL shows that BALB/c has more Tla-region class I genes, and that one of the genes absent in C57BL is gene 17.3A. PMID:3894562

Genome-Wide Identification and Expression Analysis of Homeodomain Leucine Zipper Subfamily IV (HDZ IV) Gene Family from Musa accuminata

PubMed Central

Pandey, Ashutosh; Misra, Prashant; Alok, Anshu; Kaur, Navneet; Sharma, Shivani; Lakhwani, Deepika; Asif, Mehar H.; Tiwari, Siddharth; Trivedi, Prabodh K.

2016-01-01

The homeodomain zipper family (HD-ZIP) of transcription factors is present only in plants and plays important role in the regulation of plant-specific processes. The subfamily IV of HDZ transcription factors (HD-ZIP IV) has primarily been implicated in the regulation of epidermal structure development. Though this gene family is present in all lineages of land plants, members of this gene family have not been identified in banana, which is one of the major staple fruit crops. In the present work, we identified 21 HDZIV encoding genes in banana by the computational analysis of banana genome resource. Our analysis suggested that these genes putatively encode proteins having all the characteristic domains of HDZIV transcription factors. The phylogenetic analysis of the banana HDZIV family genes further confirmed that after separation from a common ancestor, the banana, and poales lineages might have followed distinct evolutionary paths. Further, we conclude that segmental duplication played a major role in the evolution of banana HDZIV encoding genes. All the identified banana HDZIV genes expresses in different banana tissue, however at varying levels. The transcript levels of some of the banana HDZIV genes were also detected in banana fruit pulp, suggesting their putative role in fruit attributes. A large number of genes of this family showed modulated expression under drought and salinity stress. Taken together, the present work lays a foundation for elucidation of functional aspects of the banana HDZIV encoding genes and for their possible use in the banana improvement programs. PMID:26870050

Microarray Analyses of Gene Expression during Adventitious Root Development in Pinus contorta1[w

PubMed Central

Brinker, Monika; van Zyl, Leonel; Liu, Wenbin; Craig, Deborah; Sederoff, Ronald R.; Clapham, David H.; von Arnold, Sara

2004-01-01

In order to investigate the gene expression pattern during adventitious root development, RNA of Pinus contorta hypocotyls, pulse-treated with the auxin indole-3-butyric acid and harvested at distinct developmental time points of root development, was hybridized to microarrays containing 2,178 cDNAs from Pinus taeda. Over the period of observation of root development, the transcript levels of 220 genes changed significantly. During the root initiation phase, genes involved in cell replication and cell wall weakening and a transcript encoding a PINHEAD/ZWILLE-like protein were up-regulated, while genes related to auxin transport, photosynthesis, and cell wall synthesis were down-regulated. In addition, there were changes in transcript abundance of genes related to water stress. During the root meristem formation phase the transcript abundances of genes involved in auxin transport, auxin responsive transcription, and cell wall synthesis, and of a gene encoding a B-box zinc finger-like protein, increased, while those encoding proteins involved in cell wall weakening decreased. Changes of transcript abundance of genes related to water stress during the root meristem formation and root formation phase indicate that the plant roots had become functional in water transport. Simultaneously, genes involved in auxin transport were up-regulated, while genes related to cell wall modification were down-regulated. Finally, during the root elongation phase down-regulation of transcripts encoding proteins involved in cell replication and stress occurred. Based on the observed changes in transcript abundances, we suggest hypotheses about the relative importance of various physiological processes during the auxin-induced development of roots in P. contorta. PMID:15247392

Identification of a Linked Set of Genes in Serpulina hyodysenteriae (B204) Predicted To Encode Closely Related 39-Kilodalton Extracytoplasmic Proteins

PubMed Central

Gabe, Jeffrey D.; Dragon, Elizabeth; Chang, Ray-Jen; McCaman, Michael T.

1998-01-01

A tandem pair of nearly identical genes from Serpulina hyodysenteriae (B204) were cloned and sequenced. The full open reading frame of one gene and the partial open reading frame of the neighboring gene appear to encode secreted proteins which are homologous to, yet distinct from, the 39-kDa extracytoplasmic protein purified from the membrane fraction of S. hyodysenteriae. We have designated these newly identified genes vspA and vspB (for variable surface protein). PMID:9440540

Direct cloning of the trxB gene that encodes thioredoxin reductase.

PubMed Central

Russel, M; Model, P

1985-01-01

A strain was constructed which contains mutations in the genes encoding thioredoxin (trxA) and thioredoxin reductase (trxB) such that filamentous phage f1 cannot grow. The complementation of either mutation with its wild-type allele permits phage growth. We used this strain to select f1 phage which contain a cloned trxB gene. The location of the gene on the cloned fragment was determined, and its protein product was identified. Plasmid subclones that contain this gene overproduce thioredoxin reductase. Images PMID:2989245

Wound induced Beta vulgaris polygalacturonase-inhibiting protein genes encode a longer leucine-rich repeat domain and inhibit fungal polygalacturonases

USDA-ARS?s Scientific Manuscript database

Polygalacturonase-inhibiting proteins (PGIPs) are leucine-rich repeat (LRR) proteins involved in plant defense. Sugar beet (Beta vulgaris L.) PGIP genes, BvPGIP1, BvPGIP2 and BvPGIP3, were isolated from two breeding lines, F1016 and F1010. Full-length cDNA sequences of the three BvPGIP genes encod...

The role of genetics and antibodies in sepsis

PubMed Central

Giamarellos-Bourboulis, Evangelos J.

2016-01-01

During the course of sepsis when immunosuppression predominates, the concentrations of circulating immunoglobulins (IGs) are decreased and this is associated with adverse outcomes. The production of IGs as response to invasive bacterial pathogens takes place through a complex pathway starting from the recognition of the antigen (Ag) by innate immune cells that process and present Ags to T cells. The orchestration of T-helper (Th) lymphocyte responses directs specific B cells and ends with the production of IGs by plasma cells. All molecules implicated in this process are encoded by genes bearing single nucleotide polymorphisms (SNPs). Meta-analysis of case-control studies have shown that the carriage of minor frequency SNPs of CD14, TLR2 and TNF is associated with increased sepsis risk. The ambiguity of results of clinical trials studying the clinical efficacy of exogenous IG administration in sepsis suggests that efficacy of treatment should be considered after adjustment for SNPs of all implicated genes in the pathway of IG production. PMID:27713886

The role of genetics and antibodies in sepsis.

PubMed

Giamarellos-Bourboulis, Evangelos J; Opal, Steven M

2016-09-01

During the course of sepsis when immunosuppression predominates, the concentrations of circulating immunoglobulins (IGs) are decreased and this is associated with adverse outcomes. The production of IGs as response to invasive bacterial pathogens takes place through a complex pathway starting from the recognition of the antigen (Ag) by innate immune cells that process and present Ags to T cells. The orchestration of T-helper (Th) lymphocyte responses directs specific B cells and ends with the production of IGs by plasma cells. All molecules implicated in this process are encoded by genes bearing single nucleotide polymorphisms (SNPs). Meta-analysis of case-control studies have shown that the carriage of minor frequency SNPs of CD14 , TLR2 and TNF is associated with increased sepsis risk. The ambiguity of results of clinical trials studying the clinical efficacy of exogenous IG administration in sepsis suggests that efficacy of treatment should be considered after adjustment for SNPs of all implicated genes in the pathway of IG production.

PCSK1 Mutations and Human Endocrinopathies: From Obesity to Gastrointestinal Disorders.

PubMed

Stijnen, Pieter; Ramos-Molina, Bruno; O'Rahilly, Stephen; Creemers, John W M

2016-08-01

Prohormone convertase 1/3, encoded by the PCSK1 gene, is a serine endoprotease that is involved in the processing of a variety of proneuropeptides and prohormones. Humans who are homozygous or compound heterozygous for loss-of-function mutations in PCSK1 exhibit a variable and pleiotropic syndrome consisting of some or all of the following: obesity, malabsorptive diarrhea, hypogonadotropic hypogonadism, altered thyroid and adrenal function, and impaired regulation of plasma glucose levels in association with elevated circulating proinsulin-to-insulin ratio. Recently, more common variants in the PCSK1 gene have been found to be associated with alterations in body mass index, increased circulating proinsulin levels, and defects in glucose homeostasis. This review provides an overview of the endocrinopathies and other disorders observed in prohormone convertase 1/3-deficient patients, discusses the possible biochemical basis for these manifestations of the disease, and proposes a model whereby certain missense mutations in PCSK1 may result in proteins with a dominant negative action.

ACE insertion/deletion (I/D) polymorphism and diabetic nephropathy.

PubMed

Rahimi, Zohreh

2012-10-01

Angiotensin converting enzyme (ACE) gene encodes ACE, a key component of renin angiotensin system (RAS), plays an important role in blood pressure homeostasis by generating the vasoconstrictor peptide angiotensin II. Directory of Open Access Journals (DOAJ), Google Scholar, Pubmed (NLM), LISTA (EBSCO) and Web of Science have been searched. The presence of ACE insertion/deletion (I/D) polymorphism affects the plasma level of ACE. ACE DD genotype is associated with the highest systemic and renal ACE levels compared with the lowest ACE activity in carriers of II genotype. In this review focus has been performed on the study of ACE I/D polymorphism in various populations and its influence on the risk of onset and progression of diabetic nephropathy. Also, association between ACE I/D polymorphism and response to ACE inhibitor and angiotensin II receptor antagonists will be reviewed. Further, synergistic effect of this polymorphism and variants of some genes on the risk of development of diabetic nephropathy will be discussed.

Glycine and a Glycine Dehydrogenase (GLDC) SNP as Citalopram/Escitalopram Response Biomarkers in Depression: Pharmacometabolomics-informed Pharmacogenomics

PubMed Central

Ji, Yuan; Hebbring, Scott; Zhu, Hongjie; Jenkins, Gregory D; Biernacka, Joanna; Snyder, Karen; Drews, Maureen; Fiehn, Oliver; Zeng, Zhaobang; Schaid, Daniel; Mrazek, David A.; Kaddurah-Daouk, Rima; Weinshilboum, Richard M.

2011-01-01

Major depressive disorder (MDD) is a common psychiatric disease. Selective serotonin reuptake inhibitors (SSRIs) are an important class of drugs used to treat MDD. However, many patients do not respond adequately to SSRI therapy. We used a pharmacometabolomics-informed pharmacogenomic research strategy to identify citalopram/escitalopram treatment outcome biomarkers. Metabolomic assay of plasma samples from 20 escitalopram remitters and 20 non-remitters showed that glycine was negatively associated with treatment outcome (p=0.0054). That observation was pursued by genotyping tag single nucleotide polymorphisms (SNPs) for genes encoding glycine synthesis and degradation enzymes using 529 DNA samples from SSRI-treated MDD patients. The rs10975641 SNP in the glycine dehydrogenase gene was associated with treatment outcome phenotypes. Rs10975641 was then genotyped and was significant (p=0.02) in DNA from 1245 MDD patients in the STAR*D depression study. These results highlight both a possible role for glycine in SSRI response and the use of pharmacometabolomics to “inform” pharmacogenomics. PMID:21107318

COBRA encodes a putative GPI-anchored protein, which is polarly localized and necessary for oriented cell expansion in Arabidopsis.

PubMed

Schindelman, G; Morikami, A; Jung, J; Baskin, T I; Carpita, N C; Derbyshire, P; McCann, M C; Benfey, P N

2001-05-01

To control organ shape, plant cells expand differentially. The organization of the cellulose microfibrils in the cell wall is a key determinant of differential expansion. Mutations in the COBRA (COB) gene of Arabidopsis, known to affect the orientation of cell expansion in the root, are reported here to reduce the amount of crystalline cellulose in cell walls in the root growth zone. The COB gene, identified by map-based cloning, contains a sequence motif found in proteins that are anchored to the extracellular surface of the plasma membrane through a glycosylphosphatidylinositol (GPI) linkage. In animal cells, this lipid linkage is known to confer polar localization to proteins. The COB protein was detected predominately on the longitudinal sides of root cells in the zone of rapid elongation. Moreover, COB RNA levels are dramatically upregulated in cells entering the zone of rapid elongation. Based on these results, models are proposed for the role of COB as a regulator of oriented cell expansion.

COBRA encodes a putative GPI-anchored protein, which is polarly localized and necessary for oriented cell expansion in Arabidopsis

PubMed Central

Schindelman, Gary; Morikami, Atsushi; Jung, Jee; Baskin, Tobias I.; Carpita, Nicholas C.; Derbyshire, Paul; McCann, Maureen C.; Benfey, Philip N.

2001-01-01

To control organ shape, plant cells expand differentially. The organization of the cellulose microfibrils in the cell wall is a key determinant of differential expansion. Mutations in the COBRA (COB) gene of Arabidopsis, known to affect the orientation of cell expansion in the root, are reported here to reduce the amount of crystalline cellulose in cell walls in the root growth zone. The COB gene, identified by map-based cloning, contains a sequence motif found in proteins that are anchored to the extracellular surface of the plasma membrane through a glycosylphosphatidylinositol (GPI) linkage. In animal cells, this lipid linkage is known to confer polar localization to proteins. The COB protein was detected predominately on the longitudinal sides of root cells in the zone of rapid elongation. Moreover, COB RNA levels are dramatically upregulated in cells entering the zone of rapid elongation. Based on these results, models are proposed for the role of COB as a regulator of oriented cell expansion. PMID:11331607

Innate responses to gene knockouts impact overlapping gene networks and vary with respect to resistance to viral infection.

PubMed

Liu, Yonghong; Liu, Yuanyuan; Wu, Jiaming; Roizman, Bernard; Zhou, Grace Guoying

2018-04-03

Analyses of the levels of mRNAs encoding IFIT1, IFI16, RIG-1, MDA5, CXCL10, LGP2, PUM1, LSD1, STING, and IFNβ in cell lines from which the gene encoding LGP2, LSD1, PML, HDAC4, IFI16, PUM1, STING, MDA5, IRF3, or HDAC 1 had been knocked out, as well as the ability of these cell lines to support the replication of HSV-1, revealed the following: ( i ) Cell lines lacking the gene encoding LGP2, PML, or HDAC4 (cluster 1) exhibited increased levels of expression of partially overlapping gene networks. Concurrently, these cell lines produced from 5 fold to 12 fold lower yields of HSV-1 than the parental cells. ( ii ) Cell lines lacking the genes encoding STING, LSD1, MDA5, IRF3, or HDAC 1 (cluster 2) exhibited decreased levels of mRNAs of partially overlapping gene networks. Concurrently, these cell lines produced virus yields that did not differ from those produced by the parental cell line. The genes up-regulated in cell lines forming cluster 1, overlapped in part with genes down-regulated in cluster 2. The key conclusions are that gene knockouts and subsequent selection for growth causes changes in expression of multiple genes, and hence the phenotype of the cell lines cannot be ascribed to a single gene; the patterns of gene expression may be shared by multiple knockouts; and the enhanced immunity to viral replication by cluster 1 knockout cell lines but not by cluster 2 cell lines suggests that in parental cells, the expression of innate resistance to infection is specifically repressed.

Genomic characterization of an extensively-drug resistance Salmonella enterica serotype Indiana strain harboring blaNDM-1 gene isolated from a chicken carcass in China.

PubMed

Wang, Wei; Peng, Zixin; Baloch, Zulqarnain; Hu, Yujie; Xu, Jin; Zhang, Wenhui; Fanning, Séamus; Li, Fengqin

2017-11-01

The objective of this study was to genetically characterize the antimicrobial resistance mechanisms of Salmonella enterica serotype Indiana C629 isolated from a chicken carcass in China in 2014. Antimicrobial susceptibility against a panel of 23 antimicrobial agents was carried out on Salmonella enterica serotype Indiana C629 and assessed according to CLSI standards. Whole-genome sequencing of this isolate was conducted to obtain the complete genome of S. Indiana. Salmonella Indiana C629 expressed an XDR phenotype being resistant to more than 20 antimicrobial agents, including imipenem and meropenem. From the analysis of the resistance mechanisms, two mutations were identified in subunit A of DNA gyrase within the quinolone resistance determining region, in addition to the acquisition of mobile efflux pumps encoding oqxA/B/R. Additionally, four beta-lactamases resistance genes (bla CTX-M-65 , bla TEM-1 , bla OXA-1 , and bla NDM-1 ), five aminoglycosides resistance genes (aac(3)-IV, aac(6')-Ib-cr, aadA2, aadA5, and aph(4)-Ia), two phenicol resistance genes (catB3 and floR), and five trimethoprim/sulfamethoxazole resistance genes (sul1/2/3 and dfrA12/17) were also identified. A total of 191 virulence genes were identified. Among them, 57 belonged to type-three secretion system (T3SS) encoding genes, 55 belonged to fimbrial adherence encoding genes, and 39 belonged to flagella-encoding genes CONCLUSIONS: This study demonstrated that multi-resistance mechanisms consistent with an XDR-phenotype, along with various virulence encoding genes of a S. Indiana strain in China These findings highlight the importance of cooperation among different sectors in order to monitor the spread of resistant pathogens among food animal, foods of animal origin and human beings that might further take measures to protect consumers' health. Copyright © 2017 Elsevier GmbH. All rights reserved.

Molecular and functional characterization of novel fructosyltransferases and invertases from Agave tequilana.

PubMed

Cortés-Romero, Celso; Martínez-Hernández, Aída; Mellado-Mojica, Erika; López, Mercedes G; Simpson, June

2012-01-01

Fructans are the main storage polysaccharides found in Agave species. The synthesis of these complex carbohydrates relies on the activities of specific fructosyltransferase enzymes closely related to the hydrolytic invertases. Analysis of Agave tequilana transcriptome data led to the identification of ESTs encoding putative fructosyltransferases and invertases. Based on sequence alignments and structure/function relationships, two different genes were predicted to encode 1-SST and 6G-FFT type fructosyltransferases, in addition, 4 genes encoding putative cell wall invertases and 4 genes encoding putative vacuolar invertases were also identified. Probable functions for each gene, were assigned based on conserved amino acid sequences and confirmed for 2 fructosyltransferases and one invertase by analyzing the enzymatic activity of recombinant Agave protein s expressed and purified from Pichia pastoris. The genome organization of the fructosyltransferase/invertase genes, for which the corresponding cDNA contained the complete open reading frame, was found to be well conserved since all genes were shown to carry a 9 bp mini-exon and all showed a similar structure of 8 exons/7 introns with the exception of a cell wall invertase gene which has 7 exons and 6 introns. Fructosyltransferase genes were strongly expressed in the storage organs of the plants, especially in vegetative stages of development and to lower levels in photosynthetic tissues, in contrast to the invertase genes where higher levels of expression were observed in leaf tissues and in mature plants.

Molecular and Functional Characterization of Novel Fructosyltransferases and Invertases from Agave tequilana

PubMed Central

Cortés-Romero, Celso; Martínez-Hernández, Aída; Mellado-Mojica, Erika; López, Mercedes G.; Simpson, June

2012-01-01

Fructans are the main storage polysaccharides found in Agave species. The synthesis of these complex carbohydrates relies on the activities of specific fructosyltransferase enzymes closely related to the hydrolytic invertases. Analysis of Agave tequilana transcriptome data led to the identification of ESTs encoding putative fructosyltransferases and invertases. Based on sequence alignments and structure/function relationships, two different genes were predicted to encode 1-SST and 6G-FFT type fructosyltransferases, in addition, 4 genes encoding putative cell wall invertases and 4 genes encoding putative vacuolar invertases were also identified. Probable functions for each gene, were assigned based on conserved amino acid sequences and confirmed for 2 fructosyltransferases and one invertase by analyzing the enzymatic activity of recombinant Agave protein s expressed and purified from Pichia pastoris. The genome organization of the fructosyltransferase/invertase genes, for which the corresponding cDNA contained the complete open reading frame, was found to be well conserved since all genes were shown to carry a 9 bp mini-exon and all showed a similar structure of 8 exons/7 introns with the exception of a cell wall invertase gene which has 7 exons and 6 introns. Fructosyltransferase genes were strongly expressed in the storage organs of the plants, especially in vegetative stages of development and to lower levels in photosynthetic tissues, in contrast to the invertase genes where higher levels of expression were observed in leaf tissues and in mature plants. PMID:22558253

Impact assessment of bisphenol A on lignin-modifying enzymes by basidiomycete Trametes versicolor.

PubMed

Takamiya, Minako; Magan, Naresh; Warner, Philip J

2008-06-15

The impact of different concentrations of bisphenol A (BPA) was evaluated on growth of the white-rot basidiomycete, Trametes versicolor, and on the expression of genes encoding lignin-modifying enzyme (LME) activities. Effective doses (EDs) were obtained from fungal growth rate to monitor LME activities and the expression levels of their encoding genes. The fungus showed mycelial growth at concentrations of up to 300 microg ml(-1) of BPA with an ED50 value of 185 microg ml(-1). The LME activities were stimulated by BPA concentrations up to 300 microg ml(-1). The lignin peroxidase (LIP) encoding gene may be sensitive to BPA stress.

Evaluation of the effects of sdiA, a luxR homologue, on adherence and motility of Escherichia coli O157 : H7.

PubMed

Sharma, Vijay K; Bearson, Shawn M D; Bearson, Bradley L

2010-05-01

Quorum-sensing (QS) signalling pathways are important regulatory networks for controlling the expression of genes promoting adherence of enterohaemorrhagic Escherichia coli (EHEC) O157 : H7 to epithelial cells. A recent study has shown that EHEC O157 : H7 encodes a luxR homologue, called sdiA, which upon overexpression reduces the expression of genes encoding flagellar and locus of enterocyte effacement (LEE) proteins, thus negatively impacting on the motility and intimate adherence phenotypes, respectively. Here, we show that the deletion of sdiA from EHEC O157 : H7 strain 86-24, and from a hha (a negative regulator of ler) mutant of this strain, enhanced bacterial adherence to HEp-2 epithelial cells of the sdiA mutant strains relative to the strains containing a wild-type copy of sdiA. Quantitative reverse transcription PCR showed that the expression of LEE-encoded genes ler, espA and eae in strains with the sdiA deletions was not significantly different from that of the strains wild-type for sdiA. Similarly, no additional increases in the expression of LEE genes were observed in a sdiA hha double mutant strain relative to that observed in the hha deletion mutant. While the expression of fliC, which encodes flagellin, was enhanced in the sdiA mutant strain, the expression of fliC was reduced by several fold in the hha mutant strain, irrespective of the presence or absence of sdiA, indicating that the genes sdiA and hha exert opposing effects on the expression of fliC. The strains with deletions in sdiA or hha showed enhanced expression of csgA, encoding curlin of the curli fimbriae, with the expression of csgA highest in the sdiA hha double mutant, suggesting an additive effect of these two gene deletions on the expression of csgA. No significant differences were observed in the expression of the genes lpfA and fimA of the operons encoding long polar and type 1 fimbriae in the sdiA mutant strain. These data indicate that SdiA has no significant effect on the expression of LEE genes, but that it appears to act as a strong repressor of genes encoding flagella and curli fimbriae, and the alleviation of the SdiA-mediated repression of these genes in an EHEC O157 : H7 sdiA mutant strain contributes to enhanced bacterial motility and increased adherence to HEp-2 epithelial cells.

Molecular characterization and expression of the M6 gene of grass carp hemorrhage virus (GCHV), an aquareovirus.

PubMed

Qiu, T; Lu, R H; Zhang, J; Zhu, Z Y

2001-07-01

The complete nucleotide sequence of M6 gene of grass carp hemorrhage virus (GCHV) was determined. It is 2039 nucleotides in length and contains a single large open reading frame that could encode a protein of 648 amino acids with predicted molecular mass of 68.7 kDa. Amino acid sequence comparison revealed that the protein encoded by GCHV M6 is closely related to the protein mu1 of mammalian reovirus. The M6 gene, encoding the major outer-capsid protein, was expressed using the pET fusion protein vector in Escherichia coli and detected by Western blotting using chicken anti-GCHV immunoglobulin (IgY). The result indicates that the protein encoded by M6 may share a putative Asn-42-Pro-43 proteolytic cleavage site with mu1.

A [Cu]rious Ribosomal Profiling Pattern Leads to the Discovery of Ribosomal Frameshifting in the Synthesis of a Copper Chaperone.

PubMed

Atkins, John F; Loughran, Gary; Baranov, Pavel V

2017-01-19

In many bacteria, separate genes encode a copper binding chaperone and a copper efflux pump, but in some the chaperone encoding gene has been elusive. In this issue of Molecular Cell, Meydan et al. (2017) report that ribosomes translating the ORF that encodes the copper pump frequently frameshift and terminate to produce the copper chaperone. Copyright © 2017 Elsevier Inc. All rights reserved.

Transcriptome analysis of Neisseria meningitidis in human whole blood and mutagenesis studies identify virulence factors involved in blood survival.

PubMed

Echenique-Rivera, Hebert; Muzzi, Alessandro; Del Tordello, Elena; Seib, Kate L; Francois, Patrice; Rappuoli, Rino; Pizza, Mariagrazia; Serruto, Davide

2011-05-01

During infection Neisseria meningitidis (Nm) encounters multiple environments within the host, which makes rapid adaptation a crucial factor for meningococcal survival. Despite the importance of invasion into the bloodstream in the meningococcal disease process, little is known about how Nm adapts to permit survival and growth in blood. To address this, we performed a time-course transcriptome analysis using an ex vivo model of human whole blood infection. We observed that Nm alters the expression of ≈30% of ORFs of the genome and major dynamic changes were observed in the expression of transcriptional regulators, transport and binding proteins, energy metabolism, and surface-exposed virulence factors. In particular, we found that the gene encoding the regulator Fur, as well as all genes encoding iron uptake systems, were significantly up-regulated. Analysis of regulated genes encoding for surface-exposed proteins involved in Nm pathogenesis allowed us to better understand mechanisms used to circumvent host defenses. During blood infection, Nm activates genes encoding for the factor H binding proteins, fHbp and NspA, genes encoding for detoxifying enzymes such as SodC, Kat and AniA, as well as several less characterized surface-exposed proteins that might have a role in blood survival. Through mutagenesis studies of a subset of up-regulated genes we were able to identify new proteins important for survival in human blood and also to identify additional roles of previously known virulence factors in aiding survival in blood. Nm mutant strains lacking the genes encoding the hypothetical protein NMB1483 and the surface-exposed proteins NalP, Mip and NspA, the Fur regulator, the transferrin binding protein TbpB, and the L-lactate permease LctP were sensitive to killing by human blood. This increased knowledge of how Nm responds to adaptation in blood could also be helpful to develop diagnostic and therapeutic strategies to control the devastating disease cause by this microorganism.

Transcriptome Analysis of Neisseria meningitidis in Human Whole Blood and Mutagenesis Studies Identify Virulence Factors Involved in Blood Survival

PubMed Central

Del Tordello, Elena; Seib, Kate L.; Francois, Patrice; Rappuoli, Rino; Pizza, Mariagrazia; Serruto, Davide

2011-01-01

During infection Neisseria meningitidis (Nm) encounters multiple environments within the host, which makes rapid adaptation a crucial factor for meningococcal survival. Despite the importance of invasion into the bloodstream in the meningococcal disease process, little is known about how Nm adapts to permit survival and growth in blood. To address this, we performed a time-course transcriptome analysis using an ex vivo model of human whole blood infection. We observed that Nm alters the expression of ≈30% of ORFs of the genome and major dynamic changes were observed in the expression of transcriptional regulators, transport and binding proteins, energy metabolism, and surface-exposed virulence factors. In particular, we found that the gene encoding the regulator Fur, as well as all genes encoding iron uptake systems, were significantly up-regulated. Analysis of regulated genes encoding for surface-exposed proteins involved in Nm pathogenesis allowed us to better understand mechanisms used to circumvent host defenses. During blood infection, Nm activates genes encoding for the factor H binding proteins, fHbp and NspA, genes encoding for detoxifying enzymes such as SodC, Kat and AniA, as well as several less characterized surface-exposed proteins that might have a role in blood survival. Through mutagenesis studies of a subset of up-regulated genes we were able to identify new proteins important for survival in human blood and also to identify additional roles of previously known virulence factors in aiding survival in blood. Nm mutant strains lacking the genes encoding the hypothetical protein NMB1483 and the surface-exposed proteins NalP, Mip and NspA, the Fur regulator, the transferrin binding protein TbpB, and the L-lactate permease LctP were sensitive to killing by human blood. This increased knowledge of how Nm responds to adaptation in blood could also be helpful to develop diagnostic and therapeutic strategies to control the devastating disease cause by this microorganism. PMID:21589640

Simultaneous quantitation of sphingoid bases by UPLC-ESI-MS/MS with identical 13C-encoded internal standards.

PubMed

Mirzaian, M; Wisse, P; Ferraz, M J; Marques, A R A; Gaspar, P; Oussoren, S V; Kytidou, K; Codée, J D C; van der Marel, G; Overkleeft, H S; Aerts, J M

2017-03-01

Free sphingoid bases (lysosphingolipids) of primary storage sphingolipids are increased in tissues and plasma of several sphingolipidoses. As shown earlier by us, sphingoid bases can be accurately quantified using UPLC-ESI-MS/MS, particularly in combination with identical 13 C-encoded internal standards. The feasibility of simultaneous quantitation of sphingoid bases in plasma specimens spiked with a mixture of such standards is here described. The sensitivity and linearity of detection is excellent for all examined sphingoid bases (sphingosine, sphinganine, hexosyl-sphingosine (glucosylsphingosine), hexosyl 2 -sphingosine (lactosylsphingosine), hexosyl 3 -sphingosine (globotriaosylsphingosine), phosphorylcholine-sphingosine) in the relevant concentration range and the measurements show very acceptable intra- and inter-assay variation (<10% average). Plasma samples of a series of male and female Gaucher Disease and Fabry Disease patients were analyzed with the multiplex assay. The obtained data compare well to those earlier determined for plasma globotriaosylsphingosine and glucosylsphingosine in GD and FD patients. The same approach can be also applied to measure sphingolipids in the same sample. Following extraction of sphingolipids from the same sample these can be converted to sphingoid bases by microwave exposure and subsequently quantified using 13 C-encoded internal standards. Copyright © 2017 Elsevier B.V. All rights reserved.

Gene encoding acetyl-coenzyme A carboxylase

DOEpatents

Roessler, P.G.; Ohlrogge, J.B.

1996-09-24

A DNA encoding an acetyl-coenzyme A carboxylase (ACCase) from a photosynthetic organism and functional derivatives are disclosed which are resistant to inhibition from certain herbicides. This gene can be placed in organisms to increase their fatty acid content or to render them resistant to certain herbicides. 5 figs.

Mutations in iron-sulfur cluster proteins that improve xylose utilization

DOEpatents

Froehlich, Allan; Henningsen, Brooks; Covalla, Sean; Zelle, Rintze M.

2018-03-20

There is provided an engineered host cells comprising (a) one or more mutations in one or more endogenous genes encoding a protein associated with iron metabolism; and (b) at least one gene encoding a polypeptide having xylose isomerase activity, and methods of their use thereof.

Heterologous production and characterization of two glyoxal oxidases from Pycnoporus cinnabarinus

Treesearch

Marianne Daou; François Piumi; Daniel Cullen; Eric Record; Craig B. Faulds

2016-01-01

The genome of the white rot fungus Pycnoporus cinnabarinus includes a large number of genes encoding enzymes implicated in lignin degradation. Among these, three genes are predicted to encode glyoxal oxidase, an enzyme previously isolated from Phanerochaete chrysosporium. The glyoxal oxidase of P. chrysosporium...

Recent urbanization in China is correlated with a Westernized microbiome encoding increased virulence and antibiotic resistance genes.

PubMed

Winglee, Kathryn; Howard, Annie Green; Sha, Wei; Gharaibeh, Raad Z; Liu, Jiawu; Jin, Donghui; Fodor, Anthony A; Gordon-Larsen, Penny

2017-09-15

Urbanization is associated with an increased risk for a number of diseases, including obesity, diabetes, and cancer, which all also show associations with the microbiome. While microbial community composition has been shown to vary across continents and in traditional versus Westernized societies, few studies have examined urban-rural differences in neighboring communities within a single country undergoing rapid urbanization. In this study, we compared the gut microbiome, plasma metabolome, dietary habits, and health biomarkers of rural and urban people from a single Chinese province. We identified significant differences in the microbiota and microbiota-related plasma metabolites in rural versus recently urban subjects from the Hunan province of China. Microbes with higher relative abundance in Chinese urban samples have been associated with disease in other studies and were substantially more prevalent in the Human Microbiome Project cohort of American subjects. Furthermore, using whole metagenome sequencing, we found that urbanization was associated with a loss of microbial diversity and changes in the relative abundances of Viruses, Archaea, and Bacteria. Gene diversity, however, increased with urbanization, along with the proportion of reads associated with antibiotic resistance and virulence, which were strongly correlated with the presence of Escherichia and Shigella. Our data suggest that urbanization has produced convergent evolution of the gut microbial composition in American and urban Chinese populations, resulting in similar compositional patterns of abundant microbes through similar lifestyles on different continents, including a loss of potentially beneficial bacteria and an increase in potentially harmful genes via increased relative abundance of Escherichia and Shigella.

Effects of the commercial antiandrogen flutamide on the biomarkers of reproduction in male Murray rainbowfish (Melanotaenia fluviatilis).

PubMed

Bhatia, Harpreet; Kumar, Anupama; Ogino, Yukiko; Du, Jun; Gregg, Adrienne; Chapman, John; McLaughlin, Mike J; Iguchi, Taisen

2014-05-01

The endocrine responses in male Murray rainbowfish (Melanotaenia fluviatilis) were evaluated after exposures to biologically active concentrations of the nonsteroidal pharmaceutical, flutamide. Fish were exposed to nominal concentrations of 125 µg/L, 250 µg/L, 500 µg/L, and 1000 µg/L of flutamide for 7 d, after which plasma vitellogenin concentration; brain aromatase activity; and hepatic expression of the genes for vitellogenin, choriogenin, and androgen and estrogen receptors were assessed. Qualitative assessment of the testes of the fish exposed to flutamide exhibited hindrance in the transformation of spermatogonia to spermatozoa and increased testicular anomalies, such as multinucleated and pyknotic cells and interstitial fibrosis. An increase in the hepatosomatic index with respect to the controls was noted after treating the fish with flutamide at all concentrations. Vitellogenin was induced in plasma in the 1000 µg/L flutamide group. The activity of aromatase in the brain declined significantly after exposures to flutamide at all concentrations. Males exposed to 1000 µg/L of flutamide showed a downregulation in the genes encoding androgen receptors α and β. The expression of the gene for the estrogen receptor α was induced and of vitellogenin was downregulated after treatment with 250 µg/L to 1000 µg/L of flutamide. The results suggest that 7-d exposures to 125 µg/L to 1000 µg/L flutamide can impair the reproductive endocrine system in male Murray rainbowfish at multiple levels by an antiandrogenic mode of action. © 2014 SETAC.

Genomic polymorphism, recombination, and linkage disequilibrium in human major histocompatibility complex-encoded antigen-processing genes.

PubMed Central

van Endert, P M; Lopez, M T; Patel, S D; Monaco, J J; McDevitt, H O

1992-01-01

Recently, two subunits of a large cytosolic protease and two putative peptide transporter proteins were found to be encoded by genes within the class II region of the major histocompatibility complex (MHC). These genes have been suggested to be involved in the processing of antigenic proteins for presentation by MHC class I molecules. Because of the high degree of polymorphism in MHC genes, and previous evidence for both functional and polypeptide sequence polymorphism in the proteins encoded by the antigen-processing genes, we tested DNA from 27 consanguineous human cell lines for genomic polymorphism by restriction fragment length polymorphism (RFLP) analysis. These studies demonstrate a strong linkage disequilibrium between TAP1 and LMP2 RFLPs. Moreover, RFLPs, as well as a polymorphic stop codon in the telomeric TAP2 gene, appear to be in linkage disequilibrium with HLA-DR alleles and RFLPs in the HLA-DO gene. A high rate of recombination, however, seems to occur in the center of the complex, between the TAP1 and TAP2 genes. Images PMID:1360671

Functional differentiation and spatial-temporal co-expression networks of the NBS-encoding gene family in Jilin ginseng, Panax ginseng C.A. Meyer.

PubMed

Yin, Rui; Zhao, Mingzhu; Wang, Kangyu; Lin, Yanping; Wang, Yanfang; Sun, Chunyu; Wang, Yi; Zhang, Meiping

2017-01-01

Ginseng, Panax ginseng C.A. Meyer, is one of the most important medicinal plants for human health and medicine. It has been documented that over 80% of genes conferring resistance to bacteria, viruses, fungi and nematodes are contributed by the nucleotide binding site (NBS)-encoding gene family. Therefore, identification and characterization of NBS genes expressed in ginseng are paramount to its genetic improvement and breeding. However, little is known about the NBS-encoding genes in ginseng. Here we report genome-wide identification and systems analysis of the NBS genes actively expressed in ginseng (PgNBS genes). Four hundred twelve PgNBS gene transcripts, derived from 284 gene models, were identified from the transcriptomes of 14 ginseng tissues. These genes were classified into eight types, including TNL, TN, CNL, CN, NL, N, RPW8-NL and RPW8-N. Seven conserved motifs were identified in both the Toll/interleukine-1 receptor (TIR) and coiled-coil (CC) typed genes whereas six were identified in the RPW8 typed genes. Phylogenetic analysis showed that the PgNBS gene family is an ancient family, with a vast majority of its genes originated before ginseng originated. In spite of their belonging to a family, the PgNBS genes have functionally dramatically differentiated and been categorized into numerous functional categories. The expressions of the across tissues, different aged roots and the roots of different genotypes. However, they are coordinating in expression, forming a single co-expression network. These results provide a deeper understanding of the origin, evolution and functional differentiation and expression dynamics of the NBS-encoding gene family in plants in general and in ginseng particularly, and a NBS gene toolkit useful for isolation and characterization of disease resistance genes and for enhanced disease resistance breeding in ginseng and related species.

Functional differentiation and spatial-temporal co-expression networks of the NBS-encoding gene family in Jilin ginseng, Panax ginseng C.A. Meyer

PubMed Central

Wang, Kangyu; Lin, Yanping; Wang, Yanfang; Sun, Chunyu; Wang, Yi

2017-01-01

Ginseng, Panax ginseng C.A. Meyer, is one of the most important medicinal plants for human health and medicine. It has been documented that over 80% of genes conferring resistance to bacteria, viruses, fungi and nematodes are contributed by the nucleotide binding site (NBS)-encoding gene family. Therefore, identification and characterization of NBS genes expressed in ginseng are paramount to its genetic improvement and breeding. However, little is known about the NBS-encoding genes in ginseng. Here we report genome-wide identification and systems analysis of the NBS genes actively expressed in ginseng (PgNBS genes). Four hundred twelve PgNBS gene transcripts, derived from 284 gene models, were identified from the transcriptomes of 14 ginseng tissues. These genes were classified into eight types, including TNL, TN, CNL, CN, NL, N, RPW8-NL and RPW8-N. Seven conserved motifs were identified in both the Toll/interleukine-1 receptor (TIR) and coiled-coil (CC) typed genes whereas six were identified in the RPW8 typed genes. Phylogenetic analysis showed that the PgNBS gene family is an ancient family, with a vast majority of its genes originated before ginseng originated. In spite of their belonging to a family, the PgNBS genes have functionally dramatically differentiated and been categorized into numerous functional categories. The expressions of the across tissues, different aged roots and the roots of different genotypes. However, they are coordinating in expression, forming a single co-expression network. These results provide a deeper understanding of the origin, evolution and functional differentiation and expression dynamics of the NBS-encoding gene family in plants in general and in ginseng particularly, and a NBS gene toolkit useful for isolation and characterization of disease resistance genes and for enhanced disease resistance breeding in ginseng and related species. PMID:28727829

Studying the organization of genes encoding plant cell wall degrading enzymes in Chrysomela tremula provides insights into a leaf beetle genome.

PubMed

Pauchet, Y; Saski, C A; Feltus, F A; Luyten, I; Quesneville, H; Heckel, D G

2014-06-01

The ability of herbivorous beetles from the superfamilies Chrysomeloidea and Curculionoidea to degrade plant cell wall polysaccharides has only recently begun to be appreciated. The presence of plant cell wall degrading enzymes (PCWDEs) in the beetle's digestive tract makes this degradation possible. Sequences encoding these beetle-derived PCWDEs were originally identified from transcriptomes and strikingly resemble those of saprophytic and phytopathogenic microorganisms, raising questions about their origin; e.g. are they insect- or microorganism-derived? To demonstrate unambiguously that the genes encoding PCWDEs found in beetle transcriptomes are indeed of insect origin, we generated a bacterial artificial chromosome library from the genome of the leaf beetle Chrysomela tremula, containing 18 432 clones with an average size of 143 kb. After hybridizing this library with probes derived from 12 C. tremula PCWDE-encoding genes and sequencing the positive clones, we demonstrated that the latter genes are encoded by the insect's genome and are surrounded by genes possessing orthologues in the genome of Tribolium castaneum as well as in three other beetle genomes. Our analyses showed that although the level of overall synteny between C. tremula and T. castaneum seems high, the degree of microsynteny between both species is relatively low, in contrast to the more closely related Colorado potato beetle. © 2014 The Royal Entomological Society.

Identification of a Polymorphic Gene, BCL2A1, Encoding Two Novel Hematopoietic Lineage-specific Minor Histocompatibility Antigens

PubMed Central

Akatsuka, Yoshiki; Nishida, Tetsuya; Kondo, Eisei; Miyazaki, Mikinori; Taji, Hirohumi; Iida, Hiroatsu; Tsujimura, Kunio; Yazaki, Makoto; Naoe, Tomoki; Morishima, Yasuo; Kodera, Yoshihisa; Kuzushima, Kiyotaka; Takahashi, Toshitada

2003-01-01

We report the identification of two novel minor histocompatibility antigens (mHAgs), encoded by two separate single nucleotide polymorphisms on a single gene, BCL2A1, and restricted by human histocompatibility leukocyte antigen (HLA)-A*2402 (the most common HLA-A allele in Japanese) and B*4403, respectively. Two cytotoxic T lymphocyte (CTL) clones specific for these mHAgs were first isolated from two distinct recipients after hematopoietic cell transplantation. Both clones lyse only normal and malignant cells within the hematopoietic lineage. To localize the gene encoding the mHAgs, two-point linkage analysis was performed on the CTL lytic patterns of restricting HLA-transfected B lymphoblastoid cell lines obtained from Centre d'Etude du Polymorphisme Humain. Both CTL clones showed a completely identical lytic pattern for 4 pedigrees and the gene was localized within a 3.6-cM interval of 15q24.3–25.1 region that encodes at least 46 genes. Of those, only BCL2A1 has been reported to be expressed in hematopoietic cells and possess three nonsynonymous nucleotide changes. Minigene transfection and epitope reconstitution assays with synthetic peptides identified both HLA-A*2402– and B*4403-restricted mHAg epitopes to be encoded by distinct polymorphisms within BCL2A1. PMID:12771180

A Shigella flexneri Virulence Plasmid Encoded Factor Controls Production of Outer Membrane Vesicles

PubMed Central

Sidik, Saima; Kottwitz, Haila; Benjamin, Jeremy; Ryu, Julie; Jarrar, Ameer; Garduno, Rafael; Rohde, John R.

2014-01-01

Shigella spp. use a repertoire of virulence plasmid-encoded factors to cause shigellosis. These include components of a Type III Secretion Apparatus (T3SA) that is required for invasion of epithelial cells and many genes of unknown function. We constructed an array of 99 deletion mutants comprising all genes encoded by the virulence plasmid (excluding those known to be required for plasmid maintenance) of Shigella flexneri. We screened these mutants for their ability to bind the dye Congo red: an indicator of T3SA function. This screen focused our attention on an operon encoding genes that modify the cell envelope including virK, a gene of partially characterized function. We discovered that virK is required for controlled release of proteins to the culture supernatant. Mutations in virK result in a temperature-dependent overproduction of outer membrane vesicles (OMVs). The periplasmic chaperone/protease DegP, a known regulator of OMV production in Escherichia coli (encoded by a chromosomal gene), was found to similarly control OMV production in S. flexneri. Both virK and degP show genetic interactions with mxiD, a structural component of the T3SA. Our results are consistent with a model in which VirK and DegP relieve the periplasmic stress that accompanies assembly of the T3SA. PMID:25378474

Boron Toxicity Reduces Water Transport from Root to Shoot in Arabidopsis Plants. Evidence for a Reduced Transpiration Rate and Expression of Major PIP Aquaporin Genes.

PubMed

Macho-Rivero, Miguel A; Herrera-Rodríguez, M Begoña; Brejcha, Ramona; Schäffner, Anton R; Tanaka, Nobuhiro; Fujiwara, Toru; González-Fontes, Agustín; Camacho-Cristóbal, Juan J

2018-04-01

Toxic boron (B) concentrations cause impairments in several plant metabolic and physiological processes. Recently we reported that B toxicity led to a decrease in the transpiration rate of Arabidopsis plants in an ABA-dependent process within 24 h, which could indicate the occurrence of an adjustment of whole-plant water relations in response to this stress. Since plasma membrane intrinsic protein (PIP) aquaporins are key components influencing the water balance of plants because of their involvement in root water uptake and tissue hydraulic conductance, the aim of the present work was to study the effects of B toxicity on these important parameters affecting plant water status over a longer period of time. For this purpose, transpiration rate, water transport to the shoot and transcript levels of genes encoding four major PIP aquaporins were measured in Arabidopsis plants treated or not with a toxic B concentration. Our results indicate that, during the first 24 h of B toxicity, increased shoot ABA content would play a key role in reducing stomatal conductance, transpiration rate and, consequently, the water transport to the shoot. These physiological responses to B toxicity were maintained for up to 48 h of B toxicity despite shoot ABA content returning to control levels. In addition, B toxicity also caused the down-regulation of several genes encoding root and shoot aquaporins, which could reduce the cell to cell movement of water in plant tissues and, consequently, the water flux to shoot. All these changes in the water balance of plants under B toxicity could be a mechanism to prevent excess B accumulation in plant tissues.

The life-extending gene Indy encodes an exchanger for Krebs-cycle intermediates.

PubMed

Knauf, Felix; Mohebbi, Nilufar; Teichert, Carsten; Herold, Diana; Rogina, Blanka; Helfand, Stephen; Gollasch, Maik; Luft, Friedrich C; Aronson, Peter S

2006-07-01

A longevity gene called Indy (for 'I'm not dead yet'), with similarity to mammalian genes encoding sodium-dicarboxylate cotransporters, was identified in Drosophila melanogaster. Functional studies in Xenopus oocytes showed that INDY mediates the flux of dicarboxylates and citrate across the plasma membrane, but the specific transport mechanism mediated by INDY was not identified. To test whether INDY functions as an anion exchanger, we examined whether substrate efflux is stimulated by transportable substrates added to the external medium. Efflux of [14C]citrate from INDY-expressing oocytes was greatly accelerated by the addition of succinate to the external medium, indicating citrate-succinate exchange. The succinate-stimulated [14C]citrate efflux was sensitive to inhibition by DIDS (4,4'-di-isothiocyano-2,2'-disulphonic stilbene), as demonstrated previously for INDY-mediated succinate uptake. INDY-mediated efflux of [14C]citrate was also stimulated by external citrate and oxaloacetate, indicating citrate-citrate and citrate-oxaloacetate exchange. Similarly, efflux of [14C]succinate from INDY-expressing oocytes was stimulated by external citrate, alpha-oxoglutarate and fumarate, indicating succinate-citrate, succinate-alpha-oxoglutarate and succinate-fumarate exchange respectively. Conversely, when INDY-expressing Xenopus oocytes were loaded with succinate and citrate, [14C]succinate uptake was markedly stimulated, confirming succinate-succinate and succinate-citrate exchange. Exchange of internal anion for external citrate was markedly pH(o)-dependent, consistent with the concept that citrate is co-transported with a proton. Anion exchange was sodium-independent. We conclude that INDY functions as an exchanger of dicarboxylate and tricarboxylate Krebs-cycle intermediates. The effect of decreasing INDY activity, as in the long-lived Indy mutants, may be to alter energy metabolism in a manner that favours lifespan extension.

A novel family of integrases associated with prophages and genomic islands integrated within the tRNA-dihydrouridine synthase A (dusA) gene

PubMed Central

Farrugia, Daniel N.; Elbourne, Liam D. H.; Mabbutt, Bridget C.; Paulsen, Ian T.

2015-01-01

Genomic islands play a key role in prokaryotic genome plasticity. Genomic islands integrate into chromosomal loci such as transfer RNA genes and protein coding genes, whilst retaining various cargo genes that potentially bestow novel functions on the host organism. A gene encoding a putative integrase was identified at a single site within the 5′ end of the dusA gene in the genomes of over 200 bacteria. This integrase was discovered to be a component of numerous genomic islands, which appear to share a target site within the dusA gene. dusA encodes the tRNA-dihydrouridine synthase A enzyme, which catalyses the post-transcriptional reduction of uridine to dihydrouridine in tRNA. Genomic islands encoding homologous dusA-associated integrases were found at a much lower frequency within the related dusB and dusC genes, and non-dus genes. Excision of these dusA-associated islands from the chromosome as circularized intermediates was confirmed by polymerase chain reaction. Analysis of the dusA-associated islands indicated that they were highly diverse, with the integrase gene representing the only universal common feature. PMID:25883135

Transcript Profiling Identifies Gene Cohorts Controlled by Each Signal Regulating Trans-Differentiation of Epidermal Cells of Vicia faba Cotyledons to a Transfer Cell Phenotype

PubMed Central

Zhang, Hui-Ming; Wheeler, Simon L.; Xia, Xue; Colyvas, Kim; Offler, Christina E.; Patrick, John W.

2017-01-01

Transfer cells (TCs) support high rates of membrane transport of nutrients conferred by a plasma membrane area amplified by lining a wall labyrinth comprised of an uniform wall layer (UWL) upon which intricate wall ingrowth (WI) papillae are deposited. A signal cascade of auxin, ethylene, extracellular hydrogen peroxide (H2O2) and cytosolic Ca2+ regulates wall labyrinth assembly. To identify gene cohorts regulated by each signal, a RNA- sequencing study was undertaken using Vicia faba cotyledons. When cotyledons are placed in culture, their adaxial epidermal cells spontaneously undergo trans-differentiation to epidermal TCs (ETCs). Expressed genes encoding proteins central to wall labyrinth formation (signaling, intracellular organization, cell wall) and TC function of nutrient transport were assembled. Transcriptional profiles identified 9,742 annotated ETC-specific differentially expressed genes (DEGs; Log2fold change > 1; FDR p ≤ 0.05) of which 1,371 belonged to signaling (50%), intracellular organization (27%), cell wall (15%) and nutrient transporters (9%) functional categories. Expression levels of 941 ETC-specific DEGs were found to be sensitive to the known signals regulating ETC trans-differentiation. Significantly, signals acting alone, or in various combinations, impacted similar numbers of ETC-specific DEGs across the four functional gene categories. Amongst the signals acting alone, H2O2 exerted most influence affecting expression levels of 56% of the ETC-specific DEGs followed by Ca2+ (21%), auxin (18%) and ethylene (5%). The dominance by H2O2 was evident across all functional categories, but became more attenuated once trans-differentiation transitioned into WI papillae formation. Amongst the eleven signal combinations, H2O2/Ca2+ elicited the greatest impact across all functional categories accounting for 20% of the ETC-specific DEG cohort. The relative influence of the other signals acting alone, or in various combinations, varied across the four functional categories and two phases of wall labyrinth construction. These transcriptome data provide a powerful information platform from which to examine signal transduction pathways and how these regulate expression of genes encoding proteins engaged in intracellular organization, cell wall construction and nutrient transport. PMID:29234338

Transcript Profiling Identifies Gene Cohorts Controlled by Each Signal Regulating Trans-Differentiation of Epidermal Cells of Vicia faba Cotyledons to a Transfer Cell Phenotype.

PubMed

Zhang, Hui-Ming; Wheeler, Simon L; Xia, Xue; Colyvas, Kim; Offler, Christina E; Patrick, John W

2017-01-01

Transfer cells (TCs) support high rates of membrane transport of nutrients conferred by a plasma membrane area amplified by lining a wall labyrinth comprised of an uniform wall layer (UWL) upon which intricate wall ingrowth (WI) papillae are deposited. A signal cascade of auxin, ethylene, extracellular hydrogen peroxide (H 2 O 2 ) and cytosolic Ca 2+ regulates wall labyrinth assembly. To identify gene cohorts regulated by each signal, a RNA- sequencing study was undertaken using Vicia faba cotyledons. When cotyledons are placed in culture, their adaxial epidermal cells spontaneously undergo trans -differentiation to epidermal TCs (ETCs). Expressed genes encoding proteins central to wall labyrinth formation (signaling, intracellular organization, cell wall) and TC function of nutrient transport were assembled. Transcriptional profiles identified 9,742 annotated ETC-specific differentially expressed genes (DEGs; Log 2 fold change > 1; FDR p ≤ 0.05) of which 1,371 belonged to signaling (50%), intracellular organization (27%), cell wall (15%) and nutrient transporters (9%) functional categories. Expression levels of 941 ETC-specific DEGs were found to be sensitive to the known signals regulating ETC trans -differentiation. Significantly, signals acting alone, or in various combinations, impacted similar numbers of ETC-specific DEGs across the four functional gene categories. Amongst the signals acting alone, H 2 O 2 exerted most influence affecting expression levels of 56% of the ETC-specific DEGs followed by Ca 2+ (21%), auxin (18%) and ethylene (5%). The dominance by H 2 O 2 was evident across all functional categories, but became more attenuated once trans -differentiation transitioned into WI papillae formation. Amongst the eleven signal combinations, H 2 O 2 /Ca 2+ elicited the greatest impact across all functional categories accounting for 20% of the ETC-specific DEG cohort. The relative influence of the other signals acting alone, or in various combinations, varied across the four functional categories and two phases of wall labyrinth construction. These transcriptome data provide a powerful information platform from which to examine signal transduction pathways and how these regulate expression of genes encoding proteins engaged in intracellular organization, cell wall construction and nutrient transport.

Potassium supply and homeostasis in the osmotolerant non-conventional yeasts Zygosaccharomyces rouxii differ from Saccharomyces cerevisiae.

PubMed

Stříbný, Jiří; Kinclová-Zimmermannová, Olga; Sychrová, Hana

2012-12-01

Three different transport systems exist to accumulate a sufficient amount of potassium cations in yeasts. The most common of these are Trk-type transporters, which are used by all yeast species. Though most yeast species employ two different types of transporters, we only identified one gene encoding a potassium uptake system (Trk-type) in the genome of the highly osmotolerant yeast Zygosaccharomyces rouxii, and our results showed that ZrTrk1 is its major (and probably only) specific potassium uptake system. When expressed in Saccharomyces cerevisiae, the product of the ZrTRK1 gene is localized to the plasma membrane and its presence efficiently complements the phenotypes of S. cerevisiae trk1∆ trk2∆ cells. Deletion of the ZrTRK1 gene resulted in Z. rouxii cells being almost incapable of growth at low K(+) concentrations and it changed some cell physiological parameters in a way that differs from S. cerevisiae. In contrast to S. cerevisiae, Z. rouxii cells without the TRK1 gene contained less potassium than the control cells and their plasma membrane was significantly hyperpolarized compared with those of the parental strain when grown in the presence of 100 mM KCl. On the other hand, subsequent potassium starvation led to a substantial depolarization which is again different from S. cerevisiae. Plasma-membrane hyperpolarization did not prevent the efflux of potassium from Z. rouxii trk1Δ cells during potassium starvation, and the activity of ZrPma1 is less affected by the absence of ZrTRK1 than in S. cerevisiae. The use of a newly constructed Z. rouxii-specific plasmid for the expression of pHluorin showed that the intracellular pH of the Z. rouxii wild type and the trk1∆ mutant is not significantly different. Together with the fact that Z. rouxii cells contain a significantly lower amount of intracellular potassium than identically grown S. cerevisiae cells, our results suggest that this highly osmotolerant yeast species maintain its intracellular pH and potassium homeostasis in way(s) partially distinct from S. cerevisiae.

Gene disruption in Trichoderma atroviride via Agrobacterium-mediated transformation.

PubMed

Zeilinger, Susanne

2004-02-01

A modified Agrobacterium-mediated transformation method for the efficient disruption of two genes encoding signaling compounds of the mycoparasite Trichoderma atroviride is described, using the hph gene of Escherichia coli as selection marker. The transformation vectors contained about 1 kb of 5' and 3' non-coding regions from the tmk1 (encoding a MAP kinase) or tga3 (encoding an alpha-subunit of a heterotrimeric G protein) target loci flanking a selection marker. Transformation of fungal conidia and selection on hygromycin-containing media applying an overlay-based procedure, which overcomes the lack of formation of distinct single colonies by the fungus, led to stable clones for both disruption constructs. Southern and PCR analyses proved gene disruption by single-copy homologous integration with a frequency of approximately 60% for both genes; and the loss of tmk1 and tga3 transcript formation in the disruptants was demonstrated by RT-PCR.

Overexpression of the gene encoding HMG-CoA reductase in Saccharomyces cerevisiae for production of prenyl alcohols.

PubMed

Ohto, Chikara; Muramatsu, Masayoshi; Obata, Shusei; Sakuradani, Eiji; Shimizu, Sakayu

2009-04-01

To develop microbial production method for prenyl alcohols (e.g., (E,E)-farnesol (FOH), (E)-nerolidol (NOH), and (E,E,E)-geranylgeraniol (GGOH)), the genes encoding enzymes in the mevalonate and prenyl diphosphate pathways were overexpressed in Saccharomyces cerevisiae, and the resultant transformants were evaluated as to the production of these alcohols. Overexpression of the gene encoding hydroxymethylglutaryl (HMG)-CoA reductase was most effective among the genes tested. A derivative of S. cerevisiae ATCC 200589, which was selected through screening, was found to be the most suitable host for the production. On cultivation of the resultant transformant, in which the HMG-CoA reductase gene was overexpressed, in a 5-liter bench-scale jar fermenter for 7 d, the production of FOH, NOH, and GGOH reached 145.7, 98.8, and 2.46 mg/l, respectively.

prtH2, Not prtH, Is the Ubiquitous Cell Wall Proteinase Gene in Lactobacillus helveticus▿

PubMed Central

Genay, M.; Sadat, L.; Gagnaire, V.; Lortal, S.

2009-01-01

Lactobacillus helveticus strains possess an efficient proteolytic system that releases peptides which are essential for lactobacillus growth in various fermented dairy products and also affect textural properties or biological activities. Cell envelope proteinases (CEPs) are bacterial enzymes that hydrolyze milk proteins. In the case of L. helveticus, two CEPs with low percentages of amino acid identity have been described, i.e., PrtH and PrtH2. However, the distribution of the genes that encode CEPs still remains unclear, rendering it difficult to further control the formation of particular peptides. This study evaluated the diversity of genes that encode CEPs in a collection of strains of L. helveticus isolated from various biotopes, both in terms of the presence or absence of these genes and in terms of nucleotide sequence, and studied their transcription in dairy matrices. After defining three sets of primers for both the prtH and prtH2 genes, we studied the distribution of the genes by using PCR and Southern blotting experiments. The prtH2 gene was ubiquitous in the 29 strains of L. helveticus studied, whereas only 18 of them also exhibited the prtH gene. Sequencing of a 350-bp internal fragment of these genes revealed the existence of intraspecific diversity. Finally, expression of these two CEP-encoding genes was followed during the growth in dairy matrices of two strains, ITG LH77 and CNRZ32, which possess one and two CEP-encoding genes, respectively. Both genes were shown to be expressed by L. helveticus at each stage of growth in milk and at different stages of mini-Swiss-type cheese making and ripening. PMID:19286786

The Arabidopsis thaliana ortholog of a purported maize cholinesterase gene encodes a GDSL-lipase

PubMed Central

Muralidharan, Mrinalini; Buss, Kristina; Larrimore, Katherine E.; Segerson, Nicholas A.; Kannan, Latha

2013-01-01

Acetylcholinesterase is an enzyme that is intimately associated with regulation of synaptic transmission in the cholinergic nervous system and in neuromuscular junctions of animals. However the presence of cholinesterase activity has been described also in non-metazoan organisms such as slime molds, fungi and plants. More recently, a gene purportedly encoding for acetylcholinesterase was cloned from maize. We have cloned the Arabidopsis thaliana homolog of the Zea mays gene, At3g26430, and studied its biochemical properties. Our results indicate that the protein encoded by the gene exhibited lipase activity with preference to long chain substrates but did not hydrolyze choline esters. The At3g26430 protein belongs to the SGNH clan of serine hydrolases, and more specifically to the GDS(L) lipase family. PMID:23430565

Genetic analysis reveals the identity of the photoreceptor for phototaxis in hormogonium filaments of Nostoc punctiforme.

PubMed

Campbell, Elsie L; Hagen, Kari D; Chen, Rui; Risser, Douglas D; Ferreira, Daniela P; Meeks, John C

2015-02-15

In cyanobacterial Nostoc species, substratum-dependent gliding motility is confined to specialized nongrowing filaments called hormogonia, which differentiate from vegetative filaments as part of a conditional life cycle and function as dispersal units. Here we confirm that Nostoc punctiforme hormogonia are positively phototactic to white light over a wide range of intensities. N. punctiforme contains two gene clusters (clusters 2 and 2i), each of which encodes modular cyanobacteriochrome-methyl-accepting chemotaxis proteins (MCPs) and other proteins that putatively constitute a basic chemotaxis-like signal transduction complex. Transcriptional analysis established that all genes in clusters 2 and 2i, plus two additional clusters (clusters 1 and 3) with genes encoding MCPs lacking cyanobacteriochrome sensory domains, are upregulated during the differentiation of hormogonia. Mutational analysis determined that only genes in cluster 2i are essential for positive phototaxis in N. punctiforme hormogonia; here these genes are designated ptx (for phototaxis) genes. The cluster is unusual in containing complete or partial duplicates of genes encoding proteins homologous to the well-described chemotaxis elements CheY, CheW, MCP, and CheA. The cyanobacteriochrome-MCP gene (ptxD) lacks transmembrane domains and has 7 potential binding sites for bilins. The transcriptional start site of the ptx genes does not resemble a sigma 70 consensus recognition sequence; moreover, it is upstream of two genes encoding gas vesicle proteins (gvpA and gvpC), which also are expressed only in the hormogonium filaments of N. punctiforme. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

TRPP2-dependent Ca2+ signaling in dorso-lateral mesoderm is required for kidney field establishment in Xenopus.

PubMed

Futel, Mélinée; Leclerc, Catherine; Le Bouffant, Ronan; Buisson, Isabelle; Néant, Isabelle; Umbhauer, Muriel; Moreau, Marc; Riou, Jean-François

2015-03-01

In Xenopus laevis embryos, kidney field specification is dependent on retinoic acid (RA) and coincides with a dramatic increase of Ca(2+) transients, but the role of Ca(2+) signaling in the kidney field is unknown. Here, we identify TRPP2, a member of the transient receptor potential (TRP) superfamily of channel proteins encoded by the pkd2 gene, as a central component of Ca(2+) signaling in the kidney field. TRPP2 is strongly expressed at the plasma membrane where it might regulate extracellular Ca(2+) entry. Knockdown of pkd2 in the kidney field results in the downregulation of pax8, but not of other kidney field genes (lhx1, osr1 and osr2). We further show that inhibition of Ca(2+) signaling with an inducible Ca(2+) chelator also causes downregulation of pax8, and that pkd2 knockdown results in a severe inhibition of Ca(2+) transients in kidney field explants. Finally, we show that disruption of RA results both in an inhibition of intracellular Ca(2+) signaling and of TRPP2 incorporation into the plasma membrane of kidney field cells. We propose that TRPP2-dependent Ca(2+) signaling is a key component of pax8 regulation in the kidney field downstream of RA-mediated non-transcriptional control of TRPP2. © 2015. Published by The Company of Biologists Ltd.

Reversal of diabetic vasculopathy in a rat model of type 1 diabetes by opiorphin-related peptides

PubMed Central

Calenda, Giulia; Tong, Yuehong; Kanika, Nirmala D.; Tar, Moses T.; Suadicani, Sylvia O.; Zhang, Xinhua; Melman, Arnold; Rougeot, Catherine

2011-01-01

Diabetes results in a myriad of vascular complications, often referred to as diabetic vasculopathy, which encompasses both microvascular [erectile dysfunction (ED), retinopathy, neuropathy, and nephropathy] and macrovascular complications (hypertension, coronary heart disease, and myocardial infarction). In diabetic animals and patients with ED, there is decreased opiorphin or opiorphin-related gene expression in corporal tissue. Both opiorphin and the rat homologous peptide sialorphin are found circulating in the plasma. In the present study, we investigated if diabetes induced changes in plasma sialorphin levels and if changes in these levels could modulate the biochemistry and physiology of vascular smooth muscle. We show that circulating sialorphin levels are reduced in a rat model of type I diabetes. Intracorporal injection of plasmids expressing sialorphin into diabetic rats restores sialorphin levels to those seen in the blood of nondiabetic animals and results in both improved erectile function and blood pressure. Sialorphin modulated the ability of C-type natriuretic peptide to relax both corporal and aortic smooth muscle strips and of bradykinin to regulate intracellular calcium levels in both corporal and aortic smooth muscle cells. We have previously shown that expression of genes encoding opiorphins is increased when erectile function is improved. Our findings thus suggest that by affecting circulating levels of opiorphin-related peptides, proper erectile function is not only an indicator but also a modulator of overall vascular health of a man. PMID:21784987

Reversal of diabetic vasculopathy in a rat model of type 1 diabetes by opiorphin-related peptides.

PubMed

Calenda, Giulia; Tong, Yuehong; Kanika, Nirmala D; Tar, Moses T; Suadicani, Sylvia O; Zhang, Xinhua; Melman, Arnold; Rougeot, Catherine; Davies, Kelvin P

2011-10-01

Diabetes results in a myriad of vascular complications, often referred to as diabetic vasculopathy, which encompasses both microvascular [erectile dysfunction (ED), retinopathy, neuropathy, and nephropathy] and macrovascular complications (hypertension, coronary heart disease, and myocardial infarction). In diabetic animals and patients with ED, there is decreased opiorphin or opiorphin-related gene expression in corporal tissue. Both opiorphin and the rat homologous peptide sialorphin are found circulating in the plasma. In the present study, we investigated if diabetes induced changes in plasma sialorphin levels and if changes in these levels could modulate the biochemistry and physiology of vascular smooth muscle. We show that circulating sialorphin levels are reduced in a rat model of type I diabetes. Intracorporal injection of plasmids expressing sialorphin into diabetic rats restores sialorphin levels to those seen in the blood of nondiabetic animals and results in both improved erectile function and blood pressure. Sialorphin modulated the ability of C-type natriuretic peptide to relax both corporal and aortic smooth muscle strips and of bradykinin to regulate intracellular calcium levels in both corporal and aortic smooth muscle cells. We have previously shown that expression of genes encoding opiorphins is increased when erectile function is improved. Our findings thus suggest that by affecting circulating levels of opiorphin-related peptides, proper erectile function is not only an indicator but also a modulator of overall vascular health of a man.

Mutation of the Membrane-Associated M1 Protease APM1 Results in Distinct Embryonic and Seedling Developmental Defects in Arabidopsis[C][W

PubMed Central

Peer, Wendy Ann; Hosein, Fazeeda N.; Bandyopadhyay, Anindita; Makam, Srinivas N.; Otegui, Marisa S.; Lee, Gil-Je; Blakeslee, Joshua J.; Cheng, Yan; Titapiwatanakun, Boosaree; Yakubov, Bahktiyor; Bangari, Bharat; Murphy, Angus S.

2009-01-01

Aminopeptidase M1 (APM1), a single copy gene in Arabidopsis thaliana, encodes a metallopeptidase originally identified via its affinity for, and hydrolysis of, the auxin transport inhibitor 1-naphthylphthalamic acid (NPA). Mutations in this gene result in haploinsufficiency. Loss-of-function mutants show irregular, uncoordinated cell divisions throughout embryogenesis, affecting the shape and number of cotyledons and the hypophysis, and is seedling lethal at 5 d after germination due to root growth arrest. Quiescent center and cell cycle markers show no signals in apm1-1 knockdown mutants, and the ground tissue specifiers SHORTROOT and SCARECROW are misexpressed or mislocalized. apm1 mutants have multiple, fused cotyledons and hypocotyls with enlarged epidermal cells with cell adhesion defects. apm1 alleles show defects in gravitropism and auxin transport. Gravistimulation decreases APM1 expression in auxin-accumulating root epidermal cells, and auxin treatment increases expression in the stele. On sucrose gradients, APM1 occurs in unique light membrane fractions. APM1 localizes at the margins of Golgi cisternae, plasma membrane, select multivesicular bodies, tonoplast, dense intravacuolar bodies, and maturing metaxylem cells. APM1 associates with brefeldin A–sensitive endomembrane structures and the plasma membrane in cortical and epidermal cells. The auxin-related phenotypes and mislocalization of auxin efflux proteins in apm1 are consistent with biochemical interactions between APM1 and NPA. PMID:19531600

Identification of an Amino Acid Domain Encoded by the Capsid Protein Gene of Porcine Circovirus Type 2 that Modulates Viral Protein Distribution During Replication

USDA-ARS?s Scientific Manuscript database

Previous work showed that distinct amino acid motifs are encoded by the Rep, Cap and ORF3 genes of two subgroups of porcine circoviruses (PCV), PCV2a and PCV2b. At a specific location of the gene, a certain amino acid residue or sequence is preferred. Specifically, two amino acid domains located in ...

Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs

PubMed Central

MIYAGAWA, Shuji; MATSUNARI, Hitomi; WATANABE, Masahito; NAKANO, Kazuaki; UMEYAMA, Kazuhiro; SAKAI, Rieko; TAKAYANAGI, Shuko; TAKEISHI, Toki; FUKUDA, Tooru; YASHIMA, Sayaka; MAEDA, Akira; EGUCHI, Hiroshi; OKUYAMA, Hiroomi; NAGAYA, Masaki; NAGASHIMA, Hiroshi

2015-01-01

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs. PMID:26227017

Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs.

PubMed

Miyagawa, Shuji; Matsunari, Hitomi; Watanabe, Masahito; Nakano, Kazuaki; Umeyama, Kazuhiro; Sakai, Rieko; Takayanagi, Shuko; Takeishi, Toki; Fukuda, Tooru; Yashima, Sayaka; Maeda, Akira; Eguchi, Hiroshi; Okuyama, Hiroomi; Nagaya, Masaki; Nagashima, Hiroshi

2015-01-01

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs.

Effect of sypQ gene on poly-N-acetylglucosamine biosynthesis in Vibrio parahaemolyticus and its role in infection process.

PubMed

Ye, Libin; Zheng, Xiaolin; Zheng, Hongjian

2014-04-01

The syp locus includes four genes encoding putative regulators, six genes encoding glycosyltransferases, two encoding export proteins, and six other genes encoding unidentified functional proteins associated with biofilm formation and symbiotic colonization. However, the individual functions of the respective genes remain unclear. Amino acid alignment indicates that sypQ is presumably involved in biosynthesizing poly-N-acetylglucosamine (PNAG), which is proposed to be a critical virulence factor in pathogen infection and is regarded as a target for protective immunity against a variety of Gram-negative/positive pathogens. However, no evidence showing that Vibrio parahaemolyticus also produces PNAG has been reported. Herein, the V. parahaemolyticus is confirmed to possess potential for producing PNAG for the first time. Our results indicated that gene sypQ is associated with PNAG biosynthesis and PNAG is involved in pathogen colonization. We propose that the function of pgaC in Escherichia coli could be taken over by sypQ from V. parahaemolyticus. We also tested whether PNAG can be used as a target against V. parahaemolyticus when it infects Pseudosciaena crocea. Our results showed that PNAG isolated from V. parahaemolyticus is an effective agent for decreasing V. parahaemolyticus invasion, implying that PNAG could be used to develop an effective vaccine against V. parahaemolyticus infection.

Development of a polymerase chain reaction to distinguish monocellate cobra (Naja khouthia) bites from other common Thai snake species, using both venom extracts and bite-site swabs.

PubMed

Suntrarachun, S; Pakmanee, N; Tirawatnapong, T; Chanhome, L; Sitprija, V

2001-07-01

A PCR technique was used in this study to identify and distinguish monocellate cobra snake bites using snake venoms and swab specimens from snake bite-sites in mice from bites by other common Thai snakes. The sequences of nucleotide primers were selected for the cobrotoxin-encoding gene from the Chinese cobra (Naja atra) since the sequences of monocellate cobra (Naja kaouthia) venom are still unknown. However, the 113-bp fragment of cDNA of the cobrotoxin-encoding gene was detected in the monocellate cobra venom using RT-PCR. This gene was not found in the venoms of Ophiophagus hannah (king cobra), Bungarus fasciatus (banded krait), Daboia russelii siamensis (Siamese Russell's Viper, and Calloselasma rhodostoma (Malayan pit viper). Moreover, direct PCR could detect a 665-bp fragment of the cobrotoxin-encoding gene in the monocellate cobra venom but not the other snake venoms. Likewise, this gene was only observed in swab specimens from cobra snake bite-sites in mice. This is the first report demonstrating the ability of PCR to detect the cobrotoxin-encoding gene from snake venoms and swab specimens. Further studies are required for identification of this and other snakes from the bite-sites on human skin.

Transposon mutagenesis and cloning analysis of the pathways for degradation of 2,4-dichlorophenoxyacetic acid and 3-chlorobenzoate in Alcaligenes eutrophus JMP134(pJP4).

PubMed Central

Don, R H; Weightman, A J; Knackmuss, H J; Timmis, K N

1985-01-01

Plasmid pJP4 permits its host bacterium, strain JMP134, to degrade and utilize as sole sources of carbon and energy 3-chlorobenzoate and 2,4-dichlorophenoxyacetic acid (R. H. Don and J. M. Pemberton, J. Bacteriol. 145:681-686, 1981). Mutagenesis of pJP4 by transposons Tn5 and Tn1771 enabled localization of five genes for enzymes involved in these catabolic pathways. Four of the genes, tfdB, tfdC, tfdD, and tfdE, encoded 2,4-dichlorophenol hydroxylase, dichlorocatechol 1,2-dioxygenase, chloromuconate cycloisomerase, and chlorodienelactone hydrolase, respectively. No function has been assigned to the fifth gene, tfdF, although it may encode a trans-chlorodiene-lactone isomerase. Inactivation of genes tfdC, tfdD, and tfdE, which encode the transformation of dichlorocatechol to chloromaleylacetic acid, prevented host strain JMP134 from degrading both 3-chlorobenzoate and 2,4-dichlorophenoxyacetic acid, which indicates that the pathways for these two substrates utilize common enzymes for the dissimilation of chlorocatechols. Studies with cloned catabolic genes from pJP4 indicated that whereas all essential steps in the degradation of 2,4-dichlorophenoxyacetic acid are plasmid encoded, the conversion of 3-chlorobenzoate to chlorocatechol is specified by chromosomal genes. PMID:2981813

Parallel loss of nuclear-encoded mitochondrial aminoacyl-tRNA synthetases and mtDNA-encoded tRNAs in Cnidaria.

PubMed

Haen, Karri M; Pett, Walker; Lavrov, Dennis V

2010-10-01

Unlike most animal mitochondrial (mt) genomes, which encode a set of 22 transfer RNAs (tRNAs) sufficient for mt protein synthesis, those of cnidarians have only retained one or two tRNA genes. Whether the missing cnidarian mt-tRNA genes relocated outside the main mt chromosome or were lost remains unclear. It is also unknown what impact the loss of tRNA genes had on other components of the mt translational machinery. Here, we explored the nuclear genome of the cnidarian Nematostella vectensis for the presence of mt-tRNA genes and their corresponding mt aminoacyl-tRNA synthetases (mt-aaRS). We detected no candidates for mt-tRNA genes and only two mt-aaRS orthologs. At the same time, we found that all but one cytosolic aaRS appear to be targeted to mitochondria. These results indicate that the loss of mt-tRNAs in Cnidaria is genuine and occurred in parallel with the loss of nuclear-encoded mt-aaRS. Our phylogenetic analyses of individual aaRS revealed that although the nearly total loss of mt-aaRS is rare, aaRS gene deletion and replacement have occurred throughout the evolution of Metazoa.

Expression and characterization of hyperthermostable exo-polygalacturonase RmGH28 from Rhodothermus marinus

USDA-ARS?s Scientific Manuscript database

The gene RmGH28 from the organism Rhodothermus marinus putatively encoding a glycosyl hydrolase family 28 polygalacturonase was expressed in E. coli, and the enzyme purified and biochemically characterized. The gene was found to encode an exo- polygalacturonase, with galacturonic acid monomer and th...

Genome-wide comparative analysis of NBS-encoding genes in four Gossypium species

USDA-ARS?s Scientific Manuscript database

Nucleotide binding site (NBS) genes encode a large family of disease resistance (R) proteins in plants. The availability of genomic data of the two diploid cotton species, Gossypium arboreum and Gossypium raimondii, and the two allotetraploid cotton species, Gossypium hirsutum (TM-1) and Gossypium ...

The pine Pschi4 promoter directs wound-induced transcription

Treesearch

Haiguo Wu; Charles H. Michler; Liborio LaRussa; John M. Davis

1999-01-01

Mechanical wounding stimulates the accumulation of Pschi4 transcripts (encoding a putative extracellular chitinase) in pine trees. To gain insight into the transcriptional regulatory region(s) in this gymnosperm defense gene, the 5'-flanking region of Pschi4 was fused to the uidA reporter gene encoding -...

Amino acid substitutions in the VanS sensor of the VanA-type vancomycin-resistant Enterococcus strains result in high-level vancomycin resistance and low-level teicoplanin resistance.

PubMed

Hashimoto, Y; Tanimoto, K; Ozawa, Y; Murata, T; Ike, Y

2000-04-15

The vancomycin-resistant enterococci GV1, GV2 and GV3, which were isolated from droppings from broiler farms in Japan have been characterized as VanA-type VRE, which express high-level vancomycin resistance (256 or 512 microg ml(-1), MIC) and low-level teicoplanin resistance (1 or 2 microg ml(-1), MIC). The vancomycin resistances were encoded on plasmids. The vancomycin resistance conjugative plasmid pMG2 was isolated from the GV2 strain. The VanA determinant of pMG2 showed the same genetic organization as that of the VanA genes encoded on the representative transposon Tn1546, which comprises vanRSHAXYZ. The nucleotide sequences of all the genes, except the gene related to the vanS gene on Tn1546, were completely identical to the genes encoded on Tn1546. Three amino acid substitutions in the N-terminal region of the deduced VanS were detected in the nucleotide sequence of vanS encoded on pMG2. There were also three amino acid substitutions in the vanS gene of the GV1 and GV3 strains in the same positions as in the vanS gene of pMG2. Vancomycin induced the increased teicoplanin resistance in these strains.

[Cloning, mutagenesis and symbiotic phenotype of three lipid transfer protein encoding genes from Mesorhizobium huakuii 7653R].

PubMed

Li, Yanan; Zeng, Xiaobo; Zhou, Xuejuan; Li, Youguo

2016-12-04

Lipid transfer protein superfamily is involved in lipid transport and metabolism. This study aimed to construct mutants of three lipid transfer protein encoding genes in Mesorhizobium huakuii 7653R, and to study the phenotypes and function of mutations during symbiosis with Astragalus sinicus. We used bioinformatics to predict structure characteristics and biological functions of lipid transfer proteins, and conducted semi-quantitative and fluorescent quantitative real-time PCR to analyze the expression levels of target genes in free-living and symbiotic conditions. Using pK19mob insertion mutagenesis to construct mutants, we carried out pot plant experiments to observe symbiotic phenotypes. MCHK-5577, MCHK-2172 and MCHK-2779 genes encoding proteins belonged to START/RHO alpha_C/PITP/Bet_v1/CoxG/CalC (SRPBCC) superfamily, involved in lipid transport or metabolism, and were identical to M. loti at 95% level. Gene relative transcription level of the three genes all increased compared to free-living condition. We obtained three mutants. Compared with wild-type 7653R, above-ground biomass of plants and nodulenitrogenase activity induced by the three mutants significantly decreased. Results indicated that lipid transfer protein encoding genes of Mesorhizobium huakuii 7653R may play important roles in symbiotic nitrogen fixation, and the mutations significantly affected the symbiotic phenotypes. The present work provided a basis to study further symbiotic function mechanism associated with lipid transfer proteins from rhizobia.

Molecular characterization and analysis of the acrB gene of Aspergillus nidulans: a gene identified by genetic interaction as a component of the regulatory network that includes the CreB deubiquitination enzyme.

PubMed Central

Boase, Natasha A; Lockington, Robin A; Adams, Julian R J; Rodbourn, Louise; Kelly, Joan M

2003-01-01

Mutations in the acrB gene, which were originally selected through their resistance to acriflavine, also result in reduced growth on a range of sole carbon sources, including fructose, cellobiose, raffinose, and starch, and reduced utilization of omega-amino acids, including GABA and beta-alanine, as sole carbon and nitrogen sources. The acrB2 mutation suppresses the phenotypic effects of mutations in the creB gene that encodes a regulatory deubiquitinating enzyme, and in the creC gene that encodes a WD40-repeat-containing protein. Thus AcrB interacts with a regulatory network controlling carbon source utilization that involves ubiquitination and deubiquitination. The acrB gene was cloned and physically analyzed, and it encodes a novel protein that contains three putative transmembrane domains and a coiled-coil region. AcrB may play a role in the ubiquitination aspect of this regulatory network. PMID:12750323

Cloning and characterization of the nagA gene that encodes beta-n-acetylglucosaminidase from Aspergillus nidulans and its expression in Aspergillus oryzae.

PubMed

Kim, Sunhwa; Matsuo, Ichiro; Ajisaka, Katsumi; Nakajima, Harushi; Kitamoto, Katsuhiko

2002-10-01

We isolated a beta-N-acetylglucosaminidase encoding gene and its cDNA from the filamentous fungus Aspergillus nidulans, and designated it nagA. The nagA gene contained no intron and encoded a polypeptide of 603 amino acids with a putative 19-amino acid signal sequence. The deduced amino acid sequence was very similar to the sequence of Candida albicans Hex1 and Trichoderma harzianum Nag1. Yeast cells containing the nagA cDNA under the control of the GAL1 promoter expressed beta-N-acetylglucosaminidase activity. The chromosomal nagA gene of A. nidulans was disrupted by replacement with the argB marker gene. The disruptant strains expressed low levels of beta-N-acetylglucosaminidase activity and showed poor growth on a medium containing chitobiose as a carbon source. Aspergillus oryzae strain carrying the nagA gene under the control of the improved glaA promoter produced large amounts of beta-N-acetylglucosaminidase in a wheat bran solid culture.

Cloning and heterologous expression of the antibiotic peptide (ABP) genes from Rhizopus oligosporus NBRC 8631.

PubMed

Yamada, Osamu; Sakamoto, Kazutoshi; Tominaga, Mihoko; Nakayama, Tasuku; Koseki, Takuya; Fujita, Akiko; Akita, Osamu

2005-03-01

We carried out protein sequencing of purified Antibiotic Peptide (ABP), and cloned two genes encoding this peptide as abp1 and abp2, from Rhizopus oligosporus NBRC 8631. Both genes contain an almost identical 231-bp segment, with only 3 nucleotide substitutions, encoding a 77 amino acid peptide. The abp gene product comprises a 28 amino acid signal sequence and a 49 amino acid mature peptide. Northern blot analysis showed that at least one of the abp genes is transcribed in R. oligosporus NBRC 8631. A truncated form of abp1 encoding only the mature peptide was fused with the alpha-factor signal peptide and engineered for expression in Pichia pastoris SMD1168H. Culture broth of the recombinant Pichia displayed ABP activity against Bacillus subtilis NBRC 3335 after induction of heterologous gene expression. This result indicates that mature ABP formed the active structure without the aid of other factors from R. oligosporus, and was secreted.

V2R mutations and nephrogenic diabetes insipidus.

PubMed

Bichet, Daniel G

2009-01-01

Nephrogenic diabetes insipidus (NDI), which can be inherited or acquired, is characterized by an inability to concentrate urine despite normal or elevated plasma concentrations of the antidiuretic hormone, arginine vasopressin (AVP). Polyuria, with hyposthenuria, and polydipsia are the cardinal clinical manifestations of the disease. Nephrogenic failure to concentrate urine maximally may be due to a defect in vasopressin-induced water permeability of the distal tubules and collecting ducts, to insufficient buildup of the corticopapillary interstitial osmotic gradient, or to a combination of these two factors. Thus, the broadest definition of the term NDI embraces any antidiuretic hormone-resistant urinary-concentrating defect, including medullary disease with low interstitial osmolality, renal failure, and osmotic diuresis. About 90% of patients with congenital NDI are males with X-linked recessive NDI (OMIM 304800)(1) and have mutations in the AVP receptor 2 (AVPR2) gene that codes for the vasopressin V(2) receptor; the gene is located in chromosome region Xq28. In about 10% of the families studied, congenital NDI has an autosomal recessive or autosomal dominant mode of inheritance (OMIM 222000 and 125800)(1). Mutations have been identified in the aquaporin-2 gene (AQP2, OMIM 107777)(1), which is located in chromosome region 12q13 and codes for the vasopressin-sensitive water channel. NDI is clinically distinguishable from neurohypophyseal diabetes insipidus (OMIM 125700(1); also referred to as central or neurogenic diabetes insipidus) by a lack of response to exogenous AVP and by plasma levels of AVP that rise normally with increase in plasma osmolality. Hereditary neurohypophyseal diabetes insipidus is secondary to mutations in the gene encoding AVP (OMIM 192340)(1). Neurohypophyseal diabetes insipidus is also a component of autosomal recessive Wolfram syndrome 1 or DIDMOAD syndrome (diabetes insipidus, diabetes mellitus, optic atrophy, and deafness) (OMIM 222300)(1), an autosomal recessive disorder. Other inherited disorders with complex polyuro-polydipsic syndrome with loss of water, sodium, chloride, calcium, magnesium, and potassium include Bartter syndrome (OMIM 601678)(1) and cystinosis (OMIM 219800)(1), while long-term lithium administration is the main cause of acquired NDI. Here, we use the gene symbols approved by the HUGO Gene Nomenclature Committee (http://www.gene.ucl.ac.uk/nomenclature) and provide OMIM entry numbers [OMIM (Online Mendelian Inheritance in Man)(1); McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, MD) and National Center for Biotechnology Information, National Library of Medicine (Bethesda, MD), 2000; World Wide Web URL: http://www.ncbi.nlm.nih.gov/omim/]. Copyright © 2009 Elsevier Inc. All rights reserved.

Capturing novel mouse genes encoding chromosomal and other nuclear proteins.

PubMed

Tate, P; Lee, M; Tweedie, S; Skarnes, W C; Bickmore, W A

1998-09-01

The burgeoning wealth of gene sequences contrasts with our ignorance of gene function. One route to assigning function is by determining the sub-cellular location of proteins. We describe the identification of mouse genes encoding proteins that are confined to nuclear compartments by splicing endogeneous gene sequences to a promoterless betageo reporter, using a gene trap approach. Mouse ES (embryonic stem) cell lines were identified that express betageo fusions located within sub-nuclear compartments, including chromosomes, the nucleolus and foci containing splicing factors. The sequences of 11 trapped genes were ascertained, and characterisation of endogenous protein distribution in two cases confirmed the validity of the approach. Three novel proteins concentrated within distinct chromosomal domains were identified, one of which appears to be a serine/threonine kinase. The sequence of a gene whose product co-localises with splicesome components suggests that this protein may be an E3 ubiquitin-protein ligase. The majority of the other genes isolated represent novel genes. This approach is shown to be a powerful tool for identifying genes encoding novel proteins with specific sub-nuclear localisations and exposes our ignorance of the protein composition of the nucleus. Motifs in two of the isolated genes suggest new links between cellular regulatory mechanisms (ubiquitination and phosphorylation) and mRNA splicing and chromosome structure/function.

Many nonuniversal archaeal ribosomal proteins are found in conserved gene clusters

PubMed Central

WANG, JIACHEN; DASGUPTA, INDRANI; FOX, GEORGE E.

2009-01-01

The genomic associations of the archaeal ribosomal proteins, (r-proteins), were examined in detail. The archaeal versions of the universal r-protein genes are typically in clusters similar or identical and to those found in bacteria. Of the 35 nonuniversal archaeal r-protein genes examined, the gene encoding L18e was found to be associated with the conserved L13 cluster, whereas the genes for S4e, L32e and L19e were found in the archaeal version of the spc operon. Eleven nonuniversal protein genes were not associated with any common genomic context. Of the remaining 19 protein genes, 17 were convincingly assigned to one of 10 previously unrecognized gene clusters. Examination of the gene content of these clusters revealed multiple associations with genes involved in the initiation of protein synthesis, transcription or other cellular processes. The lack of such associations in the universal clusters suggests that initially the ribosome evolved largely independently of other processes. More recently it likely has evolved in concert with other cellular systems. It was also verified that a second copy of the gene encoding L7ae found in some bacteria is actually a homolog of the gene encoding L30e and should be annotated as such. PMID:19478915

Molecular Characterization of CTR-type Copper Transporters in an Oceanic Diatom, Thalassiosira oceanica 1005

NASA Astrophysics Data System (ADS)

Kong, L.; Price, N. M.

2016-02-01

Copper is an essential micronutrient for phytoplankton growth because of its role as a redox cofactor in electron transfer proteins in photosynthesis and respiration, and a potentially limiting resource in parts of the open sea. Thalassiosira oceanica 1005 can grow at inorganic copper concentrations varying from 10 fmol/L to 10 nmol/L by regulating copper uptake across plasma membrane. Four putative CTR-type copper transporter genes (ToCTR1, ToCTR2, ToCTR3.1 and ToCTR3.2) were identified by BLASTP search against the T. oceanica genome. Predicted gene models were revised by assembled mRNA sequencing transcripts and updated gene models contained all conserved features of characterized CTR-type copper transporters. ToCTR3.1 and ToCTR3.2 may arise from one another by gene duplication as they shared a sequence similarity of 97.6% with a peptide insertion of 5 amino acids at N-terminus of ToCTR3.1. The expression of ToCTR1, ToCTR2 and ToCTR3.1/3.2 was upregulated in low copper concentrations, but only ToCTR3.1/3.2 showed a significant increase (2.5 fold) in copper-starved cells. Both ToCTR3.1 and ToCTR3.2 restored growth of a yeast double mutant, Saccharomyces cerevisiae ctr1Δctr3Δ, in copper deficient medium. GFP-fused ToCTR expression showed that some ToCTR3.1 localized to the plasma membrane but a large portion was retained in the endoplasmic reticulum. Inefficient targeting of ToCTR3.1 to the yeast outer membrane may explain poorer growth compared to the Saccharomyces native ScCTR1 transformant. Thus, diatom CTR genes encoding CTR-type copper transporters show high-affinity copper uptake and their regulation may enable diatoms to survive in ocean environments containing a wide range of copper concentrations.

Identification of candidate transmission-blocking antigen genes in Theileria annulata and related vector-borne apicomplexan parasites.

PubMed

Lempereur, Laetitia; Larcombe, Stephen D; Durrani, Zeeshan; Karagenc, Tulin; Bilgic, Huseyin Bilgin; Bakirci, Serkan; Hacilarlioglu, Selin; Kinnaird, Jane; Thompson, Joanne; Weir, William; Shiels, Brian

2017-06-05

Vector-borne apicomplexan parasites are a major cause of mortality and morbidity to humans and livestock globally. The most important disease syndromes caused by these parasites are malaria, babesiosis and theileriosis. Strategies for control often target parasite stages in the mammalian host that cause disease, but this can result in reservoir infections that promote pathogen transmission and generate economic loss. Optimal control strategies should protect against clinical disease, block transmission and be applicable across related genera of parasites. We have used bioinformatics and transcriptomics to screen for transmission-blocking candidate antigens in the tick-borne apicomplexan parasite, Theileria annulata. A number of candidate antigen genes were identified which encoded amino acid domains that are conserved across vector-borne Apicomplexa (Babesia, Plasmodium and Theileria), including the Pfs48/45 6-cys domain and a novel cysteine-rich domain. Expression profiling confirmed that selected candidate genes are expressed by life cycle stages within infected ticks. Additionally, putative B cell epitopes were identified in the T. annulata gene sequences encoding the 6-cys and cysteine rich domains, in a gene encoding a putative papain-family cysteine peptidase, with similarity to the Plasmodium SERA family, and the gene encoding the T. annulata major merozoite/piroplasm surface antigen, Tams1. Candidate genes were identified that encode proteins with similarity to known transmission blocking candidates in related parasites, while one is a novel candidate conserved across vector-borne apicomplexans and has a potential role in the sexual phase of the life cycle. The results indicate that a 'One Health' approach could be utilised to develop a transmission-blocking strategy effective against vector-borne apicomplexan parasites of animals and humans.

Identification of β-propeller phytase-encoding genes in culturable Paenibacillus and Bacillus spp. from the rhizosphere of pasture plants on volcanic soils.

PubMed

Jorquera, Milko A; Crowley, David E; Marschner, Petra; Greiner, Ralf; Fernández, María Teresa; Romero, Daniela; Menezes-Blackburn, Daniel; De La Luz Mora, María

2011-01-01

Phytate is one of the most abundant sources of organic phosphorus (P) in soils, but must be mineralized by phytase-producing bacteria to release P for plant uptake. Microbial inoculants based on Bacillus spp. have been developed commercially, but few studies have evaluated the ecology of these bacteria in the rhizosphere or the types of enzymes that they produce. Here, we studied the diversity of aerobic endospore-forming bacteria (EFB) with the ability to mineralize phytate in the rhizosphere of pasture plants grown in volcanic soils of southern Chile. PCR methods were used to detect candidate phytase-encoding genes and to identify EFB bacteria that carry these genes. This study revealed that the phytate-degrading EFB populations of pasture plants included species of Paenibacillus and Bacillus, which carried genes encoding β-propeller phytase (BPP). Assays of enzymatic activity confirmed the ability of these rhizosphere isolates to degrade phytate. The phytase-encoding genes described here may prove valuable as molecular markers to evaluate the role of EFB in organic P mobilization in the rhizosphere. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Isolation and characterization of major histocompatibility complex class IIB genes from the nurse shark.

PubMed

Bartl, S; Weissman, I L

1994-01-04

The major histocompatibility complex (MHC) contains a set of linked genes which encode cell surface proteins involved in the binding of small peptide antigens for their subsequent recognition by T lymphocytes. MHC proteins share structural features and the presence and location of polymorphic residues which play a role in the binding of antigens. In order to compare the structure of these molecules and gain insights into their evolution, we have isolated two MHC class IIB genes from the nurse shark, Ginglymostoma cirratum. Two clones, most probably alleles, encode proteins which differ by 13 amino acids located in the putative antigen-binding cleft. The protein structure and the location of polymorphic residues are similar to their mammalian counterparts. Although these genes appear to encode a typical MHC protein, no T-cell-mediated responses have been demonstrated in cartilaginous fish. The nurse shark represents the most phylogenetically primitive organism in which both class IIA [Kasahara, M., Vazquez, M., Sato, K., McKinney, E.C. & Flajnik, M.F. (1992) Proc. Natl. Acad. Sci USA 89, 6688-6692] and class IIB genes, presumably encoding the alpha/beta heterodimer, have been isolated.

The Retrovirus pol Gene Encodes a Product Required for DNA Integration: Identification of a Retrovirus int Locus

NASA Astrophysics Data System (ADS)

Panganiban, Antonito T.; Temin, Howard M.

1984-12-01

We mutagenized cloned spleen necrosis virus DNA to identify a region of the retrovirus genome encoding a polypeptide required for integration of viral DNA. Five plasmids bearing different lesions in the 3' end of the pol gene were examined for the ability to integrate or replicate following transfection of chicken embryo fibroblasts. Transfection with one of these DNAs resulted in the generation of mutant virus incapable of integrating but able to replicate at low levels; this phenotype is identical to that of mutants bearing alterations in the cis-acting region, att. To determine whether the 3' end of the pol gene encodes a protein that interacts with att, we did a complementation experiment. Cells were first infected with an att- virus and then superinfected with the integration-deficient virus containing a lesion in the pol gene and a wild-type att site. The results showed that the att- virus provided a trans-acting function allowing integration of viral DNA derived from the mutant bearing a wild-type att site. Thus, the 3' end of the pol gene serves as an ``int'' locus and encodes a protein mediating integration of retrovirus DNA through interaction with att.

Cloning and characterization of largemouth bass ( Micropterus salmoides) myostatin encoding gene and its promoter

NASA Astrophysics Data System (ADS)

Li, Shengjie; Bai, Junjie; Wang, Lin

2008-08-01

Myostatin or GDF-8, a member of the transforming growth factor-β (TGF-β) superfamily, has been demonstrated to be a negative regulator of skeletal muscle mass in mammals. In the present study, we obtained a 5.64 kb sequence of myostatin encoding gene and its promoter from largemouth bass ( Micropterus salmoides). The myostatin encoding gene consisted of three exons (488 bp, 371 bp and 1779 bp, respectively) and two introns (390 bp and 855 bp, respectively). The intron-exon boundaries were conservative in comparison with those of mammalian myostatin encoding genes, whereas the size of introns was smaller than that of mammals. Sequence analysis of 1.569 kb of the largemouth bass myostatin gene promoter region revealed that it contained two TATA boxes, one CAAT box and nine putative E-boxes. Putative muscle growth response elements for myocyte enhancer factor 2 (MEF2), serum response factor (SRF), activator protein 1 (AP1), etc., and muscle-specific Mt binding site (MTBF) were also detected. Some of the transcription factor binding sites were conserved among five teleost species. This information will be useful for studying the transcriptional regulation of myostatin in fish.

A member of a new plant gene family encoding a meprin and TRAF homology (MATH) domain-containing protein is involved in restriction of long distance movement of plant viruses

PubMed Central

Cosson, Patrick; Sofer, Luc; Schurdi-Levraud, Valérie

2010-01-01

Restriction of long distance movement of several potyviruses in Arabidopsis thaliana is controlled by at least three dominant restricted TEV movement (RTM) genes, named RTM1, RTM2 and RTM3 and acts as a non-conventional resistance. RTM1 encodes a protein belonging to the jacalin family and RTM2 encodes a protein which has similarities to small heat shock proteins. The recent cloning of RTM3 which encodes a protein belonging to an unknown protein family of 29 members that has a meprin and TRAF homology (MATH) domain in its N-terminal region and a coiled-coil (CC) domain at its C-terminal end is an important breakthrough for a better understanding of this resistance process. Not only the third gene involved in this resistance has been identified and has allowed revealing a new gene family in plant but the discovery that the RTM3 protein interacts directly with RTM1 strongly suggests that the RTM proteins form a multimeric complex. However, these data also highlight striking similarities of the RTM resistance with the well known R-gene mediated resistance. PMID:20930558

Characterization of Urtica dioica agglutinin isolectins and the encoding gene family.

PubMed

Does, M P; Ng, D K; Dekker, H L; Peumans, W J; Houterman, P M; Van Damme, E J; Cornelissen, B J

1999-01-01

Urtica dioica agglutinin (UDA) has previously been found in roots and rhizomes of stinging nettles as a mixture of UDA-isolectins. Protein and cDNA sequencing have shown that mature UDA is composed of two hevein domains and is processed from a precursor protein. The precursor contains a signal peptide, two in-tandem hevein domains, a hinge region and a carboxyl-terminal chitinase domain. Genomic fragments encoding precursors for UDA-isolectins have been amplified by five independent polymerase chain reactions on genomic DNA from stinging nettle ecotype Weerselo. One amplified gene was completely sequenced. As compared to the published cDNA sequence, the genomic sequence contains, besides two basepair substitutions, two introns located at the same positions as in other plant chitinases. By partial sequence analysis of 40 amplified genes, 16 different genes were identified which encode seven putative UDA-isolectins. The deduced amino acid sequences share 78.9-98.9% identity. In extracts of roots and rhizomes of stinging nettle ecotype Weerselo six out of these seven isolectins were detected by mass spectrometry. One of them is an acidic form, which has not been identified before. Our results demonstrate that UDA is encoded by a large gene family.

L-rhamnose induction of Aspergillus nidulans α-L-rhamnosidase genes is glucose repressed via a CreA-independent mechanism acting at the level of inducer uptake.

PubMed

Tamayo-Ramos, Juan A; Flipphi, Michel; Pardo, Ester; Manzanares, Paloma; Orejas, Margarita

2012-02-21

Little is known about the structure and regulation of fungal α-L-rhamnosidase genes despite increasing interest in the biotechnological potential of the enzymes that they encode. Whilst the paradigmatic filamentous fungus Aspergillus nidulans growing on L-rhamnose produces an α-L-rhamnosidase suitable for oenological applications, at least eight genes encoding putative α-L-rhamnosidases have been found in its genome. In the current work we have identified the gene (rhaE) encoding the former activity, and characterization of its expression has revealed a novel regulatory mechanism. A shared pattern of expression has also been observed for a second α-L-rhamnosidase gene, (AN10277/rhaA). Amino acid sequence data for the oenological α-L-rhamnosidase were determined using MALDI-TOF mass spectrometry and correspond to the amino acid sequence deduced from AN7151 (rhaE). The cDNA of rhaE was expressed in Saccharomyces cerevisiae and yielded pNP-rhamnohydrolase activity. Phylogenetic analysis has revealed this eukaryotic α-L-rhamnosidase to be the first such enzyme found to be more closely related to bacterial rhamnosidases than other α-L-rhamnosidases of fungal origin. Northern analyses of diverse A. nidulans strains cultivated under different growth conditions indicate that rhaA and rhaE are induced by L-rhamnose and repressed by D-glucose as well as other carbon sources, some of which are considered to be non-repressive growth substrates. Interestingly, the transcriptional repression is independent of the wide domain carbon catabolite repressor CreA. Gene induction and glucose repression of these rha genes correlate with the uptake, or lack of it, of the inducing carbon source L-rhamnose, suggesting a prominent role for inducer exclusion in repression. The A. nidulans rhaE gene encodes an α-L-rhamnosidase phylogenetically distant to those described in filamentous fungi, and its expression is regulated by a novel CreA-independent mechanism. The identification of rhaE and the characterization of its regulation will facilitate the design of strategies to overproduce the encoded enzyme - or homologs from other fungi - for industrial applications. Moreover, A. nidulans α-L-rhamnosidase encoding genes could serve as prototypes for fungal genes coding for plant cell wall degrading enzymes regulated by a novel mechanism of CCR.

L-Rhamnose induction of Aspergillus nidulans α-L-rhamnosidase genes is glucose repressed via a CreA-independent mechanism acting at the level of inducer uptake

PubMed Central

2012-01-01

Background Little is known about the structure and regulation of fungal α-L-rhamnosidase genes despite increasing interest in the biotechnological potential of the enzymes that they encode. Whilst the paradigmatic filamentous fungus Aspergillus nidulans growing on L-rhamnose produces an α-L-rhamnosidase suitable for oenological applications, at least eight genes encoding putative α-L-rhamnosidases have been found in its genome. In the current work we have identified the gene (rhaE) encoding the former activity, and characterization of its expression has revealed a novel regulatory mechanism. A shared pattern of expression has also been observed for a second α-L-rhamnosidase gene, (AN10277/rhaA). Results Amino acid sequence data for the oenological α-L-rhamnosidase were determined using MALDI-TOF mass spectrometry and correspond to the amino acid sequence deduced from AN7151 (rhaE). The cDNA of rhaE was expressed in Saccharomyces cerevisiae and yielded pNP-rhamnohydrolase activity. Phylogenetic analysis has revealed this eukaryotic α-L-rhamnosidase to be the first such enzyme found to be more closely related to bacterial rhamnosidases than other α-L-rhamnosidases of fungal origin. Northern analyses of diverse A. nidulans strains cultivated under different growth conditions indicate that rhaA and rhaE are induced by L-rhamnose and repressed by D-glucose as well as other carbon sources, some of which are considered to be non-repressive growth substrates. Interestingly, the transcriptional repression is independent of the wide domain carbon catabolite repressor CreA. Gene induction and glucose repression of these rha genes correlate with the uptake, or lack of it, of the inducing carbon source L-rhamnose, suggesting a prominent role for inducer exclusion in repression. Conclusions The A. nidulans rhaE gene encodes an α-L-rhamnosidase phylogenetically distant to those described in filamentous fungi, and its expression is regulated by a novel CreA-independent mechanism. The identification of rhaE and the characterization of its regulation will facilitate the design of strategies to overproduce the encoded enzyme - or homologs from other fungi - for industrial applications. Moreover, A. nidulans α-L-rhamnosidase encoding genes could serve as prototypes for fungal genes coding for plant cell wall degrading enzymes regulated by a novel mechanism of CCR. PMID:22353731

Common variants of the vitamin D binding protein gene and adverse health outcomes.

PubMed

Malik, Suneil; Fu, Lei; Juras, David James; Karmali, Mohamed; Wong, Betty Y L; Gozdzik, Agnes; Cole, David E C

2013-01-01

The vitamin D binding protein (DBP) is the major plasma carrier for vitamin D and its metabolites, but it is also an actin scavenger, and is the precursor to the immunomodulatory protein, Gc-MAF. Two missense variants of the DBP gene - rs7041 encoding Asp432Glu and rs4588 encoding Thr436Lys - change the amino acid sequence and alter the protein function. They are common enough to generate population-wide constitutive differences in vitamin D status, based on assay of the serum metabolite, 25-hydroxyvitamin D (25OHD). Whether these variants also influence the role of vitamin D in an immunologic milieu is not known. However, the issue is relevant, given the immunomodulatory effects of DBP and the role of protracted innate immune-related inflammation in response to tissue injury or repeated infection. Indeed, DBP and vitamin D may jointly or independently contribute to a variety of adverse health outcomes unrelated to classical notions of their function in bone and mineral metabolism. This review summarizes the reports to date of associations between DBP variants, and various chronic and infectious diseases. The available information leads us to conclude that DBP variants are a significant and common genetic factor in some common disorders, and therefore, are worthy of closer attention. In view of the heightened interest in vitamin D as a public health target, well-designed studies that look simultaneously at vitamin D and its carrier in relation to genotypes and adverse health outcome should be encouraged.

Pht2;1 encodes a low-affinity phosphate transporter from Arabidopsis.

PubMed Central

Daram, P; Brunner, S; Rausch, C; Steiner, C; Amrhein, N; Bucher, M

1999-01-01

An Arabidopsis genomic sequence was recently shown to share similarity with bacterial and eukaryotic phosphate (Pi) transporters. We have cloned the corresponding cDNA, which we named Pht2;1, and subsequently performed gene expression studies and functional analysis of the protein product. The cDNA encodes a 61-kD protein with a putative topology of 12 transmembrane (TM) domains interrupted by a large hydrophilic loop between TM8 and TM9. Two boxes of eight and nine amino acids, located in the N- and C-terminal domains, respectively, are highly conserved among species across all kingdoms (eubacteria, archea, fungi, plants, and animals). The Pht2;1 gene is predominantly expressed in green tissue, the amount of transcript staying constant in leaves irrespective of the Pi status of the shoot; in roots, however, there is a marginal increase in mRNA amounts in response to Pi deprivation. Although the protein is highly similar to eukaryotic sodium-dependent Pi transporters, functional analysis of the Pht2;1 protein in mutant yeast cells indicates that it is a proton/Pi symporter dependent on the electrochemical gradient across the plasma membrane. Its fairly high apparent K(m) for Pi (0.4 mM) and high mRNA content in the shoot, especially in leaves, suggest a role for shoot organs in Pi loading. Pht2;1 thus differs from members of the recently described plant Pi transporter family in primary structure, affinity for Pi, and presumed function. PMID:10559441

Anthocyanins and their gut metabolites reduce the adhesion of monocyte to TNFα-activated endothelial cells at physiologically relevant concentrations.

PubMed

Krga, Irena; Monfoulet, Laurent-Emmanuel; Konic-Ristic, Aleksandra; Mercier, Sylvie; Glibetic, Maria; Morand, Christine; Milenkovic, Dragan

2016-06-01

An increasing number of evidence suggests a protective role of dietary anthocyanins against cardiovascular diseases. Anthocyanins' extensive metabolism indicates that their metabolites could be responsible for the protective effects associated with consumption of anthocyanin-rich foods. The aim of this work was to investigate the effect of plasma anthocyanins and their metabolites on the adhesion of monocytes to TNFα-activated endothelial cells and on the expression of genes encoding cell adhesion molecules. Human umbilical vein endothelial cells (HUVECs) were exposed to circulating anthocyanins: cyanidin-3-arabinoside, cyanidin-3-galactoside, cyanidin-3-glucoside, delphinidin-3-glucoside, peonidin-3-glucoside, anthocyanin degradation product: 4-hydroxybenzaldehyde, or to their gut metabolites: protocatechuic, vanillic, ferulic and hippuric acid, at physiologically-relevant concentrations (0.1-2 μM) and time of exposure. Both anthocyanins and gut metabolites decreased the adhesion of monocytes to HUVECs, with a magnitude ranging from 18.1% to 47%. The mixture of anthocyanins and that of gut metabolites also reduced monocyte adhesion. However, no significant effect on the expression of genes encoding E-selectin, ICAM1 and VCAM1 was observed, suggesting that other molecular targets are involved in the observed effect. In conclusion, this study showed the potency of anthocyanins and their gut metabolites to modulate the adhesion of monocytes to endothelial cells, the initial step in atherosclerosis development, under physiologically-relevant conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Typing of Panton-Valentine Leukocidin-Encoding Phages and lukSF-PV Gene Sequence Variation in Staphylococcus aureus from China.

PubMed

Zhao, Huanqiang; Hu, Fupin; Jin, Shu; Xu, Xiaogang; Zou, Yuhan; Ding, Baixing; He, Chunyan; Gong, Fang; Liu, Qingzhong

2016-01-01

Panton-Valentine leukocidin (PVL, encoded by lukSF-PV genes), a bi-component and pore-forming toxin, is carried by different staphylococcal bacteriophages. The prevalence of PVL in Staphylococcus aureus has been reported around the globe. However, the data on PVL-encoding phage types, lukSF-PV gene variation and chromosomal phage insertion sites for PVL-positive S. aureus are limited, especially in China. In order to obtain a more complete understanding of the molecular epidemiology of PVL-positive S. aureus, an integrated and modified PCR-based scheme was applied to detect the PVL-encoding phage types. Phage insertion locus and the lukSF-PV variant were determined by PCR and sequencing. Meanwhile, the genetic background was characterized by staphylococcal cassette chromosome mec (SCCmec) typing, staphylococcal protein A (spa) gene polymorphisms typing, pulsed-field gel electrophoresis (PFGE) typing, accessory gene regulator (agr) locus typing and multilocus sequence typing (MLST). Seventy eight (78/1175, 6.6%) isolates possessed the lukSF-PV genes and 59.0% (46/78) of PVL-positive strains belonged to CC59 lineage. Eight known different PVL-encoding phage types were detected, and Φ7247PVL/ΦST5967PVL (n = 13) and ΦPVL (n = 12) were the most prevalent among them. While 25 (25/78, 32.1%) isolates, belonging to ST30, and ST59 clones, were unable to be typed by the modified PCR-based scheme. Single nucleotide polymorphisms (SNPs) were identified at five locations in the lukSF-PV genes, two of which were non-synonymous. Maximum-likelihood tree analysis of attachment sites sequences detected six SNP profiles for attR and eight for attL, respectively. In conclusion, the PVL-positive S. aureus mainly harbored Φ7247PVL/ΦST5967PVL and ΦPVL in the regions studied. lukSF-PV gene sequences, PVL-encoding phages, and phage insertion locus generally varied with lineages. Moreover, PVL-positive clones that have emerged worldwide likely carry distinct phages.

The complete chloroplast genome sequence of Dianthus superbus var. longicalycinus.

PubMed

Gurusamy, Raman; Lee, Do-Hyung; Park, SeonJoo

2016-05-01

The complete chloroplast genome (cpDNA) sequence of Dianthus superbus var. longicalycinus is an economically important traditional Chinese medicine was reported and characterized. The cpDNA of Dianthus superbus var. longicalycinus is 149,539 bp, with 36.3% GC content. A pair of inverted repeats (IRs) of 24,803 bp is separated by a large single-copy region (LSC, 82,805 bp) and a small single-copy region (SSC, 17,128 bp). It encodes 85 protein-coding genes, 36 tRNA genes and 8 rRNA genes. Of 129 individual genes, 13 genes encoded one intron and three genes have two introns.

Repressed expression of a gene for a basic helix-loop-helix protein causes a white flower phenotype in carnation

PubMed Central

Totsuka, Akane; Okamoto, Emi; Miyahara, Taira; Kouno, Takanobu; Cano, Emilio A.; Sasaki, Nobuhiro; Watanabe, Aiko; Tasaki, Keisuke; Nishihara, Masahiro; Ozeki, Yoshihiro

2018-01-01

In a previous study, two genes responsible for white flower phenotypes in carnation were identified. These genes encoded enzymes involved in anthocyanin synthesis, namely, flavanone 3-hydroxylase (F3H) and dihydroflavonol 4-reductase (DFR), and showed reduced expression in the white flower phenotypes. Here, we identify another candidate gene for white phenotype in carnation flowers using an RNA-seq analysis followed by RT-PCR. This candidate gene encodes a transcriptional regulatory factor of the basic helix-loop-helix (bHLH) type. In the cultivar examined here, both F3H and DFR genes produced active enzyme proteins; however, expression of DFR and of genes for enzymes involved in the downstream anthocyanin synthetic pathway from DFR was repressed in the absence of bHLH expression. Occasionally, flowers of the white flowered cultivar used here have red speckles and stripes on the white petals. We found that expression of bHLH occurred in these red petal segments and induced expression of DFR and the following downstream enzymes. Our results indicate that a member of the bHLH superfamily is another gene involved in anthocyanin synthesis in addition to structural genes encoding enzymes. PMID:29681756

Distribution of Suicin Gene Clusters in Streptococcus suis Serotype 2 Belonging to Sequence Types 25 and 28.

PubMed

Athey, Taryn B T; Vaillancourt, Katy; Frenette, Michel; Fittipaldi, Nahuel; Gottschalk, Marcelo; Grenier, Daniel

2016-01-01

Recently, we reported the purification and characterization of three distinct lantibiotics (named suicin 90-1330, suicin 3908, and suicin 65) produced by Streptococcus suis . In this study, we investigated the distribution of the three suicin lantibiotic gene clusters among serotype 2 S. suis strains belonging to sequence type (ST) 25 and ST28, the two dominant STs identified in North America. The genomes of 102 strains were interrogated for the presence of suicin gene clusters encoding suicins 90-1330, 3908, and 65. The gene cluster encoding suicin 65 was the most prevalent and mainly found among ST25 strains. In contrast, none of the genes related to suicin 90-1330 production were identified in 51 ST25 strains nor in 35/51 ST28 strains. However, the complete suicin 90-1330 gene cluster was found in ten ST28 strains, although some genes in the cluster were truncated in three of these isolates. The vast majority (101/102) of S. suis strains did not possess any of the genes encoding suicin 3908. In conclusion, this study indicates heterogeneous distribution of suicin genes in S. suis .

Screening of the Enterocin-Encoding Genes and Antimicrobial Activity in Enterococcus Species.

PubMed

Ogaki, Mayara Baptistucci; Rocha, Katia Real; Terra, MÁrcia Regina; Furlaneto, MÁrcia Cristina; Maia, Luciana Furlaneto

2016-06-28

In the current study, a total of 135 enterococci strains from different sources were screened for the presence of the enterocin-encoding genes entA, entP, entB, entL50A, and entL50B. The enterocin genes were present at different frequencies, with entA occurring the most frequently, followed by entP and entB; entL50A and L50B were not detected. The occurrence of single enterocin genes was higher than the occurrence of multiple enterocin gene combinations. The 80 isolates that harbor at least one enterocin-encoding gene (denoted "Gene(+) strains") were screened for antimicrobial activity. A total of 82.5% of the Gene(+) strains inhibited at least one of the indicator strains, and the isolates harboring multiple enterocin-encoding genes inhibited a larger number of indicator strains than isolates harboring a single gene. The indicator strains that exhibited growth inhibition included Listeria innocua strain CLIP 12612 (ATCC BAA-680), Listeria monocytogenes strain CDC 4555, Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 25923, S. aureus ATCC 29213, S. aureus ATCC 6538, Salmonella enteritidis ATCC 13076, Salmonella typhimurium strain UK-1 (ATCC 68169), and Escherichia coli BAC 49LT ETEC. Inhibition due to either bacteriophage lysis or cytolysin activity was excluded. The growth inhibition of antilisterial Gene+ strains was further tested under different culture conditions. Among the culture media formulations, the MRS agar medium supplemented with 2% (w/v) yeast extract was the best solidified medium for enterocin production. Our findings extend the current knowledge of enterocin-producing enterococci, which may have potential applications as biopreservatives in the food industry due to their capability of controlling food spoilage pathogens.

Update on the molecular biology of dyslipidemias.

PubMed

Ramasamy, I

2016-02-15

Dyslipidemia is a commonly encountered clinical condition and is an important determinant of cardiovascular disease. Although secondary factors play a role in clinical expression, dyslipidemias have a strong genetic component. Familial hypercholesterolemia is usually due to loss-of-function mutations in LDLR, the gene coding for low density lipoprotein receptor and genes encoding for proteins that interact with the receptor: APOB, PCSK9 and LDLRAP1. Monogenic hypertriglyceridemia is the result of mutations in genes that regulate the metabolism of triglyceride rich lipoproteins (eg LPL, APOC2, APOA5, LMF1, GPIHBP1). Conversely familial hypobetalipoproteinemia is caused by inactivation of the PCSK9 gene which increases the number of LDL receptors and decreases plasma cholesterol. Mutations in the genes APOB, and ANGPTL3 and ANGPTL4 (that encode angiopoietin-like proteins which inhibit lipoprotein lipase activity) can further cause low levels of apoB containing lipoproteins. Abetalipoproteinemia and chylomicron retention disease are due to mutations in the microsomal transfer protein and Sar1b-GTPase genes, which affect the secretion of apoB containing lipoproteins. Dysbetalipoproteinemia stems from dysfunctional apoE and is characterized by the accumulation of remnants of chylomicrons and very low density lipoproteins. ApoE deficiency can cause a similar phenotype or rarely mutations in apoE can be associated with lipoprotein glomerulopathy. Low HDL can result from mutations in a number of genes regulating HDL production or catabolism; apoAI, lecithin: cholesterol acyltransferase and the ATP-binding cassette transporter ABCA1. Patients with cholesteryl ester transfer protein deficiency have markedly increased HDL cholesterol. Both common and rare genetic variants contribute to susceptibility to dyslipidemias. In contrast to rare familial syndromes, in most patients, dyslipidemias have a complex genetic etiology consisting of multiple genetic variants as established by genome wide association studies. Secondary factors, obesity, metabolic syndrome, diabetes, renal disease, estrogen and antipsychotics can increase the likelihood of clinical presentation of an individual with predisposed genetic susceptibility to hyperlipoproteinemia. The genetic profiles studied are far from complete and there is room for further characterization of genes influencing lipid levels. Genetic assessment can help identify patients at risk for developing dyslipidemias and for treatment decisions based on 'risk allele' profiles. This review will present the current information on the genetics and pathophysiology of disorders that cause dyslipidemias. Copyright © 2015 Elsevier B.V. All rights reserved.

Transcriptomic analysis of Saccharomyces cerevisiae upon honokiol treatment.

PubMed

Zhu, Xiaolong; Zou, Shenshen; Li, Youbin; Liang, Yongheng

2017-09-01

Honokiol (HNK), one of the main medicinal components in Magnolia officinalis, possesses antimicrobial activity against a variety of pathogenic bacteria and fungi. However, little is known of the molecular mechanisms underpinning the antimicrobial activity. To explore the molecular mechanism of its antifungal activity, we determined the effects of HNK on the mRNA expression profile of Saccharomyces cerevisiae using a DNA microarray approach. HNK markedly induced the expression of genes related to iron uptake and homeostasis. Conversely, genes associated with respiratory electron transport were downregulated, mirroring the effects of iron starvation. Meanwhile, HNK-induced growth deficiency was partly rescued by iron supplementation and HNK reacted with iron, producing iron complexes that depleted iron. These results suggest that HNK treatment induced iron starvation. Additionally, HNK treatment resulted in the upregulation of genes involved in protein synthesis and drug resistance networks. Furthermore, the deletion of PDR5, a gene encoding the plasma membrane ATP binding cassette (ABC) transporter, conferred sensitivity to HNK. Overexpression of PDR5 enhanced resistance of WT and pdr5Δ strains to HNK. Taken together, these findings suggest that HNK, which can be excluded by overexpression of Pdr5, functions in multiple cellular processes in S. cerevisiae, particularly in inducing iron starvation to inhibit cell growth. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

The MADS transcription factor XAL2/AGL14 modulates auxin transport during Arabidopsis root development by regulating PIN expression

PubMed Central

Garay-Arroyo, Adriana; Ortiz-Moreno, Enrique; de la Paz Sánchez, María; Murphy, Angus S; García-Ponce, Berenice; Marsch-Martínez, Nayelli; de Folter, Stefan; Corvera-Poiré, Adriana; Jaimes-Miranda, Fabiola; Pacheco-Escobedo, Mario A; Dubrovsky, Joseph G; Pelaz, Soraya; Álvarez-Buylla, Elena R

2013-01-01

Elucidating molecular links between cell-fate regulatory networks and dynamic patterning modules is a key for understanding development. Auxin is important for plant patterning, particularly in roots, where it establishes positional information for cell-fate decisions. PIN genes encode plasma membrane proteins that serve as auxin efflux transporters; mutations in members of this gene family exhibit smaller roots with altered root meristems and stem-cell patterning. Direct regulators of PIN transcription have remained elusive. Here, we establish that a MADS-box gene (XAANTAL2, XAL2/AGL14) controls auxin transport via PIN transcriptional regulation during Arabidopsis root development; mutations in this gene exhibit altered stem-cell patterning, root meristem size, and root growth. XAL2 is necessary for normal shootward and rootward auxin transport, as well as for maintaining normal auxin distribution within the root. Furthermore, this MADS-domain transcription factor upregulates PIN1 and PIN4 by direct binding to regulatory regions and it is required for PIN4-dependent auxin response. In turn, XAL2 expression is regulated by auxin levels thus establishing a positive feedback loop between auxin levels and PIN regulation that is likely to be important for robust root patterning. PMID:24121311

Copy number variation in metabolic phenotypes.

PubMed

Lanktree, M; Hegele, R A

2008-01-01

Despite successes in identifying genetic contributors to common metabolic phenotypes, only part of the heritable component of these traits has thus far been explained. Copy number variation (CNV) is likely to be responsible for some of the unexplained variation. As observed with single nucleotide changes, it is probable that both rare and common CNVs will contribute to susceptibility to metabolic disease. For instance, CNVs in the LDLR gene underlie a substantial portion of disease in patients with heterozygous familial hypercholesterolemia. As well, a common CNV in LPA encoding apolipoprotein(a) is the primary determinant of plasma lipoprotein(a) concentrations, a risk factor for atherosclerosis. Recent efforts to map CNVs in control populations have defined their size, frequency and distribution. Many of the identified CNVs overlap genes with important functions in metabolic pathways. The overlap of CNVs that were found in control datasets with functional candidate genes or genes with previous evidence of association with metabolic syndrome presents an important subset for future CNV association studies. Finally, we describe an approach to search for CNVs in a rare high-penetrance metabolic disorder, namely lipodystrophy. As methods to identify CNVs increase in precision and accuracy, the prospect of identifying their role in both rare Mendelian and common complex metabolic phenotypes will become a reality. Copyright 2009 S. Karger AG, Basel.

Plasma fatty acid levels and gene expression related to lipid metabolism in peripheral blood mononuclear cells: a cross-sectional study in healthy subjects.

PubMed

Larsen, Sunniva V; Holven, Kirsten B; Ottestad, Inger; Dagsland, Kine N; Myhrstad, Mari C W; Ulven, Stine M

2018-01-01

Solid evidence indicates that intake of marine n-3 fatty acids lowers serum triglycerides and that replacing saturated fatty acids (SFA) with polyunsaturated fatty acids (PUFA) reduces plasma total cholesterol and LDL cholesterol. The molecular mechanisms underlying these health beneficial effects are however not completely elucidated. The aim of this study was therefore to investigate the expression of genes related to lipid metabolism in peripheral blood mononuclear cells (PBMC) depending on the plasma levels of n-6 and n-3 fatty acids and the SFA to PUFA ratio. Fifty-four healthy subjects were grouped into tertiles ( n  = 18) based on plasma levels of n-6 and n-3 fatty acids and the SFA to PUFA ratio. The PBMC gene expression levels among subjects in the highest versus the lowest tertiles were compared. In total, 285 genes related to cholesterol and triglyceride metabolism were selected for this explorative study. Among the 285 selected genes, 161 were defined as expressed in the PBMCs. The plasma SFA to PUFA ratio was associated with the highest number of significantly different expressed genes (25 gene transcripts), followed by plasma n-6 fatty acid level (15 gene transcripts) and plasma n-3 fatty acid level (8 gene transcripts). In particular, genes involved in cholesterol homeostasis were significantly different expressed among subjects with high compared to low plasma SFA to PUFA ratio. Genes involved in lipid metabolism were differentially expressed in PBMCs depending on the plasma fatty acid levels. This finding may increase our understanding of how fatty acids influence lipid metabolism at a molecular level in humans.

Assessing the resistance and bioremediation ability of selected bacterial and protozoan species to heavy metals in metal-rich industrial wastewater

PubMed Central

2013-01-01

Background Heavy-metals exert considerable stress on the environment worldwide. This study assessed the resistance to and bioremediation of heavy-metals by selected protozoan and bacterial species in highly polluted industrial-wastewater. Specific variables (i.e. chemical oxygen demand, pH, dissolved oxygen) and the growth/die-off-rates of test organisms were measured using standard methods. Heavy-metal removals were determined in biomass and supernatant by the Inductively Couple Plasma Optical Emission Spectrometer. A parallel experiment was performed with dead microbial cells to assess the biosorption ability of test isolates. Results The results revealed that the industrial-wastewater samples were highly polluted with heavy-metal concentrations exceeding by far the maximum limits (in mg/l) of 0.05-Co, 0.2-Ni, 0.1-Mn, 0.1-V, 0.01-Pb, 0.01-Cu, 0.1-Zn and 0.005-Cd, prescribed by the UN-FAO. Industrial-wastewater had no major effects on Pseudomonas putida, Bacillus licheniformis and Peranema sp. (growth rates up to 1.81, 1.45 and 1.43 d-1, respectively) compared to other test isolates. This was also revealed with significant COD increases (p < 0.05) in culture media inoculated with living bacterial isolates (over 100%) compared to protozoan isolates (up to 24% increase). Living Pseudomonas putida demonstrated the highest removal rates of heavy metals (Co-71%, Ni-51%, Mn-45%, V-83%, Pb-96%, Ti-100% and Cu-49%) followed by Bacillus licheniformis (Al-23% and Zn-53%) and Peranema sp. (Cd-42%). None of the dead cells were able to remove more than 25% of the heavy metals. Bacterial isolates contained the genes copC, chrB, cnrA3 and nccA encoding the resistance to Cu, Cr, Co-Ni and Cd-Ni-Co, respectively. Protozoan isolates contained only the genes encoding Cu and Cr resistance (copC and chrB genes). Peranema sp. was the only protozoan isolate which had an additional resistant gene cnrA3 encoding Co-Ni resistance. Conclusion Significant differences (p < 0.05) observed between dead and living microbial cells for metal-removal and the presence of certain metal-resistant genes indicated that the selected microbial isolates used both passive (biosorptive) and active (bioaccumulation) mechanisms to remove heavy metals from industrial wastewater. This study advocates the use of Peranema sp. as a potential candidate for the bioremediation of heavy-metals in wastewater treatment, in addition to Pseudomonas putida and Bacillus licheniformis. PMID:23387904

Assessing the resistance and bioremediation ability of selected bacterial and protozoan species to heavy metals in metal-rich industrial wastewater.

PubMed

Kamika, Ilunga; Momba, Maggy N B

2013-02-06

Heavy-metals exert considerable stress on the environment worldwide. This study assessed the resistance to and bioremediation of heavy-metals by selected protozoan and bacterial species in highly polluted industrial-wastewater. Specific variables (i.e. chemical oxygen demand, pH, dissolved oxygen) and the growth/die-off-rates of test organisms were measured using standard methods. Heavy-metal removals were determined in biomass and supernatant by the Inductively Couple Plasma Optical Emission Spectrometer. A parallel experiment was performed with dead microbial cells to assess the biosorption ability of test isolates. The results revealed that the industrial-wastewater samples were highly polluted with heavy-metal concentrations exceeding by far the maximum limits (in mg/l) of 0.05-Co, 0.2-Ni, 0.1-Mn, 0.1-V, 0.01-Pb, 0.01-Cu, 0.1-Zn and 0.005-Cd, prescribed by the UN-FAO. Industrial-wastewater had no major effects on Pseudomonas putida, Bacillus licheniformis and Peranema sp. (growth rates up to 1.81, 1.45 and 1.43 d-1, respectively) compared to other test isolates. This was also revealed with significant COD increases (p < 0.05) in culture media inoculated with living bacterial isolates (over 100%) compared to protozoan isolates (up to 24% increase). Living Pseudomonas putida demonstrated the highest removal rates of heavy metals (Co-71%, Ni-51%, Mn-45%, V-83%, Pb-96%, Ti-100% and Cu-49%) followed by Bacillus licheniformis (Al-23% and Zn-53%) and Peranema sp. (Cd-42%). None of the dead cells were able to remove more than 25% of the heavy metals. Bacterial isolates contained the genes copC, chrB, cnrA3 and nccA encoding the resistance to Cu, Cr, Co-Ni and Cd-Ni-Co, respectively. Protozoan isolates contained only the genes encoding Cu and Cr resistance (copC and chrB genes). Peranema sp. was the only protozoan isolate which had an additional resistant gene cnrA3 encoding Co-Ni resistance. Significant differences (p < 0.05) observed between dead and living microbial cells for metal-removal and the presence of certain metal-resistant genes indicated that the selected microbial isolates used both passive (biosorptive) and active (bioaccumulation) mechanisms to remove heavy metals from industrial wastewater. This study advocates the use of Peranema sp. as a potential candidate for the bioremediation of heavy-metals in wastewater treatment, in addition to Pseudomonas putida and Bacillus licheniformis.

Direct Activation of Amidohydrolase Domain-Containing 1 Gene by Thyroid Hormone Implicates a Role in the Formation of Adult Intestinal Stem Cells During Xenopus Metamorphosis

PubMed Central

Okada, Morihiro; Miller, Thomas C.; Fu, Liezhen

2015-01-01

The T3-dependent anuran metamorphosis resembles postembryonic development in mammals, the period around birth when plasma T3 levels peak. In particular, the remodeling of the intestine during metamorphosis mimics neonatal intestinal maturation in mammals when the adult intestinal epithelial self-renewing system is established. We have been using intestinal metamorphosis to investigate how the organ-specific adult stem cells are formed during vertebrate development. Early studies in Xenopus laevis have shown that this process involves complete degeneration of the larval epithelium and de novo formation of adult stem cells. A tissue-specific microarray analysis of intestinal gene expression during Xenopus laevis metamorphosis has identified a number of candidate stem cell genes. Here we have carried out detailed analyses of one such gene, amidohydrolase domain containing 1 (AMDHD1) gene, which encodes an enzyme in the histidine catabolic pathway. We show that AMDHD1 is exclusively expressed in the proliferating adult epithelial stem cells during metamorphosis with little expression in other intestinal tissues. We further provide evidence that T3 activates AMDHD1 gene expression directly at the transcription level through T3 receptor binding to the AMDHD1 gene in the intestine. In addition, we have reported earlier that histidine ammonia-lyase gene, another gene in histidine catabolic pathway, is similarly regulated by T3 in the intestine. These results together suggest that histidine catabolism plays a critical role in the formation and/or proliferation of adult intestinal stem cells during metamorphosis. PMID:26086244

Direct Activation of Amidohydrolase Domain-Containing 1 Gene by Thyroid Hormone Implicates a Role in the Formation of Adult Intestinal Stem Cells During Xenopus Metamorphosis.

PubMed

Okada, Morihiro; Miller, Thomas C; Fu, Liezhen; Shi, Yun-Bo

2015-09-01

The T3-dependent anuran metamorphosis resembles postembryonic development in mammals, the period around birth when plasma T3 levels peak. In particular, the remodeling of the intestine during metamorphosis mimics neonatal intestinal maturation in mammals when the adult intestinal epithelial self-renewing system is established. We have been using intestinal metamorphosis to investigate how the organ-specific adult stem cells are formed during vertebrate development. Early studies in Xenopus laevis have shown that this process involves complete degeneration of the larval epithelium and de novo formation of adult stem cells. A tissue-specific microarray analysis of intestinal gene expression during Xenopus laevis metamorphosis has identified a number of candidate stem cell genes. Here we have carried out detailed analyses of one such gene, amidohydrolase domain containing 1 (AMDHD1) gene, which encodes an enzyme in the histidine catabolic pathway. We show that AMDHD1 is exclusively expressed in the proliferating adult epithelial stem cells during metamorphosis with little expression in other intestinal tissues. We further provide evidence that T3 activates AMDHD1 gene expression directly at the transcription level through T3 receptor binding to the AMDHD1 gene in the intestine. In addition, we have reported earlier that histidine ammonia-lyase gene, another gene in histidine catabolic pathway, is similarly regulated by T3 in the intestine. These results together suggest that histidine catabolism plays a critical role in the formation and/or proliferation of adult intestinal stem cells during metamorphosis.

Designing universal primers for the isolation of DNA sequences encoding Proanthocyanidins biosynthetic enzymes in Crataegus aronia

PubMed Central

2012-01-01

Background Hawthorn is the common name of all plant species in the genus Crataegus, which belongs to the Rosaceae family. Crataegus are considered useful medicinal plants because of their high content of proanthocyanidins (PAs) and other related compounds. To improve PAs production in Crataegus tissues, the sequences of genes encoding PAs biosynthetic enzymes are required. Findings Different bioinformatics tools, including BLAST, multiple sequence alignment and alignment PCR analysis were used to design primers suitable for the amplification of DNA fragments from 10 candidate genes encoding enzymes involved in PAs biosynthesis in C. aronia. DNA sequencing results proved the utility of the designed primers. The primers were used successfully to amplify DNA fragments of different PAs biosynthesis genes in different Rosaceae plants. Conclusion To the best of our knowledge, this is the first use of the alignment PCR approach to isolate DNA sequences encoding PAs biosynthetic enzymes in Rosaceae plants. PMID:22883984

Designing universal primers for the isolation of DNA sequences encoding Proanthocyanidins biosynthetic enzymes in Crataegus aronia.

PubMed

Zuiter, Afnan Saeid; Sawwan, Jammal; Al Abdallat, Ayed

2012-08-10

Hawthorn is the common name of all plant species in the genus Crataegus, which belongs to the Rosaceae family. Crataegus are considered useful medicinal plants because of their high content of proanthocyanidins (PAs) and other related compounds. To improve PAs production in Crataegus tissues, the sequences of genes encoding PAs biosynthetic enzymes are required. Different bioinformatics tools, including BLAST, multiple sequence alignment and alignment PCR analysis were used to design primers suitable for the amplification of DNA fragments from 10 candidate genes encoding enzymes involved in PAs biosynthesis in C. aronia. DNA sequencing results proved the utility of the designed primers. The primers were used successfully to amplify DNA fragments of different PAs biosynthesis genes in different Rosaceae plants. To the best of our knowledge, this is the first use of the alignment PCR approach to isolate DNA sequences encoding PAs biosynthetic enzymes in Rosaceae plants.

Genome-Wide Characterization and Expression Analysis of Major Intrinsic Proteins during Abiotic and Biotic Stresses in Sweet Orange (Citrus sinensis L. Osb.)

PubMed Central

de Paula Santos Martins, Cristina; Pedrosa, Andresa Muniz; Du, Dongliang; Gonçalves, Luana Pereira; Yu, Qibin; Gmitter, Frederick G.; Costa, Marcio Gilberto Cardoso

2015-01-01

The family of aquaporins (AQPs), or major intrinsic proteins (MIPs), includes integral membrane proteins that function as transmembrane channels for water and other small molecules of physiological significance. MIPs are classified into five subfamilies in higher plants, including plasma membrane (PIPs), tonoplast (TIPs), NOD26-like (NIPs), small basic (SIPs) and unclassified X (XIPs) intrinsic proteins. This study reports a genome-wide survey of MIP encoding genes in sweet orange (Citrus sinensis L. Osb.), the most widely cultivated Citrus spp. A total of 34 different genes encoding C. sinensis MIPs (CsMIPs) were identified and assigned into five subfamilies (CsPIPs, CsTIPs, CsNIPs, CsSIPs and CsXIPs) based on sequence analysis and also on their phylogenetic relationships with clearly classified MIPs of Arabidopsis thaliana. Analysis of key amino acid residues allowed the assessment of the substrate specificity of each CsMIP. Gene structure analysis revealed that the CsMIPs possess an exon-intron organization that is highly conserved within each subfamily. CsMIP loci were precisely mapped on every sweet orange chromosome, indicating a wide distribution of the gene family in the sweet orange genome. Investigation of their expression patterns in different tissues and upon drought and salt stress treatments, as well as with ‘Candidatus Liberibacter asiaticus’ infection, revealed a tissue-specific and coordinated regulation of the different CsMIP isoforms, consistent with the organization of the stress-responsive cis-acting regulatory elements observed in their promoter regions. A special role in regulating the flow of water and nutrients is proposed for CsTIPs and CsXIPs during drought stress, and for most CsMIPs during salt stress and the development of HLB disease. These results provide a valuable reference for further exploration of the CsMIPs functions and applications to the genetic improvement of both abiotic and biotic stress tolerance in citrus. PMID:26397813

Genome-Wide Characterization and Expression Analysis of Major Intrinsic Proteins during Abiotic and Biotic Stresses in Sweet Orange (Citrus sinensis L. Osb.).

PubMed

Martins, Cristina de Paula Santos; Pedrosa, Andresa Muniz; Du, Dongliang; Gonçalves, Luana Pereira; Yu, Qibin; Gmitter, Frederick G; Costa, Marcio Gilberto Cardoso

2015-01-01

The family of aquaporins (AQPs), or major intrinsic proteins (MIPs), includes integral membrane proteins that function as transmembrane channels for water and other small molecules of physiological significance. MIPs are classified into five subfamilies in higher plants, including plasma membrane (PIPs), tonoplast (TIPs), NOD26-like (NIPs), small basic (SIPs) and unclassified X (XIPs) intrinsic proteins. This study reports a genome-wide survey of MIP encoding genes in sweet orange (Citrus sinensis L. Osb.), the most widely cultivated Citrus spp. A total of 34 different genes encoding C. sinensis MIPs (CsMIPs) were identified and assigned into five subfamilies (CsPIPs, CsTIPs, CsNIPs, CsSIPs and CsXIPs) based on sequence analysis and also on their phylogenetic relationships with clearly classified MIPs of Arabidopsis thaliana. Analysis of key amino acid residues allowed the assessment of the substrate specificity of each CsMIP. Gene structure analysis revealed that the CsMIPs possess an exon-intron organization that is highly conserved within each subfamily. CsMIP loci were precisely mapped on every sweet orange chromosome, indicating a wide distribution of the gene family in the sweet orange genome. Investigation of their expression patterns in different tissues and upon drought and salt stress treatments, as well as with 'Candidatus Liberibacter asiaticus' infection, revealed a tissue-specific and coordinated regulation of the different CsMIP isoforms, consistent with the organization of the stress-responsive cis-acting regulatory elements observed in their promoter regions. A special role in regulating the flow of water and nutrients is proposed for CsTIPs and CsXIPs during drought stress, and for most CsMIPs during salt stress and the development of HLB disease. These results provide a valuable reference for further exploration of the CsMIPs functions and applications to the genetic improvement of both abiotic and biotic stress tolerance in citrus.

Recognitional specificity and evolution in the tomato-Cladosporium fulvum pathosystem.

PubMed

Wulff, B B H; Chakrabarti, A; Jones, D A

2009-10-01

The interactions between plants and many biotrophic or hemibiotrophic pathogens are controlled by receptor proteins in the host and effector proteins delivered by the pathogen. Pathogen effectors facilitate pathogen growth through the suppression of host defenses and the manipulation of host metabolism, but recognition of a pathogen-effector protein by a host receptor enables the host to activate a suite of defense mechanisms that limit pathogen growth. In the tomato (Lycopersicon esculentum syn. Solanum lycopersicum)-Cladosporium fulvum (leaf mold fungus syn. Passalora fulva) pathosystem, the host receptors are plasma membrane-anchored, leucine-rich repeat, receptor-like proteins encoded by an array of Cf genes conferring resistance to C. fulvum. The pathogen effectors are mostly small, secreted, cysteine-rich, but otherwise largely dissimilar, extracellular proteins encoded by an array of avirulence (Avr) genes, so called because of their ability to trigger resistance and limit pathogen growth when the corresponding Cf gene is present in tomato. A number of Cf and Avr genes have been isolated, and details of the complex molecular interplay between tomato Cf proteins and C. fulvum effector proteins are beginning to emerge. Each effector appears to have a different role; probably most bind or modify different host proteins, but at least one has a passive role masking the pathogen. It is, therefore, not surprising that each effector is probably detected in a distinct and specific manner, some by direct binding, others as complexes with host proteins, and others via their modification of host proteins. The two papers accompanying this review contribute further to our understanding of the molecular specificity underlying effector perception by Cf proteins. This review, therefore, focuses on our current understanding of recognitional specificity in the tomato-C. fulvum pathosystem and highlights some of the critical questions that remain to be addressed. It also addresses the evolutionary causes and consequences of this specificity.

Differential effects of saturated and unsaturated fatty acid diets on cardiomyocyte apoptosis, adipose distribution, and serum leptin.

PubMed

Okere, Isidore C; Chandler, Margaret P; McElfresh, Tracy A; Rennison, Julie H; Sharov, Victor; Sabbah, Hani N; Tserng, Kou-Yi; Hoit, Brian D; Ernsberger, Paul; Young, Martin E; Stanley, William C

2006-07-01

Fatty acids are the primary fuel for the heart and are ligands for peroxisome proliferator-activated receptors (PPARs), which regulate the expression of genes encoding proteins involved in fatty acid metabolism. Saturated fatty acids, particularly palmitate, can be converted to the proapoptotic lipid intermediate ceramide. This study assessed cardiac function, expression of PPAR-regulated genes, and cardiomyocyte apoptosis in rats after 8 wk on either a low-fat diet [normal chow control (NC); 10% fat calories] or high-fat diets composed mainly of either saturated (Sat) or unsaturated fatty acids (Unsat) (60% fat calories) (n = 10/group). The Sat group had lower plasma insulin and leptin concentrations compared with the NC or Unsat groups. Cardiac function and mass and body mass were not different. Cardiac triglyceride content was increased in the Sat and Unsat groups compared with NC (P < 0.05); however, ceramide content was higher in the Sat group compared with the Unsat group (2.9 +/- 0.2 vs. 1.4 +/- 0.2 nmol/g; P < 0.05), whereas the NC group was intermediate (2.3 +/- 0.3 nmol/g). The number of apoptotic myocytes, assessed by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling staining, was higher in the Sat group compared with the Unsat group (0.28 +/- 0.05 vs. 0.17 +/- 0.04 apoptotic cells/1,000 nuclei; P < 0.04) and was positively correlated to ceramide content (P < 0.02). Both high-fat diets increased the myocardial mRNA expression of the PPAR-regulated genes encoding uncoupling protein-3 and pyruvate dehydrogenase kinase-4, but only the Sat diet upregulated medium-chain acyl-CoA dehydrogenase. In conclusion, dietary fatty acid composition affects cardiac ceramide accumulation, cardiomyocyte apoptosis, and expression of PPAR-regulated genes independent of cardiac mass or function.

CCN2 plays a key role in extracellular matrix gene expression in severe hypertrophic cardiomyopathy and heart failure.

PubMed

Tsoutsman, Tatiana; Wang, Xiaoyu; Garchow, Kendra; Riser, Bruce; Twigg, Stephen; Semsarian, Christopher

2013-09-01

Hypertrophic cardiomyopathy (HCM) is the most common inherited primary myocardial disorder. HCM is characterized by interstitial fibrosis and excessive accumulation of extracellular matrix (ECM) proteins. Fibrosis in HCM has been associated with impaired cardiac function and heart failure, and has been considered a key substrate for ventricular arrhythmias and sudden death. The molecular triggers underpinning ECM production are not well established. We have previously developed a double-mutant mouse model of HCM that recapitulates the phenotype seen in humans with multiple mutations, including earlier onset of the disease, progression to a dilated phenotype, severe heart failure and premature mortality. The present study investigated the expression of ECM-encoding genes in severe HCM and heart failure. Significant upregulation of structural Fn1, regulatory Mmp14, Timp1, Serpin3A, SerpinE1, SerpineE2, Tgfβ1, and Tgfβ2; and matricellular Ccn2, Postn, Spp1, Thbs1, Thbs4, and Tnc was evident from the early, pre-phenotype stage. Non-myocytes expressed ECM genes at higher levels than cardiomyocytes in normal and diseased hearts. Synchronous increase of secreted CCN2 and TIMP1 plasma levels and decrease of MMP3 levels were observed in end-stage disease. CCN2 protein expression was increased from early disease in double-mutant hearts and played an important role in ECM responses. It was a powerful modulator of ECM regulatory (Timp1 and SerpinE1) and matricellular protein-encoding (Spp1, Thbs1, Thbs4 and Tnc) gene expression in cardiomyocytes when added exogenously in vitro. Modulation of CCN2 (CTGF, connective tissue growth factor) and associated early ECM changes may represent a new therapeutic target in the treatment and prevention of heart failure in HCM. Copyright © 2013 Elsevier Ltd. All rights reserved.

The Association of High Prevalence of Trophozoites in Peripheral Blood with Lower Antibody Response to P. falciparum Infected Erythrocytes among Asymptomatic Children in Sudan.

PubMed

Mohamed, Sara N; Hassan, Dina A; El Hussein, Abdelrahim M; Osman, Ihssan M; Ibrahim, Muntasir E; Mohamed, Hiba S; Nour, Bakri Y; Abdulhadi, Nasreldin H

2016-01-01

Background. The most prominent variant surface antigens (VSAs) of Plasmodium falciparum are the var gene-encoded Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family, which serves as a parasite-sequestering ligand to endothelial cells. In this study we have examined the antibody reactivity of autologous plasma from symptomatic and asymptomatic malaria infected children against the infected erythrocytes' surface antigens using flow cytometry. Methods. Ethidium-bromide-labelled erythrocytic mature forms of P. falciparum parasites obtained from symptomatic and asymptomatic children were sequentially incubated with autologous plasma and fluorescein isothiocyanate-conjugated (FITC) antihuman IgG. Plasma antibody reactivity was detected by flow cytometry. Results. Asymptomatic children had more prevalence of trophozoites in peripheral blood (66%) compared to symptomatic children (16%), p = 0.002. The mean percentage of infected RBCs reacting with autologous sera was 89.78 among symptomatic children compared to 79.62 among asymptomatic children (p = 0.09). Moreover, the mean fluorescence intensity (MFI) in the asymptomatic was significantly higher compared to symptomatic children (p value = 0.040). Conclusion. Variant surface antigens on Plasmodium falciparum infected RBCs from symptomatic malaria children tend to be better recognized by IgG antibodies. This may suggest a role of some IgG antibodies in severity of malaria.

The role of brassinosteroids in the regulation of the plasma membrane H+-ATPase and NADPH oxidase under cadmium stress.

PubMed

Jakubowska, Dagmara; Janicka, Małgorzata

2017-11-01

The present research aim was to define the role of brassinosteroids (BRs) in plant adaptation to cadmium stress. We observed a stimulating effect of exogenous BR on the activity of two plasma membrane enzymes which play a key role in plants adaptation to cadmium stress, H + -ATPase (EC 3.6.3.14) and NADPH oxidase (EC 1.6.3.1). Using anti-phosphothreonine antibody we showed that modification of PM H + -ATPase activity under BR action could result from phosphorylation of the enzyme protein. Also the relative expression of genes encoding both PM H + -ATPase and NADPH oxidase was affected by BR. To confirm the role of BR in the cadmium stimulating effect on activity of both studied plasma membrane enzymes, an assay in the presence of a BR biosynthesis inhibitor (propiconazole) was performed. Moreover, as a tool in our work we used commercially available plant mutants unable to BR biosynthesis or with dysfunctional BR signaling pathway, to further confirm participation of BR in plant adaptation to heavy metal stress. Presented results demonstrate some elements of the brassinosteroid-induced pathway activated under cadmium stress, wherein H + -ATPase and NADPH oxidase are key factors. Copyright © 2017 Elsevier B.V. All rights reserved.

The euryhaline yeast Debaryomyces hansenii has two catalase genes encoding enzymes with differential activity profile.

PubMed

Segal-Kischinevzky, Claudia; Rodarte-Murguía, Beatriz; Valdés-López, Victor; Mendoza-Hernández, Guillermo; González, Alicia; Alba-Lois, Luisa

2011-03-01

Debaryomyces hansenii is a spoilage yeast able to grow in a variety of ecological niches, from seawater to dairy products. Results presented in this article show that (i) D. hansenii has an inherent resistance to H2O2 which could be attributed to the fact that this yeast has a basal catalase activity which is several-fold higher than that observed in Saccharomyces cerevisiae under the same culture conditions, (ii) D. hansenii has two genes (DhCTA1 and DhCTT1) encoding two catalase isozymes with a differential enzymatic activity profile which is not strictly correlated with a differential expression profile of the encoding genes.

Draft genome sequence of Xylaria sp., the causal agent of taproot decline of soybean in the southern United States.

PubMed

Sharma, Sandeep; Zaccaron, Alex Z; Ridenour, John B; Allen, Tom W; Conner, Kassie; Doyle, Vinson P; Price, Trey; Sikora, Edward; Singh, Raghuwinder; Spurlock, Terry; Tomaso-Peterson, Maria; Wilkerson, Tessie; Bluhm, Burton H

2018-04-01

The draft genome of Xylaria sp. isolate MSU_SB201401, causal agent of taproot decline of soybean in the southern U.S., is presented here. The genome assembly was 56.7 Mb in size with an L50 of 246. A total of 10,880 putative protein-encoding genes were predicted, including 647 genes encoding carbohydrate-active enzymes and 1053 genes encoding secreted proteins. This is the first draft genome of a plant-pathogenic Xylaria sp. associated with soybean. The draft genome of Xylaria sp. isolate MSU_SB201401 will provide an important resource for future experiments to determine the molecular basis of pathogenesis.

Cloning and characterization of a mouse gene with homology to the human von Hippel-Lindau disease tumor suppressor gene: implications for the potential organization of the human von Hippel-Lindau disease gene.

PubMed

Gao, J; Naglich, J G; Laidlaw, J; Whaley, J M; Seizinger, B R; Kley, N

1995-02-15

The human von Hippel-Lindau disease (VHL) gene has recently been identified and, based on the nucleotide sequence of a partial cDNA clone, has been predicted to encode a novel protein with as yet unknown functions [F. Latif et al., Science (Washington DC), 260: 1317-1320, 1993]. The length of the encoded protein and the characteristics of the cellular expressed protein are as yet unclear. Here we report the cloning and characterization of a mouse gene (mVHLh1) that is widely expressed in different mouse tissues and shares high homology with the human VHL gene. It predicts a protein 181 residues long (and/or 162 amino acids, considering a potential alternative start codon), which across a core region of approximately 140 residues displays a high degree of sequence identity (98%) to the predicted human VHL protein. High stringency DNA and RNA hybridization experiments and protein expression analyses indicate that this gene is the most highly VHL-related mouse gene, suggesting that it represents the mouse VHL gene homologue rather than a related gene sharing a conserved functional domain. These findings provide new insights into the potential organization of the VHL gene and nature of its encoded protein.

Plasmid-Encoded Tetracycline Efflux Pump Protein Alters Bacterial Stress Responses and Ecological Fitness of Acinetobacter oleivorans

PubMed Central

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment. PMID:25229538

Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster.

PubMed

Robertson, Hugh M; Warr, Coral G; Carlson, John R

2003-11-25

The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods.

Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster

PubMed Central

Robertson, Hugh M.; Warr, Coral G.; Carlson, John R.

2003-01-01

The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods. PMID:14608037

Plasmid-encoded tetracycline efflux pump protein alters bacterial stress responses and ecological fitness of Acinetobacter oleivorans.

PubMed

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment.

Gene Transfection Method Using Atmospheric Pressure Dielectric-Barrier Discharge Plasmas

NASA Astrophysics Data System (ADS)

Sasaki, Shota; Kanzaki, Makoto; Kaneko, Toshiro

2013-09-01

Gene transfection which is the process of deliberately introducing nucleic acids into cells is expected to play an important role in medical treatment because the process is necessary for gene therapy and creation of induced pluripotent stem (iPS) cells. However, the conventional transfection methods have some problems, so we focus attention on promising transfection methods by atmospheric pressure dielectric-barrier discharge (AP-DBD) plasmas. AP-DBD He plasmas are irradiated to the living cell covered with genes. Preliminarily, we use fluorescent dye YOYO-1 instead of the genes and use LIVE/DEAD Stain for cell viability test, and we analyze the transfection efficiency and cell viability under the various conditions. It is clarified that the transfection efficiency is strongly dependence on the plasma irradiation time and cell viability rates is high rates (>90%) regardless of long plasma irradiation time. These results suggest that ROS (Reactive Oxygen Species) and electric field generated by the plasma affect the gene transfection. In addition to this (the plasma irradiation time) dependency, we now investigate the effect of the plasma irradiation under the various conditions.

The Streptococcus pyogenes serotype M49 Nra-Ralp3 transcriptional regulatory network and its control of virulence factor expression from the novel eno ralp3 epf sagA pathogenicity region.

PubMed

Kreikemeyer, Bernd; Nakata, Masanobu; Köller, Thomas; Hildisch, Hendrikje; Kourakos, Vassilios; Standar, Kerstin; Kawabata, Shigetada; Glocker, Michael O; Podbielski, Andreas

2007-12-01

Many Streptococcus pyogenes (group A streptococcus [GAS]) virulence factor- and transcriptional regulator-encoding genes cluster together in discrete genomic regions. Nra is a central regulator of the FCT region. Previous studies exclusively described Nra as a transcriptional repressor of adhesin and toxin genes. Here transcriptome and proteome analysis of a serotype M49 GAS strain and an isogenic Nra mutant of this strain revealed the complete Nra regulon profile. Nra is active in all growth phases tested, with the largest regulon in the transition phase. Almost exclusively, virulence factor-encoding genes are repressed by Nra; these genes include the GAS pilus operon, the capsule synthesis operon, the cytolysin-mediated translocation system genes, all Mga region core virulence genes, and genes encoding other regulators, like the Ihk/Irr system, Rgg, and two additional RofA-like protein family regulators. Surprisingly, our experiments revealed that Nra additionally acts as a positive regulator, mostly for genes encoding proteins and enzymes with metabolic functions. Epidemiological investigations revealed strong genetic linkage of one particular Nra-repressed regulator, Ralp3 (SPy0735), with a gene encoding Epf (extracellular protein factor from Streptococcus suis). In a serotype-specific fashion, this ralp3 epf gene block is integrated, most likely via transposition, into the eno sagA virulence gene block, which is present in all GAS serotypes. In GAS serotypes M1, M4, M12, M28, and M49 this novel discrete genetic region is therefore designated the eno ralp3 epf sagA (ERES) pathogenicity region. Functional experiments showed that Epf is a novel GAS plasminogen-binding protein and revealed that Ralp3 activity counteracts Nra and MsmR regulatory activity. In addition to the Mga and FCT regions, the ERES region is the third discrete chromosomal pathogenicity region. All of these regions are transcriptionally linked, adding another level of complexity to the known GAS growth phase-dependent regulatory network.

The Streptococcus pyogenes Serotype M49 Nra-Ralp3 Transcriptional Regulatory Network and Its Control of Virulence Factor Expression from the Novel eno ralp3 epf sagA Pathogenicity Region▿ †

PubMed Central

Kreikemeyer, Bernd; Nakata, Masanobu; Köller, Thomas; Hildisch, Hendrikje; Kourakos, Vassilios; Standar, Kerstin; Kawabata, Shigetada; Glocker, Michael O.; Podbielski, Andreas

2007-01-01

Many Streptococcus pyogenes (group A streptococcus [GAS]) virulence factor- and transcriptional regulator-encoding genes cluster together in discrete genomic regions. Nra is a central regulator of the FCT region. Previous studies exclusively described Nra as a transcriptional repressor of adhesin and toxin genes. Here transcriptome and proteome analysis of a serotype M49 GAS strain and an isogenic Nra mutant of this strain revealed the complete Nra regulon profile. Nra is active in all growth phases tested, with the largest regulon in the transition phase. Almost exclusively, virulence factor-encoding genes are repressed by Nra; these genes include the GAS pilus operon, the capsule synthesis operon, the cytolysin-mediated translocation system genes, all Mga region core virulence genes, and genes encoding other regulators, like the Ihk/Irr system, Rgg, and two additional RofA-like protein family regulators. Surprisingly, our experiments revealed that Nra additionally acts as a positive regulator, mostly for genes encoding proteins and enzymes with metabolic functions. Epidemiological investigations revealed strong genetic linkage of one particular Nra-repressed regulator, Ralp3 (SPy0735), with a gene encoding Epf (extracellular protein factor from Streptococcus suis). In a serotype-specific fashion, this ralp3 epf gene block is integrated, most likely via transposition, into the eno sagA virulence gene block, which is present in all GAS serotypes. In GAS serotypes M1, M4, M12, M28, and M49 this novel discrete genetic region is therefore designated the eno ralp3 epf sagA (ERES) pathogenicity region. Functional experiments showed that Epf is a novel GAS plasminogen-binding protein and revealed that Ralp3 activity counteracts Nra and MsmR regulatory activity. In addition to the Mga and FCT regions, the ERES region is the third discrete chromosomal pathogenicity region. All of these regions are transcriptionally linked, adding another level of complexity to the known GAS growth phase-dependent regulatory network. PMID:17893125

Molecular cloning and characterization of two novel genes from hexaploid wheat that encode double PR-1 domains coupled with a receptor-like protein kinase

USDA-ARS?s Scientific Manuscript database

Hexaploid wheat (Triticum aestivum L.) contains at least 23 TaPr-1 genes encoding the group 1 pathogenesis-related (PR-1) proteins as identified in our previous work. Here we report the cloning and characterization of TaPr-1-rk1 and TaPr-1-rk2, two novel genes closely related to the wheat PR-1 famil...

Draft Genome Sequence of Enterohemorrhagic Escherichia coli O157:H7 Strain MC2 Isolated from Cattle in France

PubMed Central

Auffret, Pauline; Segura, Audrey; Klopp, Christophe; Bouchez, Olivier; Kérourédan, Monique; Bibbal, Delphine; Brugère, Hubert; Forano, Evelyne

2017-01-01

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) with serotype O157:H7 is a major foodborne pathogen. Here, we report the draft genome sequence of EHEC O157:H7 strain MC2 isolated from cattle in France. The assembly contains 5,400,376 bp that encoded 5,914 predicted genes (5,805 protein-encoding genes and 109 RNA genes). PMID:28983004

Evolution of Prdm Genes in Animals: Insights from Comparative Genomics

PubMed Central

Vervoort, Michel; Meulemeester, David; Béhague, Julien; Kerner, Pierre

2016-01-01

Prdm genes encode transcription factors with a subtype of SET domain known as the PRDF1-RIZ (PR) homology domain and a variable number of zinc finger motifs. These genes are involved in a wide variety of functions during animal development. As most Prdm genes have been studied in vertebrates, especially in mice, little is known about the evolution of this gene family. We searched for Prdm genes in the fully sequenced genomes of 93 different species representative of all the main metazoan lineages. A total of 976 Prdm genes were identified in these species. The number of Prdm genes per species ranges from 2 to 19. To better understand how the Prdm gene family has evolved in metazoans, we performed phylogenetic analyses using this large set of identified Prdm genes. These analyses allowed us to define 14 different subfamilies of Prdm genes and to establish, through ancestral state reconstruction, that 11 of them are ancestral to bilaterian animals. Three additional subfamilies were acquired during early vertebrate evolution (Prdm5, Prdm11, and Prdm17). Several gene duplication and gene loss events were identified and mapped onto the metazoan phylogenetic tree. By studying a large number of nonmetazoan genomes, we confirmed that Prdm genes likely constitute a metazoan-specific gene family. Our data also suggest that Prdm genes originated before the diversification of animals through the association of a single ancestral SET domain encoding gene with one or several zinc finger encoding genes. PMID:26560352

Exploiting algal NADPH oxidase for biophotovoltaic energy

DOE PAGES

Anderson, Alexander; Laohavisit, Anuphon; Blaby, Ian K.; ...

2015-01-29

Photosynthetic microbes exhibit light-dependent electron export across the cell membrane, which can generate electricity in biological photovoltaic (BPV) devices. How electrons are exported remains to be determined; the identification of mechanisms would help selection or generation of photosynthetic microbes capable of enhanced electrical output. We show that plasma membrane NADPH oxidase activity is a significant component of light-dependent generation of electricity by the unicellular green alga Chlamydomonas reinhardtii. NADPH oxidases export electrons across the plasma membrane to form superoxide anion from oxygen. The C. reinhardtii mutant lacking the NADPH oxidase encoded by RBO1 is impaired in both extracellular superoxide anionmore » production and current generation in a BPV device. Complementation with the wild-type gene restores both capacities, demonstrating the role of the enzyme in electron export. Monitoring light-dependent extracellular superoxide production with a colorimetric assay is shown to be an effective way of screening for electrogenic potential of candidate algal strains. Furthermore, the results show that algal NADPH oxidases are important for superoxide anion production and open avenues for optimizing the biological component of these devices.« less

Investigation of the Lipid Binding Properties of the Marburg Virus Matrix Protein VP40.

PubMed

Wijesinghe, Kaveesha J; Stahelin, Robert V

2015-12-30

Marburg virus (MARV), which belongs to the virus family Filoviridae, causes hemorrhagic fever in humans and nonhuman primates that is often fatal. MARV is a lipid-enveloped virus that during the replication process extracts its lipid coat from the plasma membrane of the host cell it infects. MARV carries seven genes, one of which encodes its matrix protein VP40 (mVP40), which regulates the assembly and budding of the virions. Currently, little information is available on mVP40 lipid binding properties. Here, we have investigated the in vitro and cellular mechanisms by which mVP40 associates with lipid membranes. mVP40 associates with anionic membranes in a nonspecific manner that is dependent upon the anionic charge density of the membrane. These results are consistent with recent structural determination of mVP40, which elucidated an mVP40 dimer with a flat and extensive cationic lipid binding interface. Marburg virus (MARV) is a lipid-enveloped filamentous virus from the family Filoviridae. MARV was discovered in 1967, and yet little is known about how its seven genes are used to assemble and form a new viral particle in the host cell it infects. The MARV matrix protein VP40 (mVP40) underlies the inner leaflet of the virus and regulates budding from the host cell plasma membrane. In vitro and cellular assays in this study investigated the mechanism by which mVP40 associates with lipids. The results demonstrate that mVP40 interactions with lipid vesicles or the inner leaflet of the plasma membrane are electrostatic but nonspecific in nature and are dependent on the anionic charge density of the membrane surface. Small molecules that can disrupt lipid trafficking or reduce the anionic charge of the plasma membrane interface may be useful in inhibiting assembly and budding of MARV. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

IL1RN Coding Variant Is Associated with Lower Risk of Acute Respiratory Distress Syndrome and Increased Plasma IL-1 Receptor Antagonist

PubMed Central

Feng, Rui; Li, Mingyao; Zhao, Yang; Sheu, Chau-Chyun; Tejera, Paula; Gallop, Robert; Bellamy, Scarlett; Rushefski, Melanie; Lanken, Paul N.; Aplenc, Richard; O’Keefe, Grant E.; Wurfel, Mark M.; Christiani, David C.; Christie, Jason D.

2013-01-01

Rationale: Acute respiratory distress syndrome (ARDS) behaves as a complex genetic trait, yet knowledge of genetic susceptibility factors remains incomplete. Objectives: To identify genetic risk variants for ARDS using large scale genotyping. Methods: A multistage genetic association study was conducted of three critically ill populations phenotyped for ARDS. Stage I, a trauma cohort study (n = 224), was genotyped with a 50K gene-centric single-nucleotide polymorphism (SNP) array. We tested SNPs associated with ARDS at P < 5 × 10−4 for replication in stage II, a trauma case–control population (n = 778). SNPs replicating their association in stage II (P < 0.005) were tested in a stage III nested case–control population of mixed subjects in the intensive care unit (n = 2,063). Logistic regression was used to adjust for potential clinical confounders. We performed ELISA to test for an association between ARDS-associated genotype and plasma protein levels. Measurements and Main Results: A total of 12 SNPs met the stage I threshold for an association with ARDS. rs315952 in the IL1RN gene encoding IL-1 receptor antagonist (IL1RA) replicated its association with reduced ARDS risk in stages II (P < 0.004) and III (P < 0.02), and was robust to clinical adjustment (combined odds ratio = 0.81; P = 4.2 × 10−5). Plasma IL1RA level was associated with rs315952C in a subset of critically ill subjects. The effect of rs315952 was independent from the tandem repeat variant in IL1RN. Conclusions: The IL1RN SNP rs315952C is associated with decreased risk of ARDS in three populations with heterogeneous ARDS risk factors, and with increased plasma IL1RA response. IL1RA may attenuate ARDS risk. PMID:23449693

Identification, expression and function of apolipoprotein E in annual fish Nothobranchius guentheri: implication for an aging marker.

PubMed

Wang, Xia; Shang, Xiaomei; Luan, Jing; Zhang, Shicui

2014-06-01

Apolipoprotein E (apoE) is a lipid-associated protein present in both plasma and in central nervous system. Variation in apoE gene has been reported to be associated with longevity in humans as well as with aged diseases such as atherosclerosis, Alzheimer's disease, and Parkinson's disease. However, information regarding the function and structure-activity relationship of apoE in lower vertebrates is rather limited. In this study we show that the apoE gene from the annual fish Nothobranchius guentheri, NapoE, encodes a protein of 262 amino acids, which shares common structural features characteristic of mammalian apoE. We also show that like human apoE, recombinant NapoE is able to inhibit LDL oxidation, and it is the N-terminal domain of NapoE with lysine or arginine residues that plays a key role in inhibition of LDL oxidation. NapoE is predominantly expressed in the liver of N. guentheri, consistent with that in mammalian species. More importantly, we demonstrate an age-dependent down-regulation of NapoE gene, rendering it a suitable biomarker of aging. This lays a foundation for further study of the role of apoE in the aging process of fish.

STEAP: A prostate-specific cell-surface antigen highly expressed in human prostate tumors

PubMed Central

Hubert, Rene S.; Vivanco, Igor; Chen, Emily; Rastegar, Shiva; Leong, Kahan; Mitchell, Steve C.; Madraswala, Rashida; Zhou, Yanhong; Kuo, James; Raitano, Arthur B.; Jakobovits, Aya; Saffran, Douglas C.; Afar, Daniel E. H.

1999-01-01

In search of novel genes expressed in metastatic prostate cancer, we subtracted cDNA isolated from benign prostatic hypertrophic tissue from cDNA isolated from a prostate cancer xenograft model that mimics advanced disease. One novel gene that is highly expressed in advanced prostate cancer encodes a 339-amino acid protein with six potential membrane-spanning regions flanked by hydrophilic amino- and carboxyl-terminal domains. This structure suggests a potential function as a channel or transporter protein. This gene, named STEAP for six-transmembrane epithelial antigen of the prostate, is expressed predominantly in human prostate tissue and is up-regulated in multiple cancer cell lines, including prostate, bladder, colon, ovarian, and Ewing sarcoma. Immunohistochemical analysis of clinical specimens demonstrates significant STEAP expression at the cell–cell junctions of the secretory epithelium of prostate and prostate cancer cells. Little to no staining was detected at the plasma membranes of normal, nonprostate human tissues, except for bladder tissue, which expressed low levels of STEAP at the cell membrane. Protein analysis located STEAP at the cell surface of prostate-cancer cell lines. Our results support STEAP as a cell-surface tumor-antigen target for prostate cancer therapy and diagnostic imaging. PMID:10588738

GTP cyclohydrolase I gene polymorphisms are associated with endothelial dysfunction and oxidative stress in patients with type 2 diabetes mellitus.

PubMed

Wolkow, Pawel P; Kosiniak-Kamysz, Wladyslaw; Osmenda, Grzegorz; Wilk, Grzegorz; Bujak-Gizycka, Beata; Ignacak, Adam; Kanitkar, Mihir; Walus-Miarka, Malgorzata; Harrison, David G; Korbut, Ryszard; Malecki, Maciej T; Guzik, Tomasz J

2014-01-01

The genetic background of atherosclerosis in type 2 diabetes mellitus (T2DM) is complex and poorly understood. Studying genetic components of intermediate phenotypes, such as endothelial dysfunction and oxidative stress, may aid in identifying novel genetic components for atherosclerosis in diabetic patients. Five polymorphisms forming two haplotype blocks within the GTP cyclohydrolase 1 gene, encoding a rate limiting enzyme in tetrahydrobiopterin synthesis, were studied in the context of flow and nitroglycerin mediated dilation (FMD and NMD), intima-media thickness (IMT), and plasma concentrations of von Willebrand factor (vWF) and malondialdehyde (MDA). Rs841 was associated with FMD (p = 0.01), while polymorphisms Rs10483639, Rs841, Rs3783641 (which form a single haplotype) were associated with both MDA (p = 0.012, p = 0.0015 and p = 0.003, respectively) and vWF concentrations (p = 0.016, p = 0.03 and p = 0.045, respectively). In addition, polymorphism Rs8007267 was also associated with MDA (p = 0.006). Haplotype analysis confirmed the association of both haplotypes with studied variables. Genetic variation of the GCH1 gene is associated with endothelial dysfunction and oxidative stress in T2DM patients.

The Aspergillus niger faeB gene encodes a second feruloyl esterase involved in pectin and xylan degradation and is specifically induced in the presence of aromatic compounds.

PubMed

de Vries, Ronald P; vanKuyk, Patricia A; Kester, Harry C M; Visser, Jaap

2002-04-15

The faeB gene encoding a second feruloyl esterase from Aspergillus niger has been cloned and characterized. It consists of an open reading frame of 1644 bp containing one intron. The gene encodes a protein of 521 amino acids that has sequence similarity to that of an Aspergillus oryzae tannase. However, the encoded enzyme, feruloyl esterase B (FAEB), does not have tannase activity. Comparison of the physical characteristics and substrate specificity of FAEB with those of a cinnamoyl esterase from A. niger [Kroon, Faulds and Williamson (1996) Biotechnol. Appl. Biochem. 23, 255-262] suggests that they are in fact the same enzyme. The expression of faeB is specifically induced in the presence of certain aromatic compounds, but not in the presence of other constituents present in plant-cell-wall polysaccharides such as arabinoxylan or pectin. The expression profile of faeB in the presence of aromatic compounds was compared with the expression of A. niger faeA, encoding feruloyl esterase A (FAEA), and A. niger bphA, the gene encoding a benzoate-p-hydroxylase. All three genes have different subsets of aromatic compounds that induce their expression, indicating the presence of different transcription activating systems in A. niger that respond to aromatic compounds. Comparison of the activity of FAEA and FAEB on sugar-beet pectin and wheat arabinoxylan demonstrated that they are both involved in the degradation of both polysaccharides, but have opposite preferences for these substrates. FAEA is more active than FAEB towards wheat arabinoxylan, whereas FAEB is more active than FAEA towards sugar-beet pectin.

The Aspergillus niger faeB gene encodes a second feruloyl esterase involved in pectin and xylan degradation and is specifically induced in the presence of aromatic compounds.

PubMed Central

de Vries, Ronald P; vanKuyk, Patricia A; Kester, Harry C M; Visser, Jaap

2002-01-01

The faeB gene encoding a second feruloyl esterase from Aspergillus niger has been cloned and characterized. It consists of an open reading frame of 1644 bp containing one intron. The gene encodes a protein of 521 amino acids that has sequence similarity to that of an Aspergillus oryzae tannase. However, the encoded enzyme, feruloyl esterase B (FAEB), does not have tannase activity. Comparison of the physical characteristics and substrate specificity of FAEB with those of a cinnamoyl esterase from A. niger [Kroon, Faulds and Williamson (1996) Biotechnol. Appl. Biochem. 23, 255-262] suggests that they are in fact the same enzyme. The expression of faeB is specifically induced in the presence of certain aromatic compounds, but not in the presence of other constituents present in plant-cell-wall polysaccharides such as arabinoxylan or pectin. The expression profile of faeB in the presence of aromatic compounds was compared with the expression of A. niger faeA, encoding feruloyl esterase A (FAEA), and A. niger bphA, the gene encoding a benzoate-p-hydroxylase. All three genes have different subsets of aromatic compounds that induce their expression, indicating the presence of different transcription activating systems in A. niger that respond to aromatic compounds. Comparison of the activity of FAEA and FAEB on sugar-beet pectin and wheat arabinoxylan demonstrated that they are both involved in the degradation of both polysaccharides, but have opposite preferences for these substrates. FAEA is more active than FAEB towards wheat arabinoxylan, whereas FAEB is more active than FAEA towards sugar-beet pectin. PMID:11931668

Heterologous expression of a Tpo1 homolog from Arabidopsis thaliana confers resistance to the herbicide 2,4-D and other chemical stresses in yeast.

PubMed

Cabrito, Tânia R; Teixeira, Miguel C; Duarte, Alexandra A; Duque, Paula; Sá-Correia, Isabel

2009-10-01

The understanding of the molecular mechanisms underlying acquired herbicide resistance is crucial in dealing with the emergence of resistant weeds. Saccharomyces cerevisiae has been used as a model system to gain insights into the mechanisms underlying resistance to the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The TPO1 gene, encoding a multidrug resistance (MDR) plasma membrane transporter of the major facilitator superfamily (MFS), was previously found to confer resistance to 2,4-D in yeast and to be transcriptionally activated in response to the herbicide. In this work, we demonstrate that Tpo1p is required to reduce the intracellular concentration of 2,4-D. ScTpo1p homologs encoding putative plasma membrane MFS transporters from the plant model Arabidopsis thaliana were analyzed for a possible role in 2,4-D resistance. At5g13750 was chosen for further analysis, as its transcript levels were found to increase in 2,4-D stressed plants. The functional heterologous expression of this plant open reading frame in yeast was found to confer increased resistance to the herbicide in Deltatpo1 and wild-type cells, through the reduction of the intracellular concentration of 2,4-D. Heterologous expression of At5g13750 in yeast also leads to increased resistance to indole-3-acetic acid (IAA), Al(3+) and Tl(3+). At5g13750 is the first plant putative MFS transporter to be suggested as possibly involved in MDR.

Cloning and characterization of the metE gene encoding S-adenosylmethionine synthetase from Bacillus subtilis.

PubMed Central

Yocum, R R; Perkins, J B; Howitt, C L; Pero, J

1996-01-01

The metE gene, encoding S-adenosylmethionine synthetase (EC 2.5.1.6) from Bacillus subtilis, was cloned in two steps by normal and inverse PCR. The DNA sequence of the metE gene contains an open reading frame which encodes a 400-amino-acid sequence that is homologous to other known S-adenosylmethionine synthetases. The cloned gene complements the metE1 mutation and integrates at or near the chromosomal site of metE1. Expression of S-adenosylmethionine synthetase is reduced by only a factor of about 2 by exogenous methioinine. Overproduction of S-adenosylmethionine synthetase from a strong constitutive promoter leads to methionine auxotrophy in B. subtilis, suggesting that S-adenosylmethionine is a corepressor of methionine biosynthesis in B. subtilis, as others have already shown for Escherichia coli. PMID:8755891

Cloning and characterization of the metE gene encoding S-adenosylmethionine synthetase from Bacillus subtilis.

PubMed

Yocum, R R; Perkins, J B; Howitt, C L; Pero, J

1996-08-01

The metE gene, encoding S-adenosylmethionine synthetase (EC 2.5.1.6) from Bacillus subtilis, was cloned in two steps by normal and inverse PCR. The DNA sequence of the metE gene contains an open reading frame which encodes a 400-amino-acid sequence that is homologous to other known S-adenosylmethionine synthetases. The cloned gene complements the metE1 mutation and integrates at or near the chromosomal site of metE1. Expression of S-adenosylmethionine synthetase is reduced by only a factor of about 2 by exogenous methioinine. Overproduction of S-adenosylmethionine synthetase from a strong constitutive promoter leads to methionine auxotrophy in B. subtilis, suggesting that S-adenosylmethionine is a corepressor of methionine biosynthesis in B. subtilis, as others have already shown for Escherichia coli.

GDP-L-fucose: {beta}-D-galactoside 2-{alpha}-Lfucosyltransferases, DNA sequences encoding the same, method for producing the same and a method of genotyping a person

DOEpatents

Lowe, J.B.; Lennon, G.; Rouquier, S.; Giorgi, D.; Kelly, R.J.

1998-09-15

The gene encoding GDP-L-fucose: {beta}-D-Galactoside 2-{alpha}-Lfucosyltransferase has been cloned, and a mutation in this gene has been found to be responsible for an individual being a non-secretor. 30 figs.

Similarity and functional analyses of expressed parasitism genes in Heterodera schachtii and Heterodera glycines

USDA-ARS?s Scientific Manuscript database

The secreted proteins encoded by “parasitism genes” expressed within the esophageal glands cells of cyst nematodes play important roles in plant parasitism. Homologous transcripts and encoded proteins of the Heterodera glycines pioneer parasitism genes Hgsyv46, Hg4e02 and Hg5d08 were identified and ...

Two Bombyx mori acetylcholinesterase genes influence motor control and development in different ways

USDA-ARS?s Scientific Manuscript database

Among its other biological roles, acetylcholinesterase (AChE, EC 3.1.1.7), encoded by two ace genes in most insects, catalyses the breakdown of acetylcholine, thereby terminating synaptic transmission. ace1 encodes the synaptic enzyme and ace2 has other essential actions in many insect species, such...

Large-scale analysis of antisense transcription in wheat using the Affymetrix GeneChip Wheat Genome Array

USDA-ARS?s Scientific Manuscript database

Natural antisense transcripts (NATs) are transcripts of the opposite DNA strand to the sense-strand either at the same locus (cis-encoded) or a different locus (trans-encoded). They can affect gene expression at multiple stages including transcription, RNA processing and transport, and translation....

Influence of energy supply on expression of genes encoding for lipogenic enzymes and regulatory proteins in growing beef steers

USDA-ARS?s Scientific Manuscript database

Forty crossbred beef steers were used to determine the effects metabolizable energy (ME) intake and of site and complexity of carbohydrate (CHO) infusion on expression of genes encoding lipogenic enzymes and regulatory proteins in subcutaneous (SC), mesenteric (MES) and omental (OM) adipose. Treatm...

GDP-L-fucose: .beta.-D-galactoside 2-.alpha.-L-fucosyltransferases, DNA sequences encoding the same, method for producing the same and a method of genotyping a person

DOEpatents

Lowe, John B.; Lennon, Gregory; Rouquier, Sylvie; Giorgi, Dominique; Kelly, Robert J.

1998-01-01

The gene encoding GDP-L-fucose: .beta.-D-Galactoside 2-.alpha.-L-fucosyltransferase has been cloned, and a mutation in this gene has been found to be responsible for an individual being a non-secretor.

Circulating exosomes potentiate tumor malignant properties in a mouse model of chronic sleep fragmentation

PubMed Central

Khalyfa, Abdelnaby; Almendros, Isaac; Gileles-Hillel, Alex; Akbarpour, Mahzad; Trzepizur, Wojciech; Mokhlesi, Babak; Huang, Lei; Andrade, Jorge; Farré, Ramon; Gozal, David

2016-01-01

Background Chronic sleep fragmentation (SF) increases cancer aggressiveness in mice. Exosomes exhibit pleiotropic biological functions, including immune regulatory functions, antigen presentation, intracellular communication and inter-cellular transfer of RNA and proteins. We hypothesized that SF-induced alterations in biosynthesis and cargo of plasma exosomes may affect tumor cell properties. Results SF-derived exosomes increased tumor cell proliferation (~13%), migration (~2.3-fold) and extravasation (~10%) when compared to exosomes from SC-exposed mice. Similarly, Pre exosomes from OSA patients significantly enhanced proliferation and migration of human adenocarcinoma cells compared to Post. SF-exosomal cargo revealed 3 discrete differentially expressed miRNAs, and exploration of potential mRNA targets in TC1 tumor cells uncovered 132 differentially expressed genes that encode for multiple cancer-related pathways. Methods Plasma-derived exosomes from C57/B6 mice exposed to 6 wks of SF or sleep control (SC), and from adult human patients with obstructive sleep apnea (OSA) before (Pre) and after adherent treatment for 6 wks (Post) were co-cultured with mouse lung TC1 or human adenocarcinoma tumor cell lines, respectively. Proliferation, migration, invasion, endothelial barrier integrity and extravasation assays of tumor cells were performed. Plasma mouse exosomal miRNAs were profiled with arrays, and transcriptomic assessments of TC1 cells exposed to SF or SC exosomes were conducted to identify gene targets. Conclusions Chronic SF induces alterations in exosomal miRNA cargo that alter the biological properties of TC1 lung tumor cells to enhance their proliferative, migratory and extravasation properties, and similar findings occur in OSA patients, in whom SF is a constitutive component of their sleep disorder. Thus, exosomes could participate, at least in part, in the adverse cancer outcomes observed in OSA. PMID:27419627

Methyltransferases acquired by lactococcal 936-type phage provide protection against restriction endonuclease activity.

PubMed

Murphy, James; Klumpp, Jochen; Mahony, Jennifer; O'Connell-Motherway, Mary; Nauta, Arjen; van Sinderen, Douwe

2014-10-01

So-called 936-type phages are among the most frequently isolated phages in dairy facilities utilising Lactococcus lactis starter cultures. Despite extensive efforts to control phage proliferation and decades of research, these phages continue to negatively impact cheese production in terms of the final product quality and consequently, monetary return. Whole genome sequencing and in silico analysis of three 936-type phage genomes identified several putative (orphan) methyltransferase (MTase)-encoding genes located within the packaging and replication regions of the genome. Utilising SMRT sequencing, methylome analysis was performed on all three phages, allowing the identification of adenine modifications consistent with N-6 methyladenine sequence methylation, which in some cases could be attributed to these phage-encoded MTases. Heterologous gene expression revealed that M.Phi145I/M.Phi93I and M.Phi93DAM, encoded by genes located within the packaging module, provide protection against the restriction enzymes HphI and DpnII, respectively, representing the first functional MTases identified in members of 936-type phages. SMRT sequencing technology enabled the identification of the target motifs of MTases encoded by the genomes of three lytic 936-type phages and these MTases represent the first functional MTases identified in this species of phage. The presence of these MTase-encoding genes on 936-type phage genomes is assumed to represent an adaptive response to circumvent host encoded restriction-modification systems thereby increasing the fitness of the phages in a dynamic dairy environment.

Dynamics of biomass partitioning, stem gene expression, cell wall biosynthesis, and sucrose accumulation during development of Sorghum bicolor.

PubMed

McKinley, Brian; Rooney, William; Wilkerson, Curtis; Mullet, John

2016-11-01

Biomass accumulated preferentially in leaves of the sweet sorghum Della until floral initiation, then stems until anthesis, followed by panicles until grain maturity, and apical tillers. Sorghum stem RNA-seq transcriptome profiles and composition data were collected for approximately 100 days of development beginning at floral initiation. The analysis identified >200 differentially expressed genes involved in stem growth, cell wall biology, and sucrose accumulation. Genes encoding expansins and xyloglucan endotransglucosylase/hydrolases were differentially expressed in growing stem internodes. Genes encoding enzymes involved in the synthesis of cellulose, lignin, and glucuronoarabinoxylan were expressed at elevated levels in stems until approximately 7 days before anthesis and then down-regulated. CESA genes involved in primary and secondary cell wall synthesis showed different temporal patterns of expression. Following floral initiation, the level of sucrose and other non-structural carbohydrates increased to approximately 50% of the stem's dry weight. Stem sucrose accumulation was inversely correlated with >100-fold down-regulation of SbVIN1, a gene encoding a vacuolar invertase. Accumulation of stem sucrose was also correlated with cessation of leaf and stem growth at anthesis, decreased expression of genes involved in stem cell wall synthesis, and approximately 10-fold lower expression of SbSUS4, a gene encoding sucrose synthase that generates UDP-glucose from sucrose for cell wall biosynthesis. Genes for mixed linkage glucan synthesis (CSLF) and turnover were expressed at high levels in stems throughout development. Overall, the stem transcription profile resource and the genes and regulatory dynamics identified in this study will be useful for engineering sorghum stem composition for improved conversion to biofuels and bio-products. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.