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Sample records for distinct subunit species

  1. A distinct holoenzyme organization for two-subunit pyruvate carboxylase

    PubMed Central

    Choi, Philip H.; Jo, Jeanyoung; Lin, Yu-Cheng; Lin, Min-Han; Chou, Chi-Yuan; Dietrich, Lars E. P.; Tong, Liang

    2016-01-01

    Pyruvate carboxylase (PC) has important roles in metabolism and is crucial for virulence for some pathogenic bacteria. PC contains biotin carboxylase (BC), carboxyltransferase (CT) and biotin carboxyl carrier protein (BCCP) components. It is a single-chain enzyme in eukaryotes and most bacteria, and functions as a 500 kD homo-tetramer. In contrast, PC is a two-subunit enzyme in a collection of Gram-negative bacteria, with the α subunit containing the BC and the β subunit the CT and BCCP domains, and it is believed that the holoenzyme has α4β4 stoichiometry. We report here the crystal structures of a two-subunit PC from Methylobacillus flagellatus. Surprisingly, our structures reveal an α2β4 stoichiometry, and the overall architecture of the holoenzyme is strikingly different from that of the homo-tetrameric PCs. Biochemical and mutagenesis studies confirm the stoichiometry and other structural observations. Our functional studies in Pseudomonas aeruginosa show that its two-subunit PC is important for colony morphogenesis. PMID:27708276

  2. Identification of Four Distinct Subunit Types in the Unique 6×6 Hemocyanin of the Centipede Scutigera coleoptrata

    NASA Astrophysics Data System (ADS)

    Gebauer, W.; Markl, J.

    We isolated 6×6 hemocyanin, dissociated it into subunits, and examined it by electron microscopy. The subunits were separated by native polyacrylamide gel electrophoresis (PAGE), sodium dodecyl sulfate PAGE, and crossed immunoelectrophoresis. Single subunits were isolated by gel cutting from native PAGE and identified as hemocyanin by measuring their ultraviolet spectrum. A total of four distinct hemocyanin subunits were identified, and the subunit pattern of the three electrophoresis systems assigned to each other. The relative proportion of subunits a:b:c:d were 2 : 2 :>: 1 as determined by densitometry. Presumably, c and d act as linkers between hexamers.

  3. Telomeric ORFs (TLOs) in Candida spp. Encode Mediator Subunits That Regulate Distinct Virulence Traits

    PubMed Central

    Hokamp, Karsten; Yeomans, Tim; Liu, Zhongle; Church, Michael; Fleming, Alastair B.; Anderson, Matthew Z.; Berman, Judith; Myers, Lawrence C.; Sullivan, Derek J.; Moran, Gary P.

    2014-01-01

    The TLO genes are a family of telomere-associated ORFs in the fungal pathogens Candida albicans and C. dubliniensis that encode a subunit of the Mediator complex with homology to Med2. The more virulent pathogen C. albicans has 15 copies of the gene whereas the less pathogenic species C. dubliniensis has only two (CdTLO1 and CdTLO2). In this study we used C. dubliniensis as a model to investigate the role of TLO genes in regulating virulence and also to determine whether TLO paralogs have evolved to regulate distinct functions. A C. dubliniensis tlo1Δ/tlo2Δ mutant is unable to form true hyphae, has longer doubling times in galactose broth, is more susceptible to oxidative stress and forms increased levels of biofilm. Transcript profiling of the tlo1Δ/tlo2Δ mutant revealed increased expression of starvation responses in rich medium and retarded expression of hypha-induced transcripts in serum. ChIP studies indicated that Tlo1 binds to many ORFs including genes that exhibit high and low expression levels under the conditions analyzed. The altered expression of these genes in the tlo1Δ/tlo2Δ null mutant indicates roles for Tlo proteins in transcriptional activation and repression. Complementation of the tlo1Δ/tlo2Δ mutant with TLO1, but not TLO2, restored wild-type filamentous growth, whereas only TLO2 fully suppressed biofilm growth. Complementation with TLO1 also had a greater effect on doubling times in galactose broth. The different abilities of TLO1 and TLO2 to restore wild-type functions was supported by transcript profiling studies that showed that only TLO1 restored expression of hypha-specific genes (UME6, SOD5) and galactose utilisation genes (GAL1 and GAL10), whereas TLO2 restored repression of starvation-induced gene transcription. Thus, Tlo/Med2 paralogs encoding Mediator subunits regulate different virulence properties in Candida spp. and their expansion may account for the increased adaptability of C. albicans relative to other Candida species

  4. Distinct Structural Pathways Coordinate the Activation of AMPA Receptor-Auxiliary Subunit Complexes

    PubMed Central

    Dawe, G. Brent; Musgaard, Maria; Aurousseau, Mark R.P.; Nayeem, Naushaba; Green, Tim; Biggin, Philip C.; Bowie, Derek

    2016-01-01

    Summary Neurotransmitter-gated ion channels adopt different gating modes to fine-tune signaling at central synapses. At glutamatergic synapses, high and low activity of AMPA receptors (AMPARs) is observed when pore-forming subunits coassemble with or without auxiliary subunits, respectively. Whether a common structural pathway accounts for these different gating modes is unclear. Here, we identify two structural motifs that determine the time course of AMPAR channel activation. A network of electrostatic interactions at the apex of the AMPAR ligand-binding domain (LBD) is essential for gating by pore-forming subunits, whereas a conserved motif on the lower, D2 lobe of the LBD prolongs channel activity when auxiliary subunits are present. Accordingly, channel activity is almost entirely abolished by elimination of the electrostatic network but restored via auxiliary protein interactions at the D2 lobe. In summary, we propose that activation of native AMPAR complexes is coordinated by distinct structural pathways, favored by the association/dissociation of auxiliary subunits. PMID:26924438

  5. Distinct Structural Pathways Coordinate the Activation of AMPA Receptor-Auxiliary Subunit Complexes.

    PubMed

    Dawe, G Brent; Musgaard, Maria; Aurousseau, Mark R P; Nayeem, Naushaba; Green, Tim; Biggin, Philip C; Bowie, Derek

    2016-03-16

    Neurotransmitter-gated ion channels adopt different gating modes to fine-tune signaling at central synapses. At glutamatergic synapses, high and low activity of AMPA receptors (AMPARs) is observed when pore-forming subunits coassemble with or without auxiliary subunits, respectively. Whether a common structural pathway accounts for these different gating modes is unclear. Here, we identify two structural motifs that determine the time course of AMPAR channel activation. A network of electrostatic interactions at the apex of the AMPAR ligand-binding domain (LBD) is essential for gating by pore-forming subunits, whereas a conserved motif on the lower, D2 lobe of the LBD prolongs channel activity when auxiliary subunits are present. Accordingly, channel activity is almost entirely abolished by elimination of the electrostatic network but restored via auxiliary protein interactions at the D2 lobe. In summary, we propose that activation of native AMPAR complexes is coordinated by distinct structural pathways, favored by the association/dissociation of auxiliary subunits. PMID:26924438

  6. A Single Mutation in the Acetylcholine Receptor δ-Subunit Causes Distinct Effects in Two Types of Neuromuscular Synapses

    PubMed Central

    Park, Jee-Young; Mott, Meghan; Williams, Tory; Ikeda, Hiromi; Wen, Hua; Linhoff, Michael

    2014-01-01

    Mutations in AChR subunits, expressed as pentamers in neuromuscular junctions (NMJs), cause various types of congenital myasthenic syndromes. In AChR pentamers, the adult ε subunit gradually replaces the embryonic γ subunit as the animal develops. Because of this switch in subunit composition, mutations in specific subunits result in synaptic phenotypes that change with developmental age. However, a mutation in any AChR subunit is considered to affect the NMJs of all muscle fibers equally. Here, we report a zebrafish mutant of the AChR δ subunit that exhibits two distinct NMJ phenotypes specific to two muscle fiber types: slow or fast. Homozygous fish harboring a point mutation in the δ subunit form functional AChRs in slow muscles, whereas receptors in fast muscles are nonfunctional. To test the hypothesis that different subunit compositions in slow and fast muscles underlie distinct phenotypes, we examined the presence of ε/γ subunits in NMJs using specific antibodies. Both wild-type and mutant larvae lacked ε/γ subunits in slow muscle synapses. These findings in zebrafish suggest that some mutations in human congenital myasthenic syndromes may affect slow and fast muscle fibers differently. PMID:25080583

  7. Thermostable Cross-Protective Subunit Vaccine against Brucella Species

    PubMed Central

    Barabé, Nicole D.; Grigat, Michelle L.; Lee, William E.; Poirier, Robert T.; Jager, Scott J.; Berger, Bradley J.

    2014-01-01

    A subunit vaccine candidate was produced from Brucella suis 145 (biovar 4; expressing both the A antigen of Brucella abortus and the M antigen of Brucella melitensis). The preparation consisted mostly of polysaccharide (PS; >90% [wt/wt]; both cell-associated PS and exo-PS were combined) and a small amount of protein (1 to 3%) with no apparent nucleic acids. Vaccinated mice were protected (these had a statistically significant reduction in bacterial colonization compared to that of unvaccinated controls) when challenged with representative strains of three Brucella species most pathogenic for humans, i.e., B. abortus, B. melitensis, and B. suis. As little as 1 ng of the vaccine, without added adjuvant, protected mice against B. suis 145 infection (5 × 105 CFU), and a single injection of 1 μg of this subunit vaccine protected mice from B. suis 145 challenge for at least 14 months. A single immunization induced a serum IgG response to Brucella antigens that remained elevated for up to 9 weeks. The use of heat (i.e., boiling-water bath, autoclaving) in the vaccine preparation showed that it was thermostable. This method also ensured safety and security. The vaccine produced was immunogenic and highly protective against multiple strains of Brucella and represents a promising candidate for further evaluation. PMID:25320267

  8. Class II ribonuclease H comigrates with, but is distinct from, the third largest subunit of calf thymus RNA polymerase I.

    PubMed

    Vonwirth, H; Frank, P; Kedinger, C; Büsen, W

    1990-09-10

    It has been reported (Iborra et al. (1979) J. Biol. Chem. 254, 10920-10924) that the third and the fifth largest subunit of yeast RNA polymerase I exhibit ribonuclease H activity. The authors suggested that the third largest subunit is identical with the chromatin-associated ribonuclease H49, the putative yeast equivalent of bovine ribonuclease H IIb. Although the third largest subunit of calf thymus RNA polymerase I and ribonuclease H IIb display nearly identical molecular masses under denaturing conditions, serological analysis reveals that, in contrast to their counterparts in yeast, these mammalian proteins are distinct entities. Interestingly, sera from some patients with mixed connective tissue disease which contain antibodies directed against RNA polymerase I, also react with ribonuclease H IIb epitopes. This observation suggests that a protein displaying ribonuclease H IIb antigenicity could be associated with RNA polymerase I. Additional indications supporting this conclusion are discussed.

  9. Distinct Subunit Domains Govern Synaptic Stability and Specificity of the Kainate Receptor.

    PubMed

    Straub, Christoph; Noam, Yoav; Nomura, Toshihiro; Yamasaki, Miwako; Yan, Dan; Fernandes, Herman B; Zhang, Ping; Howe, James R; Watanabe, Masahiko; Contractor, Anis; Tomita, Susumu

    2016-07-12

    Synaptic communication between neurons requires the precise localization of neurotransmitter receptors to the correct synapse type. Kainate-type glutamate receptors restrict synaptic localization that is determined by the afferent presynaptic connection. The mechanisms that govern this input-specific synaptic localization remain unclear. Here, we examine how subunit composition and specific subunit domains contribute to synaptic localization of kainate receptors. The cytoplasmic domain of the GluK2 low-affinity subunit stabilizes kainate receptors at synapses. In contrast, the extracellular domain of the GluK4/5 high-affinity subunit synergistically controls the synaptic specificity of kainate receptors through interaction with C1q-like proteins. Thus, the input-specific synaptic localization of the native kainate receptor complex involves two mechanisms that underlie specificity and stabilization of the receptor at synapses.

  10. Distinct double- and single-stranded DNA binding of E. coli replicative DNA polymerase III alpha subunit.

    PubMed

    McCauley, Micah J; Shokri, Leila; Sefcikova, Jana; Venclovas, Ceslovas; Beuning, Penny J; Williams, Mark C

    2008-09-19

    The alpha subunit of the replicative DNA polymerase III of Escherichia coli is the active polymerase of the 10-subunit bacterial replicase. The C-terminal region of the alpha subunit is predicted to contain an oligonucleotide binding (OB-fold) domain. In a series of optical tweezers experiments, the alpha subunit is shown to have an affinity for both double- and single-stranded DNA, in distinct subdomains of the protein. The portion of the protein that binds to double-stranded DNA stabilizes the DNA helix, because protein binding must be at least partially disrupted with increasing force to melt DNA. Upon relaxation, the DNA fails to fully reanneal, because bound protein interferes with the reformation of the double helix. In addition, the single-stranded DNA binding component appears to be passive, as the protein does not facilitate melting but instead binds to single-stranded regions already separated by force. From DNA stretching measurements we determine equilibrium association constants for the binding of alpha and several fragments to dsDNA and ssDNA. The results demonstrate that ssDNA binding is localized to the C-terminal region that contains the OB-fold domain, while a tandem helix-hairpin-helix (HhH) 2 motif contributes significantly to dsDNA binding. PMID:18652472

  11. DNA barcoding of Sri Lankan phlebotomine sand flies using cytochrome c oxidase subunit I reveals the presence of cryptic species.

    PubMed

    Gajapathy, Kanapathy; Tharmasegaram, Tharmatha; Eswaramohan, Thampoe; Peries, Lalanthika B S L; Jayanetti, Raveendra; Surendran, Sinnathamby N

    2016-09-01

    Sri Lanka is known for high diversity of phlebotomine sand flies and prevalence of cutaneous and visceral leishmaniasis; a disease vectored by sand flies. The taxonomy of phlebotomine sand flies is complicated and often the diversity is over/underrated. The current study aims to use the cytochrome c oxidase gene subunit 1 (COI) sequence and formulate a barcode for the sand fly species in Sri Lanka. A total of 70 samples comprising seven species morphologically identified and collected from dry zone districts of Hambantota, Anuradhapura, Vavuniya, Trincomalee and Jaffna were processed. Neighbour-joining (NJ) tree created using the sequences revealed the species identity is compatible with the current morphology based identification. Further the analysis delineated morphologically identified Se. bailyi, Se babu babu and Se babu insularis into genetically distinct groups. PMID:27180216

  12. Identification of small subunits of mammalian serine palmitoyltransferase that confer distinct acyl-CoA substrate specificities.

    PubMed

    Han, Gongshe; Gupta, Sita D; Gable, Kenneth; Niranjanakumari, Somashekarappa; Moitra, Prasun; Eichler, Florian; Brown, Robert H; Harmon, Jeffrey M; Dunn, Teresa M

    2009-05-19

    Serine palmitoyltransferase (SPT) catalyzes the first committed step in sphingolipid biosynthesis. In yeast, SPT is composed of a heterodimer of 2 highly-related subunits, Lcb1p and Lcb2p, and a third subunit, Tsc3p, which increases enzyme activity markedly and is required for growth at elevated temperatures. Higher eukaryotic orthologs of Lcb1p and Lcb2p have been identified, but SPT activity is not highly correlated with coexpression of these subunits and no ortholog of Tsc3p has been identified. Here, we report the discovery of 2 proteins, ssSPTa and ssSPTb, which despite sharing no homology with Tsc3p, each substantially enhance the activity of mammalian SPT expressed in either yeast or mammalian cells and therefore define an evolutionarily conserved family of low molecular weight proteins that confer full enzyme activity. The 2 ssSPT isoforms share a conserved hydrophobic central domain predicted to reside in the membrane, and each interacts with both hLCB1 and hLCB2 as assessed by positive split ubiquitin 2-hybrid analysis. The presence of these small subunits, along with 2 hLCB2 isofoms, suggests that there are 4 distinct human SPT isozymes. When each SPT isozyme was expressed in either yeast or CHO LyB cells lacking endogenous SPT activity, characterization of their in vitro enzymatic activities, and long-chain base (LCB) profiling revealed differences in acyl-CoA preference that offer a potential explanation for the observed diversity of LCB seen in mammalian cells. PMID:19416851

  13. Assembly of the Arp5 (Actin-related Protein) Subunit Involved in Distinct INO80 Chromatin Remodeling Activities.

    PubMed

    Yao, Wei; Beckwith, Sean L; Zheng, Tina; Young, Thomas; Dinh, Van T; Ranjan, Anand; Morrison, Ashby J

    2015-10-16

    ATP-dependent chromatin remodeling, which repositions and restructures nucleosomes, is essential to all DNA-templated processes. The INO80 chromatin remodeling complex is an evolutionarily conserved complex involved in diverse cellular processes, including transcription, DNA repair, and replication. The functional diversity of the INO80 complex can, in part, be attributed to specialized activities of distinct subunits that compose the complex. Furthermore, structural analyses have identified biochemically discrete subunit modules that assemble along the Ino80 ATPase scaffold. Of particular interest is the Saccharomyces cerevisiae Arp5-Ies6 module located proximal to the Ino80 ATPase and the Rvb1-Rvb2 helicase module needed for INO80-mediated in vitro activity. In this study we demonstrate that the previously uncharacterized Ies2 subunit is required for Arp5-Ies6 association with the catalytic components of the INO80 complex. In addition, Arp5-Ies6 module assembly with the INO80 complex is dependent on distinct conserved domains within Arp5, Ies6, and Ino80, including the spacer region within the Ino80 ATPase domain. Arp5-Ies6 interacts with chromatin via assembly with the INO80 complex, as IES2 and INO80 deletion results in loss of Arp5-Ies6 chromatin association. Interestingly, ectopic addition of the wild-type Arp5-Ies6 module stimulates INO80-mediated ATP hydrolysis and nucleosome sliding in vitro. However, the addition of mutant Arp5 lacking unique insertion domains facilitates ATP hydrolysis in the absence of nucleosome sliding. Collectively, these results define the requirements of Arp5-Ies6 assembly, which are needed to couple ATP hydrolysis to productive nucleosome movement.

  14. A distinct three-helix centipede toxin SSD609 inhibits Iks channels by interacting with the KCNE1 auxiliary subunit

    PubMed Central

    Sun, Peibei; Wu, Fangming; Wen, Ming; Yang, Xingwang; Wang, Chenyang; Li, Yiming; He, Shufang; Zhang, Longhua; Zhang, Yun; Tian, Changlin

    2015-01-01

    KCNE1 is a single-span transmembrane auxiliary protein that modulates the voltage-gated potassium channel KCNQ1. The KCNQ1/KCNE1 complex in cardiomyocytes exhibited slow activated potassium (Iks) currents. Recently, a novel 47-residue polypeptide toxin SSD609 was purified from Scolopendra subspinipes dehaani venom and showed Iks current inhibition. Here, chemically synthesized SSD609 was shown to exert Iks inhibition in extracted guinea pig cardiomyocytes and KCNQ1/KCNE1 current attenuation in CHO cells. The K+ current attenuation of SSD609 showed decent selectivity among different auxiliary subunits. Solution nuclear magnetic resonance analysis of SSD609 revealed a distinctive three-helix conformation that was stabilized by a new disulfide bonding pattern as well as segregated surface charge distribution. Structure-activity studies demonstrated that negatively charged Glu19 in the amphipathic extracellular helix of KCNE1 was the key residue that interacted with SSD609. The distinctive three-helix centipede toxin SSD609 is known to be the first polypeptide toxin acting on channel auxiliary subunit KCNE1, which suggests a new type of pharmacological regulation for ion channels in cardiomyocytes. PMID:26307551

  15. Molecular evidence for species-level distinctions in clouded leopards.

    PubMed

    Buckley-Beason, Valerie A; Johnson, Warren E; Nash, Willliam G; Stanyon, Roscoe; Menninger, Joan C; Driscoll, Carlos A; Howard, JoGayle; Bush, Mitch; Page, John E; Roelke, Melody E; Stone, Gary; Martelli, Paolo P; Wen, Ci; Ling, Lin; Duraisingam, Ratna K; Lam, Phan V; O'Brien, Stephen J

    2006-12-01

    Among the 37 living species of Felidae, the clouded leopard (Neofelis nebulosa) is generally classified as a monotypic genus basal to the Panthera lineage of great cats. This secretive, mid-sized (16-23 kg) carnivore, now severely endangered, is traditionally subdivided into four southeast Asian subspecies (Figure 1A). We used molecular genetic methods to re-evaluate subspecies partitions and to quantify patterns of population genetic variation among 109 clouded leopards of known geographic origin (Figure 1A, Tables S1 ans S2 in the Supplemental Data available online). We found strong phylogeographic monophyly and large genetic distances between N. n. nebulosa (mainland) and N. n. diardi (Borneo; n = 3 individuals) with mtDNA (771 bp), nuclear DNA (3100 bp), and 51 microsatellite loci. Thirty-six fixed mitochondrial and nuclear nucleotide differences and 20 microsatellite loci with nonoverlapping allele-size ranges distinguished N. n. nebulosa from N. n. diardi. Along with fixed subspecies-specific chromosomal differences, this degree of differentiation is equivalent to, or greater than, comparable measures among five recognized Panthera species (lion, tiger, leopard, jaguar, and snow leopard). These distinctions increase the urgency of clouded leopard conservation efforts, and if affirmed by morphological analysis and wider sampling of N. n. diardi in Borneo and Sumatra, would support reclassification of N. n. diardi as a new species (Neofelis diardi).

  16. Molecular evidence for species-level distinctions in clouded leopards.

    PubMed

    Buckley-Beason, Valerie A; Johnson, Warren E; Nash, Willliam G; Stanyon, Roscoe; Menninger, Joan C; Driscoll, Carlos A; Howard, JoGayle; Bush, Mitch; Page, John E; Roelke, Melody E; Stone, Gary; Martelli, Paolo P; Wen, Ci; Ling, Lin; Duraisingam, Ratna K; Lam, Phan V; O'Brien, Stephen J

    2006-12-01

    Among the 37 living species of Felidae, the clouded leopard (Neofelis nebulosa) is generally classified as a monotypic genus basal to the Panthera lineage of great cats. This secretive, mid-sized (16-23 kg) carnivore, now severely endangered, is traditionally subdivided into four southeast Asian subspecies (Figure 1A). We used molecular genetic methods to re-evaluate subspecies partitions and to quantify patterns of population genetic variation among 109 clouded leopards of known geographic origin (Figure 1A, Tables S1 ans S2 in the Supplemental Data available online). We found strong phylogeographic monophyly and large genetic distances between N. n. nebulosa (mainland) and N. n. diardi (Borneo; n = 3 individuals) with mtDNA (771 bp), nuclear DNA (3100 bp), and 51 microsatellite loci. Thirty-six fixed mitochondrial and nuclear nucleotide differences and 20 microsatellite loci with nonoverlapping allele-size ranges distinguished N. n. nebulosa from N. n. diardi. Along with fixed subspecies-specific chromosomal differences, this degree of differentiation is equivalent to, or greater than, comparable measures among five recognized Panthera species (lion, tiger, leopard, jaguar, and snow leopard). These distinctions increase the urgency of clouded leopard conservation efforts, and if affirmed by morphological analysis and wider sampling of N. n. diardi in Borneo and Sumatra, would support reclassification of N. n. diardi as a new species (Neofelis diardi). PMID:17141620

  17. Partial protection against four species of chicken coccidia induced by multivalent subunit vaccine.

    PubMed

    Song, Xiaokai; Gao, Yunlu; Xu, Lixin; Yan, Ruofeng; Li, Xiangrui

    2015-09-15

    In this study, a multivalent subunit vaccine was designed to protect chickens against simultaneous infection by several Eimeria species. This vaccine contains recombinant proteins from four Eimeria species - E. tenella, E. necatrix, E. acervulina and E. maxima - and was evaluated for efficacy in animals. To produce this vaccine, candidate antigens from each Eimeria species were first screened in chickens via intramuscular inoculation and subsequent challenge. Antigens tested include recombinant proteins TA4 and SO7 from E. tenella, NA4 and NPmz19 from E. necatrix, LDH, 3-1E and MIF from E. acervulina, and Em6 and Em8 (two portions of EmTFP250) from E. maxima. A homologous challenge was then performed to identify which antigen from each species conferred the best protection. The antigens identified as most protective against its species were then challenged by heterologous species. Finally, the selected recombinant proteins from each of the four respective species were mixed with the final concentration of 400 μg/ml (100 μg of each protein/ml) to form the multivalent subunit vaccine, which was tested for efficacy in animals. The results indicated that TA4 from E. tenella, NA4 from E. necatrix, LDH from E. acervulina, and Em8 from E. maxima each induced the most effective protection from homologous challenge. Cross-protection results showed that TA4 provided partial cross-protection against E. necatrix, NA4 provided partial cross-protection against E. tenella and E. acervulina, LDH provided partial cross-protection against E. tenella and E. necatrix, and Em8 provided partial cross-protection against E. tenella and E. acervulina. The multivalent subunit vaccine provided partial protection against E. tenella, E. necatrix, E. acervulina and E. maxima challenge, and resulted in ACIs of more than 170. These results suggest that our candidate multivalent vaccine could protect chickens against simultaneous infection by several Eimeria species.

  18. Lungfish aestivating activities are locked in distinct encephalic γ-aminobutyric acid type A receptor α subunits.

    PubMed

    Giusi, Giuseppina; Crudo, Michele; Di Vito, Anna; Facciolo, Rosa Maria; Garofalo, Filippo; Chew, Shit Fun; Ip, Yuen Kwong; Canonaco, Marcello

    2011-03-01

    Ammonia in dipnoans plays a crucial role on neuronal homeostasis, especially for those brain areas that maintain torpor and awakening states in equilibrium. In the present study, specific α subunits of the major neuroreceptor inhibitory complex (GABA(A) R), which predominated during some phases of aestivation of the lungfish Protopterus annectens, turned out to be key adaptive factors of this species. From the isolation, for the first time, of the encoding sequence for GABA(A) R α₁, α₄ , and α₅ subunits in Protopterus annectens, qPCR and in situ hybridization levels of α₄ transcript in thalamic (P < 0.001) and mesencephalic (P < 0.01) areas proved to be significantly higher during long aestivating maintenance states. Very evident α₅ mRNA levels were detected in diencephalon during short inductive aestivating states, whereas an α₄ /α₁ turnover characterized the arousal state. Contextually, the recovery of physiological activities appeared to be tightly related to an evident up-regulation of α₁ transcripts in telencephalic and cerebellar sites. Surprisingly, TUNEL and amino cupric silver methods corroborated apoptotic and neurodegenerative cellular events, respectively, above all in telencephalon and cerebellum of lungfish exposed to long maintenance aestivating conditions. Overall, these results tend to underlie a novel GABAergic-related ON/OFF molecular switch operating during aestivation of the lungfish, which might have a bearing on sleeping disorders.

  19. Separation of distinct photoexcitation species in femtosecond transient absorption microscopy

    DOE PAGES

    Xiao, Kai; Ma, Ying -Zhong; Simpson, Mary Jane; Doughty, Benjamin; Yang, Bing

    2016-02-03

    Femtosecond transient absorption microscopy is a novel chemical imaging capability with simultaneous high spatial and temporal resolution. Although several powerful data analysis approaches have been developed and successfully applied to separate distinct chemical species in such images, the application of such analysis to distinguish different photoexcited species is rare. In this paper, we demonstrate a combined approach based on phasor and linear decomposition analysis on a microscopic level that allows us to separate the contributions of both the excitons and free charge carriers in the observed transient absorption response of a composite organometallic lead halide perovskite film. We found spatialmore » regions where the transient absorption response was predominately a result of excitons and others where it was predominately due to charge carriers, and regions consisting of signals from both contributors. Lastly, quantitative decomposition of the transient absorption response curves further enabled us to reveal the relative contribution of each photoexcitation to the measured response at spatially resolved locations in the film.« less

  20. Distinct requirements for signal peptidase processing and function in the stable signal peptide subunit of the Junin virus envelope glycoprotein

    SciTech Connect

    York, Joanne; Nunberg, Jack H. . E-mail: jack.nunberg@umontana.edu

    2007-03-01

    The arenavirus envelope glycoprotein (GP-C) retains a cleaved and stable signal peptide (SSP) as an essential subunit of the mature complex. This 58-amino-acid residue peptide serves as a signal sequence and is additionally required to enable transit of the assembled GP-C complex to the Golgi, and for pH-dependent membrane fusion activity. We have investigated the C-terminal region of the Junin virus SSP to study the role of the cellular signal peptidase (SPase) in generating SSP. Site-directed mutagenesis at the cleavage site (positions - 1 and - 3) reveals a pattern of side-chain preferences consistent with those of SPase. Although position - 2 is degenerate for SPase cleavage, this residue in the arenavirus SSP is invariably a cysteine. In the Junin virus, this cysteine is not involved in disulfide bonding. We show that replacement with alanine or serine is tolerated for SPase cleavage but prevents the mutant SSP from associating with GP-C and enabling transport to the cell surface. Conversely, an arginine mutation at position - 1 that prevents SPase cleavage is fully compatible with GP-C-mediated membrane fusion activity when the mutant SSP is provided in trans. These results point to distinct roles of SSP sequences in SPase cleavage and GP-C biogenesis. Further studies of the unique structural organization of the GP-C complex will be important in identifying novel opportunities for antiviral intervention against arenaviral hemorrhagic disease.

  1. Distinct expression patterns of glycoprotein hormone subunits in the lophotrochozoan Aplysia: implications for the evolution of neuroendocrine systems in animals.

    PubMed

    Heyland, Andreas; Plachetzki, David; Donelly, Evonne; Gunaratne, Dinuka; Bobkova, Yelena; Jacobson, John; Kohn, Andrea B; Moroz, Leonid L

    2012-11-01

    Glycoprotein hormones (GPHs) comprise a group of signaling molecules critical for major metabolic and reproductive functions. In vertebrates they include chorionic gonadotropin, LH, FSH, and TSH. The active hormones are characterized by heterodimerization between a common α and hormone-specific β subunit, which activate leucine-rich repeat-containing G protein coupled receptors. To date, genes referred to as GPHα2 and GPHβ5 have been the only glycoprotein hormone subunits identified in invertebrates, suggesting that other GPHα and GPHβ subunits diversified during vertebrate evolution. Still the functions of GPHα2 and GPHβ5 remain largely unknown for both vertebrates and invertebrates. To further understand the evolution and putative function of these subunits, we cloned and analyzed phylogenetically two glycoprotein subunits, AcaGPHα and AcaGPHβ, from the sea hare Aplysia californica. Model based three-dimensional predictions of AcaGPHβ confirm the presence of a complete cysteine knot, two hairpin loops, and a long loop. As in the human GPHβ5 subunit the seatbelt structure is absent in AcaGPHβ. We also found that AcaGPHα and AcaGPHβ subunits are expressed in larval stages of Aplysia, and we present a detailed expression map of the subunits in the adult central nervous system using in situ hybridizations. Both subunits are expressed in subpopulations of pleural and buccal mechanosensory neurons, suggesting a neuronal modulatory function of these subunits in Aplysia. Furthermore it supports the model of a relatively diffuse neuroendocrine-like system in molluscs, where specific primary sensory neurons release peptides extrasynaptically (paracrine secretion). This is in contrast to vertebrates and insects, in which releasing and stimulating factor from centralized sensory regions of the central nervous system ultimately regulate hormone release in peripheral glands.

  2. Distinct Contributions of T1R2 and T1R3 Taste Receptor Subunits to the Detection of Sweet Stimuli

    SciTech Connect

    Nie,Y.; Vigues, S.; Hobbs, J.; Conn, G.; Munger, S.

    2005-01-01

    The molecular mechanisms by which G protein-coupled receptor (GPCR)-type chemosensory receptors of animals selectively interact with their cognate ligands remain poorly understood. There is growing evidence that many chemosensory receptors exist in multimeric complexes, though little is known about the relative contributions of individual subunits to receptor functions. This study showed that each of the two subunits in the mammalian heteromeric T1R2:T1R3 sweet taste receptor binds sweet stimuli, though with distinct affinities and conformational changes. Furthermore, ligand affinities for T1R3 are drastically reduced by the introduction of a single amino acid change associated with decreased sweet taste sensitivity in mice. Thus, individual T1R subunits increase the receptive range of the sweet taste receptor, offering a functional mechanism for phenotypic variations in sweet taste.

  3. Distinct Effects of Different Phosphatidylglycerol Species on Mouse Keratinocyte Proliferation

    PubMed Central

    Xie, Ding; Seremwe, Mutsa; Edwards, John G.; Podolsky, Robert; Bollag, Wendy B.

    2014-01-01

    We have previously shown that liposomes composed of egg-derived phosphatidylglycerol (PG), with a mixed fatty acid composition (comprising mainly palmitate and oleate), inhibit the proliferation and promote the differentiation of rapidly dividing keratinocytes, and stimulate the growth of slowly proliferating epidermal cells. To determine the species of PG most effective at modulating keratinocyte proliferation, primary mouse keratinocytes were treated with different PG species, and proliferation was measured. PG species containing polyunsaturated fatty acids were effective at inhibiting rapidly proliferating keratinocytes, whereas PG species with monounsaturated fatty acids were effective at promoting proliferation in slowly dividing cells. Thus, palmitoyl-arachidonyl-PG (16∶0/20∶4), palmitoyl-linoleoyl-PG (16∶0/18∶2), dilinoleoyl-PG (18∶2/18∶2) and soy PG (a PG mixture with a large percentage of polyunsaturated fatty acids) were particularly effective at inhibiting proliferation in rapidly dividing keratinocytes. Conversely, palmitoyl-oleoyl-PG (16∶0/18∶1) and dioleoyl-PG (18∶1/18∶1) were especially effective proproliferative PG species. This result represents the first demonstration of opposite effects of different species of a single class of phospholipid and suggests that these different PG species may signal to diverse effector enzymes to differentially affect keratinocyte proliferation and normalize keratinocyte proliferation. Thus, different PG species may be useful for treating skin diseases characterized by excessive or insufficient proliferation. PMID:25233484

  4. p87 and p101 subunits are distinct regulators determining class IB phosphoinositide 3-kinase (PI3K) specificity.

    PubMed

    Shymanets, Aliaksei; Prajwal; Bucher, Kirsten; Beer-Hammer, Sandra; Harteneck, Christian; Nürnberg, Bernd

    2013-10-25

    Class IB phosphoinositide 3-kinase γ (PI3Kγ) comprises a single catalytic p110γ subunit, which binds to two non-catalytic subunits, p87 or p101, and controls a plethora of fundamental cellular responses. The non-catalytic subunits are assumed to be redundant adaptors for Gβγ enabling G-protein-coupled receptor-mediated regulation of PI3Kγ. Growing experimental data provide contradictory evidence. To elucidate the roles of the non-catalytic subunits in determining the specificity of PI3Kγ, we tested the impact of p87 and p101 in heterodimeric p87-p110γ and p101-p110γ complexes on the modulation of PI3Kγ activity in vitro and in living cells. RT-PCR, biochemical, and imaging data provide four lines of evidence: (i) specific expression patterns of p87 and p101, (ii) up-regulation of p101, providing the basis to consider p87 as a protein forming a constitutively and p101 as a protein forming an inducibly expressed PI3Kγ, (iii) differences in basal and stimulated enzymatic activities, and (iv) differences in complex stability, all indicating apparent diversity within class IB PI3Kγ. In conclusion, expression and activities of PI3Kγ are modified differently by p87 and p101 in vitro and in living cells, arguing for specific regulatory roles of the non-catalytic subunits in the differentiation of PI3Kγ signaling pathways. PMID:24014027

  5. Biochemical and functional properties of distinct nicotinic acetylcholine receptors in the superior cervical ganglion of mice with targeted deletions of nAChR subunit genes.

    PubMed

    David, Reinhard; Ciuraszkiewicz, Anna; Simeone, Xenia; Orr-Urtreger, Avi; Papke, Roger L; McIntosh, J M; Huck, Sigismund; Scholze, Petra

    2010-03-01

    Nicotinic acetylcholine receptors (nAChRs) mediate fast synaptic transmission in ganglia of the autonomic nervous system. Here, we determined the subunit composition of hetero-pentameric nAChRs in the mouse superior cervical ganglion (SCG), the function of distinct receptors (obtained by deletions of nAChR subunit genes) and mechanisms at the level of nAChRs that might compensate for the loss of subunits. As shown by immunoprecipitation and Western blots, wild-type (WT) mice expressed: alpha 3 beta 4 (55%), alpha 3 beta 4 alpha 5 (24%) and alpha 3 beta 4 beta 2 (21%) nAChRs. nAChRs in beta 4 knockout (KO) mice were reduced to < 15% of controls and no longer contained the alpha 5 subunit. Compound action potentials, recorded from the postganglionic (internal carotid) nerve and induced by preganglionic nerve stimulation, did not differ between alpha 5 beta 4 KO and WT mice, suggesting that the reduced number of receptors in the KO mice did not impair transganglionic transmission. Deletions of alpha 5 or beta2 did not affect the overall number of receptors and we found no evidence that the two subunits substitute for each other. In addition, dual KOs allowed us to study the functional properties of distinct alpha 3 beta4 and alpha 3 beta 2 receptors that have previously only been investigated in heterologous expression systems. The two receptors strikingly differed in the decay of macroscopic currents, the efficacy of cytisine, and their responses to the alpha-conotoxins AuIB and MII. Our data, based on biochemical and functional experiments and several mouse KO models, clarify and significantly extend previous observations on the function of nAChRs in heterologous systems and the SCG. PMID:20377613

  6. Co-existing grass species have distinctive arbuscular mycorrhizal communities.

    PubMed

    Vandenkoornhuyse, P; Ridgway, K P; Watson, I J; Fitter, A H; Young, J P W

    2003-11-01

    Arbuscular mycorrhizal (AM) fungi are biotrophic symbionts colonizing the majority of land plants, and are of major importance in plant nutrient supply. Their diversity is suggested to be an important determinant of plant community structure, but the influence of host-plant and environmental factors on AM fungal community in plant roots is poorly documented. Using the terminal restriction fragment length polymorphism (T-RFLP) strategy, the diversity of AM fungi was assessed in 89 roots of three grass species (Agrostis capillaris, Festuca rubra, Poa pratensis) that co-occurred in the same plots of a field experiment. The impact of different soil amendments (nitrogen, lime, nitrogen and lime) and insecticide application on AM fungal community was also studied. The level of diversity found in AM fungal communities using the T-RFLP strategy was consistent with previous studies based on clone libraries. Our results clearly confirm that an AM fungal host-plant preference exists, even between different grass species. AM communities colonizing A. capillaris were statistically different from the others (P < 0.05). Although grass species evenness changed in amended soils, AM fungal community composition in roots of a given grass species remained stable. Conversely, in plots where insecticide was applied, we found higher AM fungal diversity and, in F. rubra roots, a statistically different AM fungal community.

  7. Subunits of the Heterotrimeric Transcription Factor NF-Y Are Imported into the Nucleus by Distinct Pathways Involving Importin β and Importin 13

    PubMed Central

    Kahle, Joerg; Baake, Matthias; Doenecke, Detlef; Albig, Werner

    2005-01-01

    The transcriptional activator NF-Y is a heterotrimeric complex composed of NF-YA, NF-YB, and NF-YC, which specifically binds the CCAAT consensus present in about 30% of eukaryotic promoters. All three subunits contain evolutionarily conserved core regions, which comprise a histone fold motif (HFM) in the case of NF-YB and NF-YC. Our results of in vitro binding studies and nuclear import assays reveal two different transport mechanisms for NF-Y subunits. While NF-YA is imported by an importin β-mediated pathway, the NF-YB/NF-YC heterodimer is translocated into the nucleus in an importin 13-dependent manner. We define a nonclassical nuclear localization signal (ncNLS) in NF-YA, and mutational analysis indicates that positively charged amino acid residues in the ncNLS are required for nuclear targeting of NF-YA. Importin β binding is restricted to the monomeric, uncomplexed NF-YA subunit. In contrast, the nuclear import of NF-YB and NF-YC requires dimer formation. Only the NF-YB/NF-YC dimer, but not the monomeric components, are recognized by importin 13 and are imported into the nucleus. Importin 13 competes with NF-YA for binding to the NF-YB/NF-YC dimer. Our data suggest that a distinct binding platform derived from the HFM of both subunits, NF-YB/NF-YC, mediates those interactions. PMID:15964792

  8. Distribution of Plasmids in Distinct Leptospira Pathogenic Species.

    PubMed

    Wang, Yanzhuo; Zhuang, Xuran; Zhong, Yi; Zhang, Cuicai; Zhang, Yan; Zeng, Lingbing; Zhu, Yongzhang; He, Ping; Dong, Ke; Pal, Utpal; Guo, Xiaokui; Qin, Jinhong

    2015-11-01

    Leptospirosis, caused by pathogenic Leptospira, is a worldwide zoonotic infection. The genus Leptospira includes at least 21 species clustered into three groups--pathogens, non-pathogens, and intermediates--based on 16S rRNA phylogeny. Research on Leptospira is difficult due to slow growth and poor transformability of the pathogens. Recent identification of extrachromosomal elements besides the two chromosomes in L. interrogans has provided new insight into genome complexity of the genus Leptospira. The large size, low copy number, and high similarity of the sequence of these extrachromosomal elements with the chromosomes present challenges in isolating and detecting them without careful genome assembly. In this study, two extrachromosomal elements were identified in L. borgpetersenii serovar Ballum strain 56604 through whole genome assembly combined with S1 nuclease digestion following pulsed-field gel electrophoresis (S1-PFGE) analysis. Further, extrachromosomal elements in additional 15 Chinese epidemic strains of Leptospira, comprising L. borgpetersenii, L. weilii, and L. interrogans, were successfully separated and identified, independent of genome sequence data. Southern blot hybridization with extrachromosomal element-specific probes, designated as lcp1, lcp2 and lcp3-rep, further confirmed their occurrences as extrachromosomal elements. In total, 24 plasmids were detected in 13 out of 15 tested strains, among which 11 can hybridize with the lcp1-rep probe and 11 with the lcp2-rep probe, whereas two can hybridize with the lcp3-rep probe. None of them are likely to be species-specific. Blastp search of the lcp1, lcp2, and lcp3-rep genes with a nonredundant protein database of Leptospira species genomes showed that their homologous sequences are widely distributed among clades of pathogens but not non-pathogens or intermediates. These results suggest that the plasmids are widely distributed in Leptospira species, and further elucidation of their biological

  9. Distribution of Plasmids in Distinct Leptospira Pathogenic Species.

    PubMed

    Wang, Yanzhuo; Zhuang, Xuran; Zhong, Yi; Zhang, Cuicai; Zhang, Yan; Zeng, Lingbing; Zhu, Yongzhang; He, Ping; Dong, Ke; Pal, Utpal; Guo, Xiaokui; Qin, Jinhong

    2015-11-01

    Leptospirosis, caused by pathogenic Leptospira, is a worldwide zoonotic infection. The genus Leptospira includes at least 21 species clustered into three groups--pathogens, non-pathogens, and intermediates--based on 16S rRNA phylogeny. Research on Leptospira is difficult due to slow growth and poor transformability of the pathogens. Recent identification of extrachromosomal elements besides the two chromosomes in L. interrogans has provided new insight into genome complexity of the genus Leptospira. The large size, low copy number, and high similarity of the sequence of these extrachromosomal elements with the chromosomes present challenges in isolating and detecting them without careful genome assembly. In this study, two extrachromosomal elements were identified in L. borgpetersenii serovar Ballum strain 56604 through whole genome assembly combined with S1 nuclease digestion following pulsed-field gel electrophoresis (S1-PFGE) analysis. Further, extrachromosomal elements in additional 15 Chinese epidemic strains of Leptospira, comprising L. borgpetersenii, L. weilii, and L. interrogans, were successfully separated and identified, independent of genome sequence data. Southern blot hybridization with extrachromosomal element-specific probes, designated as lcp1, lcp2 and lcp3-rep, further confirmed their occurrences as extrachromosomal elements. In total, 24 plasmids were detected in 13 out of 15 tested strains, among which 11 can hybridize with the lcp1-rep probe and 11 with the lcp2-rep probe, whereas two can hybridize with the lcp3-rep probe. None of them are likely to be species-specific. Blastp search of the lcp1, lcp2, and lcp3-rep genes with a nonredundant protein database of Leptospira species genomes showed that their homologous sequences are widely distributed among clades of pathogens but not non-pathogens or intermediates. These results suggest that the plasmids are widely distributed in Leptospira species, and further elucidation of their biological

  10. Sequence homology between the subunits of two immunologically and functionally distinct types of fimbriae of Actinomyces spp.

    PubMed Central

    Yeung, M K; Cisar, J O

    1990-01-01

    Nucleotide sequencing of the type 1 fimbrial subunit gene of Actinomyces viscosus T14V revealed a consensus ribosome-binding site followed by an open reading frame of 1,599 nucleotides. The encoded protein of 533 amino acids (Mr = 56,899) was predominantly hydrophilic except for an amino-terminal signal peptide and a carboxy-terminal region identified as a potential membrane-spanning segment. Edman degradation of the cloned protein expressed in Escherichia coli and the type 1 fimbriae of A. viscosus T14V showed that both began with alanine at position 31 of the deduced amino acid sequence. The amino acid compositions of the cloned protein and fimbriae also were comparable and in close agreement with the composition of the deduced protein. The amino acid sequence of the A. viscosus T14V type 1 fimbrial subunit showed no significant global homology with various other proteins, including the pilins of gram-negative bacteria. However, 34% amino acid sequence identity was noted between the type 1 fimbrial subunit of strain T14V and the type 2 fimbrial subunit of Actinomyces naeslundii WVU45 (M. K. Yeung and J. O. Cisar, J. Bacteriol. 170:3803-3809, 1988). This homology included several different conserved sequences of up to eight identical amino acids that were distributed in both the amino- and carboxy-terminal thirds of each Actinomyces fimbrial subunit. These findings indicate that the different types of fimbriae on these gram-positive bacteria share a common ancestry. PMID:1970561

  11. Bordetella pertussis and Bordetella parapertussis: two immunologically distinct species.

    PubMed Central

    Khelef, N; Danve, B; Quentin-Millet, M J; Guiso, N

    1993-01-01

    Bordetella pertussis and Bordetella parapertussis are closely related species. Both are responsible for outbreaks of whooping cough in humans and produce similar virulence factors, with the exception of pertussis toxin, specific to B. pertussis. Current pertussis whole-cell vaccine will soon be replaced by acellular vaccines containing major adhesins (filamentous hemagglutinin and pertactin) and major toxin (pertussis toxin). All of these factors are antigens that stimulate a protective immune response in the murine respiratory model and in clinical assays. In the present study, we examined the protective efficacies of these factors, and that of adenylate cyclase-hemolysin, another B. pertussis toxin, against B. parapertussis infection in a murine respiratory model. As expected, pertussis toxin did not protect against B. parapertussis infection, since this bacterium did not express this protein, but the surprising result was that none of the other factors were protective against B. parapertussis infection. Furthermore, B. parapertussis adenylate cyclase-hemolysin, although it protected against B. parapertussis infection, did not protect against B. pertussis infection. Despite a high degree of homology between both B. pertussis and B. parapertussis species, no cross-protection was observed. Our results outline the fact that, as in other gram-negative bacteria, Bordetella surface proteins vary immunologically. Images PMID:8423077

  12. IgA binding lectins isolated from distinct Artocarpus species demonstrate differential specificity.

    PubMed

    Hashim, O H; Ng, C L; Gendeh, S; Nik Jaafar, M I

    1991-01-01

    The discovery of jacalin, a group of lectins from jackfruit seeds (Artocarpus heterophyllus), has attracted considerable attention due to its numerous interesting immunological properties as well as its usefulness in the isolation of various serum proteins. We have further identified a similar lectin from the seeds of Champedak (Artocarpus integer) which we refer to as lectin-C and performed comparative studies with two types of jacalin isolated from different batches of the Malaysian jackfruit seeds (jacalin-M1 and jacalin-M2). The three purified lectins demonstrated equivalent apparent Mr of about 52,500, each of which comprised of a combination of two types of non-covalently-linked subunits with apparent Mr of approximately 13,300 and 16,000. The lectins demonstrated equal haemagglutinating activity against human erythrocytes of blood groups A, B, AB and O. Our data also demonstrated that lectin-C, jacalin-M1 and jacalin-M2 are similar by selectively precipitating human serum IgA1 and colostral sIgA but not IgA2, IgD, IgG and IgM. When immunoelectrophoresis was performed on normal human sera and reacted with the lectins, single precipitin arcs corresponding to IgA immunoprecipitates were detected with lectin-C and jacalin-MI. Jacalin-M2, however, exhibited two closely associated precipitin arcs. The binding of these lectins with IgA was pronouncedly inhibited in the presence of p-nitrophenyl-beta-D-galactopyranoside, 1-o-methyl-alpha-D-galactopyranoside, D-melibiose, N-acetyl-D-galactosamine and D-galactose. The data therefore provide evidence on the differential specificity of IgA binding lectins isolated from seeds of similar as well as distinct Artocarpus species.

  13. Fast Photochemistry of Prototypical Phytochromes—A Species vs. Subunit Specific Comparison

    PubMed Central

    Ihalainen, Janne A.; Takala, Heikki; Lehtivuori, Heli

    2015-01-01

    Phytochromes are multi-domain red light photosensor proteins, which convert red light photons to biological activity utilizing the multitude of structural and chemical reactions. The steady increase in structural information obtained from various bacteriophytochromes has increased understanding about the functional mechanism of the photochemical processes of the phytochromes. Furthermore, a number of spectroscopic studies have revealed kinetic information about the light-induced reactions. The spectroscopic changes are, however, challenging to connect with the structural changes of the chromophore and the protein environment, as the excited state properties of the chromophores are very sensitive to the small structural and chemical changes of their environment. In this article, we concentrate on the results of ultra-fast spectroscopic experiments which reveal information about the important initial steps of the photoreactions of the phytochromes. We survey the excited state properties obtained during the last few decades. The differences in kinetics between different research laboratories are traditionally related to the differences of the studied species. However, we notice that the variation in the excited state properties depends on the subunit composition of the protein as well. This observation illustrates a feedback mechanism from the other domains to the chromophore. We propose that two feedback routes exist in phytochromes between the chromophore and the remotely located effector domain. The well-known connection between the subunits is the so-called tongue region, which changes its secondary structure while changing the light-activated state of the system. The other feedback route which we suggest is less obvious, it is made up of several water molecules ranging from the dimer interface to the vicinity of the chromophore, allowing even proton transfer reactions nearby the chromophore. PMID:26779488

  14. Effects of Salt Stress on Three Ecologically Distinct Plantago Species

    PubMed Central

    Al Hassan, Mohamad; Pacurar, Andrea; López-Gresa, María P.; Donat-Torres, María P.; Llinares, Josep V.; Boscaiu, Monica; Vicente, Oscar

    2016-01-01

    Comparative studies on the responses to salt stress of taxonomically related taxa should help to elucidate relevant mechanisms of stress tolerance in plants. We have applied this strategy to three Plantago species adapted to different natural habitats, P. crassifolia and P. coronopus–both halophytes–and P. major, considered as salt-sensitive since it is never found in natural saline habitats. Growth inhibition measurements in controlled salt treatments indicated, however, that P. major is quite resistant to salt stress, although less than its halophytic congeners. The contents of monovalent ions and specific osmolytes were determined in plant leaves after four-week salt treatments. Salt-treated plants of the three taxa accumulated Na+ and Cl- in response to increasing external NaCl concentrations, to a lesser extent in P. major than in the halophytes; the latter species also showed higher ion contents in the non-stressed plants. In the halophytes, K+ concentration decreased at moderate salinity levels, to increase again under high salt conditions, whereas in P. major K+ contents were reduced only above 400 mM NaCl. Sorbitol contents augmented in all plants, roughly in parallel with increasing salinity, but the relative increments and the absolute values reached did not differ much in the three taxa. On the contrary, a strong (relative) accumulation of proline in response to high salt concentrations (600–800 mM NaCl) was observed in the halophytes, but not in P. major. These results indicate that the responses to salt stress triggered specifically in the halophytes, and therefore the most relevant for tolerance in the genus Plantago are: a higher efficiency in the transport of toxic ions to the leaves, the capacity to use inorganic ions as osmotica, even under low salinity conditions, and the activation, in response to very high salt concentrations, of proline accumulation and K+ transport to the leaves of the plants. PMID:27490924

  15. Effects of Salt Stress on Three Ecologically Distinct Plantago Species.

    PubMed

    Al Hassan, Mohamad; Pacurar, Andrea; López-Gresa, María P; Donat-Torres, María P; Llinares, Josep V; Boscaiu, Monica; Vicente, Oscar

    2016-01-01

    Comparative studies on the responses to salt stress of taxonomically related taxa should help to elucidate relevant mechanisms of stress tolerance in plants. We have applied this strategy to three Plantago species adapted to different natural habitats, P. crassifolia and P. coronopus-both halophytes-and P. major, considered as salt-sensitive since it is never found in natural saline habitats. Growth inhibition measurements in controlled salt treatments indicated, however, that P. major is quite resistant to salt stress, although less than its halophytic congeners. The contents of monovalent ions and specific osmolytes were determined in plant leaves after four-week salt treatments. Salt-treated plants of the three taxa accumulated Na+ and Cl- in response to increasing external NaCl concentrations, to a lesser extent in P. major than in the halophytes; the latter species also showed higher ion contents in the non-stressed plants. In the halophytes, K+ concentration decreased at moderate salinity levels, to increase again under high salt conditions, whereas in P. major K+ contents were reduced only above 400 mM NaCl. Sorbitol contents augmented in all plants, roughly in parallel with increasing salinity, but the relative increments and the absolute values reached did not differ much in the three taxa. On the contrary, a strong (relative) accumulation of proline in response to high salt concentrations (600-800 mM NaCl) was observed in the halophytes, but not in P. major. These results indicate that the responses to salt stress triggered specifically in the halophytes, and therefore the most relevant for tolerance in the genus Plantago are: a higher efficiency in the transport of toxic ions to the leaves, the capacity to use inorganic ions as osmotica, even under low salinity conditions, and the activation, in response to very high salt concentrations, of proline accumulation and K+ transport to the leaves of the plants.

  16. Effects of Salt Stress on Three Ecologically Distinct Plantago Species.

    PubMed

    Al Hassan, Mohamad; Pacurar, Andrea; López-Gresa, María P; Donat-Torres, María P; Llinares, Josep V; Boscaiu, Monica; Vicente, Oscar

    2016-01-01

    Comparative studies on the responses to salt stress of taxonomically related taxa should help to elucidate relevant mechanisms of stress tolerance in plants. We have applied this strategy to three Plantago species adapted to different natural habitats, P. crassifolia and P. coronopus-both halophytes-and P. major, considered as salt-sensitive since it is never found in natural saline habitats. Growth inhibition measurements in controlled salt treatments indicated, however, that P. major is quite resistant to salt stress, although less than its halophytic congeners. The contents of monovalent ions and specific osmolytes were determined in plant leaves after four-week salt treatments. Salt-treated plants of the three taxa accumulated Na+ and Cl- in response to increasing external NaCl concentrations, to a lesser extent in P. major than in the halophytes; the latter species also showed higher ion contents in the non-stressed plants. In the halophytes, K+ concentration decreased at moderate salinity levels, to increase again under high salt conditions, whereas in P. major K+ contents were reduced only above 400 mM NaCl. Sorbitol contents augmented in all plants, roughly in parallel with increasing salinity, but the relative increments and the absolute values reached did not differ much in the three taxa. On the contrary, a strong (relative) accumulation of proline in response to high salt concentrations (600-800 mM NaCl) was observed in the halophytes, but not in P. major. These results indicate that the responses to salt stress triggered specifically in the halophytes, and therefore the most relevant for tolerance in the genus Plantago are: a higher efficiency in the transport of toxic ions to the leaves, the capacity to use inorganic ions as osmotica, even under low salinity conditions, and the activation, in response to very high salt concentrations, of proline accumulation and K+ transport to the leaves of the plants. PMID:27490924

  17. Two Drosophila DEG/ENaC channel subunits have distinct functions in gustatory neurons that activate male courtship.

    PubMed

    Liu, Tong; Starostina, Elena; Vijayan, Vinoy; Pikielny, Claudio W

    2012-08-22

    Trimeric sodium channels of the DEG/ENaC family have important roles in neurons, but the specific functions of different subunits present in heteromeric channels are poorly understood. We previously reported that the Drosophila DEG/ENaC subunit Ppk25 is essential in a small subset of gustatory neurons for activation of male courtship behavior, likely through detection of female pheromones. Here we show that, like mutations in ppk25, mutations in another Drosophila DEG/ENaC subunit gene, nope, specifically impair male courtship of females. nope regulatory sequences drive reporter gene expression in gustatory neurons of the labellum wings, and legs, including all gustatory neurons in which ppk25 function is required for male courtship of females. In addition, gustatory-specific knockdown of nope impairs male courtship. Further, the impaired courtship response of nope mutant males to females is rescued by targeted expression of nope in the subset of gustatory neurons in which ppk25 functions. However, nope and ppk25 have nonredundant functions, as targeted expression of ppk25 does not compensate for the lack of nope and vice versa. Moreover, Nope and Ppk25 form specific complexes when coexpressed in cultured cells. Together, these data indicate that the Nope and Ppk25 polypeptides have specific, nonredundant functions in a subset of gustatory neurons required for activation of male courtship in response to females, and suggest the hypothesis that Nope and Ppk25 function as subunits of a heteromeric DEG/ENaC channel required for gustatory detection of female pheromones. PMID:22915128

  18. Rice G-protein subunits qPE9-1 and RGB1 play distinct roles in abscisic acid responses and drought adaptation.

    PubMed

    Zhang, Dong-Ping; Zhou, Yong; Yin, Jian-Feng; Yan, Xue-Jiao; Lin, Sheng; Xu, Wei-Feng; Baluška, František; Wang, Yi-Ping; Xia, Yi-Ji; Liang, Guo-hua; Liang, Jian-Sheng

    2015-10-01

    Heterotrimeric GTP-binding protein (G-protein)-mediated abscisic acid (ABA) and drought-stress responses have been documented in numerous plant species. However, our understanding of the function of rice G-protein subunits in ABA signalling and drought tolerance is limited. In this study, the function of G-protein subunits in ABA response and drought resistance in rice plants was explored. It was found that the transcription level of qPE9-1 (rice Gγ subunit) gradually decreased with increasing ABA concentration and the lack of qPE9-1 showed an enhanced drought tolerance in rice plants. In contrast, mRNA levels of RGB1 (rice Gβ subunit) were significantly upregulated by ABA treatment and the lack of RGB1 led to reduced drought tolerance. Furthermore, the results suggested that qPE9-1 negatively regulates the ABA response by suppressing the expression of key transcription factors involved in ABA and stress responses, while RGB1 positively regulates ABA biosynthesis by upregulating NCED gene expression under both normal and drought stress conditions. Taken together, it is proposed that RGB1 is a positive regulator of the ABA response and drought adaption in rice plants, whereas qPE9-1 is modulated by RGB1 and functions as a negative regulator in the ABA-dependent drought-stress responses.

  19. Rice G-protein subunits qPE9-1 and RGB1 play distinct roles in abscisic acid responses and drought adaptation.

    PubMed

    Zhang, Dong-Ping; Zhou, Yong; Yin, Jian-Feng; Yan, Xue-Jiao; Lin, Sheng; Xu, Wei-Feng; Baluška, František; Wang, Yi-Ping; Xia, Yi-Ji; Liang, Guo-hua; Liang, Jian-Sheng

    2015-10-01

    Heterotrimeric GTP-binding protein (G-protein)-mediated abscisic acid (ABA) and drought-stress responses have been documented in numerous plant species. However, our understanding of the function of rice G-protein subunits in ABA signalling and drought tolerance is limited. In this study, the function of G-protein subunits in ABA response and drought resistance in rice plants was explored. It was found that the transcription level of qPE9-1 (rice Gγ subunit) gradually decreased with increasing ABA concentration and the lack of qPE9-1 showed an enhanced drought tolerance in rice plants. In contrast, mRNA levels of RGB1 (rice Gβ subunit) were significantly upregulated by ABA treatment and the lack of RGB1 led to reduced drought tolerance. Furthermore, the results suggested that qPE9-1 negatively regulates the ABA response by suppressing the expression of key transcription factors involved in ABA and stress responses, while RGB1 positively regulates ABA biosynthesis by upregulating NCED gene expression under both normal and drought stress conditions. Taken together, it is proposed that RGB1 is a positive regulator of the ABA response and drought adaption in rice plants, whereas qPE9-1 is modulated by RGB1 and functions as a negative regulator in the ABA-dependent drought-stress responses. PMID:26175353

  20. Changes in the Lung Microbiome following Lung Transplantation Include the Emergence of Two Distinct Pseudomonas Species with Distinct Clinical Associations

    PubMed Central

    Dickson, Robert P.; Erb-Downward, John R.; Freeman, Christine M.; Walker, Natalie; Scales, Brittan S.; Beck, James M.; Martinez, Fernando J.; Curtis, Jeffrey L.; Lama, Vibha N.; Huffnagle, Gary B.

    2014-01-01

    Background Multiple independent culture-based studies have identified the presence of Pseudomonas aeruginosa in respiratory samples as a positive risk factor for bronchiolitis obliterans syndrome (BOS). Yet, culture-independent microbiological techniques have identified a negative association between Pseudomonas species and BOS. Our objective was to investigate whether there may be a unifying explanation for these apparently dichotomous results. Methods We performed bronchoscopies with bronchoalveolar lavage (BAL) on lung transplant recipients (46 procedures in 33 patients) and 26 non-transplant control subjects. We analyzed bacterial communities in the BAL fluid using qPCR and pyrosequencing of 16S rRNA gene amplicons and compared the culture-independent data with the clinical metadata and culture results from these subjects. Findings Route of bronchoscopy (via nose or via mouth) was not associated with changes in BAL microbiota (p = 0.90). Among the subjects with positive Pseudomonas bacterial culture, P. aeruginosa was also identified by culture-independent methods. In contrast, a distinct Pseudomonas species, P. fluorescens, was often identified in asymptomatic transplant subjects by pyrosequencing but not detected via standard bacterial culture. The subject populations harboring these two distinct pseudomonads differed significantly with respect to associated symptoms, BAL neutrophilia, bacterial DNA burden and microbial diversity. Despite notable differences in culturability, a global database search of UM Hospital Clinical Microbiology Laboratory records indicated that P. fluorescens is commonly isolated from respiratory specimens. Interpretation We have reported for the first time that two prominent and distinct Pseudomonas species (P. fluorescens and P. aeruginosa) exist within the post-transplant lung microbiome, each with unique genomic and microbiologic features and widely divergent clinical associations, including presence during acute infection

  1. Two Distinct Mechanisms of Inactivation of the Class Ic Ribonucleotide Reductase from Chlamydia trachomatis by Hydroxyurea: Implications for the Protein Gating of Inter-subunit Electron Transfer†

    PubMed Central

    Jiang, Wei; Xie, Jiajia; Varano, Paul T.; Krebs, Carsten; Bollinger, J. Martin

    2010-01-01

    Catalysis by a class I ribonucleotide reductase (RNR) begins when a cysteine (C) residue in the α2 subunit is oxidized to a thiyl radical (C•) by a cofactor ~ 35 Å away in the β2 subunit. In a class Ia or Ib RNR, a stable tyrosyl radical (Y•) is the C oxidant, whereas a MnIV/FeIII cluster serves this function in the class Ic enzyme from Chlamydia trachomatis (Ct). It is thought that, in either case, a chain of Y residues spanning the two subunits mediates C oxidation by forming transient "pathway" Y•s in a multi-step electron transfer (ET) process that is "gated" by the protein so as to occur only in the ready holoenzyme complex. The drug hydroxyurea (HU) inactivates both Ia/b and Ic β2s by reducing their C oxidants. Reduction of the stable cofactor Y• (Y122•) in Escherichia coli class Ia β2 is faster in the presence of α2 and substrate (CDP), leading to speculation that HU might intercept a transient ET-pathway Y• under these turnover conditions. Here we show that this mechanism is one of two that are operant in HU inactivation of the Ct enzyme. HU reacts with the MnIV/FeIII cofactor to give two distinct products: the previously described homogeneous MnIII/FeIII-β2 complex forms only under turnover conditions (in the presence α2 and the substrate), and a distinct, diamagnetic Mn/Fe cluster forms ~900-fold less rapidly as a second phase in the reaction under turnover conditions and as the sole outcome in the reaction of MnIV/FeIII-β2 only. Formation of MnIII/FeIII-β2 also requires (i) either Y338, its subunit-interfacial ET pathway residue, or Y222, the surface residue that relays the “extra electron” to the MnIV/FeIV intermediate during activation of β2 but is not part of the catalytic ET pathway, and (ii) W51, the cofactor-proximal residue required for efficient ET between either Y222 or Y338 and the cofactor. The combined requirements for the catalytic subunit, the substrate, and, most importantly, a functional surface

  2. A Streamlined System for Species Diagnosis in Caenorhabditis (Nematoda: Rhabditidae) with Name Designations for 15 Distinct Biological Species

    PubMed Central

    Félix, Marie-Anne; Braendle, Christian; Cutter, Asher D.

    2014-01-01

    The rapid pace of species discovery outstrips the rate of species description in many taxa. This problem is especially acute for Caenorhabditis nematodes, where the naming of distinct species would greatly improve their visibility and usage for biological research, given the thousands of scientists studying Caenorhabditis. Species description and naming has been hampered in Caenorhabditis, in part due to the presence of morphologically cryptic species despite complete biological reproductive isolation and often enormous molecular divergence. With the aim of expediting species designations, here we propose and apply a revised framework for species diagnosis and description in this group. Our solution prioritizes reproductive isolation over traditional morphological characters as the key feature in delineating and diagnosing new species, reflecting both practical considerations and conceptual justifications. DNA sequence divergence criteria help prioritize crosses for establishing patterns of reproductive isolation among the many species of Caenorhabditis known to science, such as with the ribosomal internal transcribed spacer-2 (ITS2) DNA barcode. By adopting this approach, we provide new species name designations for 15 distinct biological species, thus increasing the number of named Caenorhabditis species in laboratory culture by nearly 3-fold. We anticipate that the improved accessibility of these species to the research community will expand the opportunities for study and accelerate our understanding of diverse biological phenomena. PMID:24727800

  3. A distinct three-helix centipede toxin SSD609 inhibits I(ks) channels by interacting with the KCNE1 auxiliary subunit.

    PubMed

    Sun, Peibei; Wu, Fangming; Wen, Ming; Yang, Xingwang; Wang, Chenyang; Li, Yiming; He, Shufang; Zhang, Longhua; Zhang, Yun; Tian, Changlin

    2015-08-26

    KCNE1 is a single-span transmembrane auxiliary protein that modulates the voltage-gated potassium channel KCNQ1. The KCNQ1/KCNE1 complex in cardiomyocytes exhibited slow activated potassium (I(ks)) currents. Recently, a novel 47-residue polypeptide toxin SSD609 was purified from Scolopendra subspinipes dehaani venom and showed I(ks) current inhibition. Here, chemically synthesized SSD609 was shown to exert I(ks) inhibition in extracted guinea pig cardiomyocytes and KCNQ1/KCNE1 current attenuation in CHO cells. The K(+) current attenuation of SSD609 showed decent selectivity among different auxiliary subunits. Solution nuclear magnetic resonance analysis of SSD609 revealed a distinctive three-helix conformation that was stabilized by a new disulfide bonding pattern as well as segregated surface charge distribution. Structure-activity studies demonstrated that negatively charged Glu19 in the amphipathic extracellular helix of KCNE1 was the key residue that interacted with SSD609. The distinctive three-helix centipede toxin SSD609 is known to be the first polypeptide toxin acting on channel auxiliary subunit KCNE1, which suggests a new type of pharmacological regulation for ion channels in cardiomyocytes.

  4. A distinct three-helix centipede toxin SSD609 inhibits I(ks) channels by interacting with the KCNE1 auxiliary subunit.

    PubMed

    Sun, Peibei; Wu, Fangming; Wen, Ming; Yang, Xingwang; Wang, Chenyang; Li, Yiming; He, Shufang; Zhang, Longhua; Zhang, Yun; Tian, Changlin

    2015-01-01

    KCNE1 is a single-span transmembrane auxiliary protein that modulates the voltage-gated potassium channel KCNQ1. The KCNQ1/KCNE1 complex in cardiomyocytes exhibited slow activated potassium (I(ks)) currents. Recently, a novel 47-residue polypeptide toxin SSD609 was purified from Scolopendra subspinipes dehaani venom and showed I(ks) current inhibition. Here, chemically synthesized SSD609 was shown to exert I(ks) inhibition in extracted guinea pig cardiomyocytes and KCNQ1/KCNE1 current attenuation in CHO cells. The K(+) current attenuation of SSD609 showed decent selectivity among different auxiliary subunits. Solution nuclear magnetic resonance analysis of SSD609 revealed a distinctive three-helix conformation that was stabilized by a new disulfide bonding pattern as well as segregated surface charge distribution. Structure-activity studies demonstrated that negatively charged Glu19 in the amphipathic extracellular helix of KCNE1 was the key residue that interacted with SSD609. The distinctive three-helix centipede toxin SSD609 is known to be the first polypeptide toxin acting on channel auxiliary subunit KCNE1, which suggests a new type of pharmacological regulation for ion channels in cardiomyocytes. PMID:26307551

  5. Gill (Na+,K+)-ATPase in diadromous, freshwater palaemonid shrimps: species-specific kinetic characteristics and alpha-subunit expression.

    PubMed

    Santos, L C F; Belli, N M; Augusto, A; Masui, D C; Leone, F A; McNamara, J C; Furriel, R P M

    2007-09-01

    To better comprehend physiological adaptation to dilute media and the molecular mechanisms underlying ammonia excretion in palaemonid shrimps, we characterized the (Na+,K+)-ATPase from Macrobrachium amazonicum gills, disclosing high- (K(0.5) = 4.2+/-0.2 micromol L(-1); V = 33.9+/-1.9 U mg(-1)) and low-affinity (K(0.5) = 0.144+/-0.010 mmol L(-1); V = 232.9+/-15.3 U mg(-1)) ATP hydrolyzing sites. Stimulation by Na+ (K(0.5) = 5.5+/-0.3 mmol L(-1); V = 275.1+/-15.1 U mg(-1)), Mg2+ (K(0.5) = 0.79+/-0.06 mmol L(-1); V = 261.9+/-18.3 U mg(-1)), K+ (K(M) = 0.88+/-0.04 mmol L(-1); V = 271.8+/-10.9 U mg(-1)) and NH4(+) (K(M) = 5.0+/-0.2 mmol L(-1); V = 385.9+/-15.8 U mg(-1)) obeys single saturation curves, activity being stimulated synergistically by NH4(+) and K+. There is a single K+ binding site, NH4(+) binding to a second, exclusive site, stimulating activity by 33%, modulating K+ affinity. (Na+,K+)-ATPase activity constitutes approximately 80% of total ATPase activity (K(Iouabain) = 147.5+/-8.9 micromol L(-1)); Na+-, K+-, Ca2+-, V- and F(o)F(1)-ATPases are also present. M. amazonicum microsomal fractions possess approximately 2-fold less (Na+,K+)-ATPase alpha-subunit than M. olfersi, consistent with a 2.6-fold lower specific activity. These differences in (Na+, K+)-ATPase stimulation by ATP and ions, and specific activities of other ATPases, suggest the presence of distinct biochemical adaptations to life in fresh water in these related species.

  6. Culturing-based Temperature Calibration of a Genetically Distinct, Alkenone-producing Haptophyte Species isolated from Lake George, ND

    NASA Astrophysics Data System (ADS)

    Zheng, Y.; Andersen, R. A.; Huang, Y.; Amaral-Zettler, L. A.

    2014-12-01

    Lacustrine alkenones are rapidly becoming an important tool for continental paleoclimate reconstructions. However, DNA sequencing of 18S ribosomal RNA marker genes has uncovered multiple species of haptophytes in different lakes. To date, there are only two isolated lacustrine species Chrysotila lamellosa and Isochrysis galbana available for culture studies. In our study, we report the isolation of a new haptophyte species from Lake George (LG) that, based on analyses of partial large subunit rRNA gene sequences, is genetically distinct from both Chrysotila lamellosa and Isochrysis galbana. We examined alkenone unsaturation index UK37 values for the LG species at 4°C, 10°C, 15°C, 20°C and 25°C as a function of temperature in a culture experiment. The temperature sensitivity of the new species was significantly higher than previously cultured Isochrysis galbana and Chrysotila lamellosa strains, with a slope that was 25 to 100 % higher. We found that the best linear relationship was obtained when two double-bond alkenones were excluded from the calibration (we developed an index termed UK''37 = [C37:4] / [C37:3+C37:4]). In particular, UK''37 is more linear to the growth temperature than UK37 at low (4-10°C) and high (20-25°C) temperature ranges. Our experiments show that both UK37 and UK''37 of this new alkenone-produced species is strongly controlled by culture temperature and can be used for paleoclimate reconstruction. However, we recommend the use of UK''37 index to reconstruct temperature if the haptophyte's growing environment falls within temperature extremes (4-10°C and 20-25°C). This newly cultivated species broadens our ability of applying lacustrine haptophyte calibrations to continental paleothermometry.

  7. Distinctive interactions of the Arabidopsis homolog of the 30 kD subunit of the cleavage and polyadenylation specificity factor (AtCPSF30) with other polyadenylation factor subunits

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: The Arabidopsis ortholog of the 30 kD subunit of the mammalian Cleavage and Polyadenylation Specificity Factor (AtCPSF30) is an RNA-binding endonuclease that is associated with other Arabidopsis CPSF subunits (orthologs of the 160, 100, and 73 kD subunits of CPSF). In order to better u...

  8. Efficient distinction of invasive aquatic plant species from non-invasive related species using DNA barcoding.

    PubMed

    Ghahramanzadeh, R; Esselink, G; Kodde, L P; Duistermaat, H; van Valkenburg, J L C H; Marashi, S H; Smulders, M J M; van de Wiel, C C M

    2013-01-01

    Biological invasions are regarded as threats to global biodiversity. Among invasive aliens, a number of plant species belonging to the genera Myriophyllum, Ludwigia and Cabomba, and to the Hydrocharitaceae family pose a particular ecological threat to water bodies. Therefore, one would try to prevent them from entering a country. However, many related species are commercially traded, and distinguishing invasive from non-invasive species based on morphology alone is often difficult for plants in a vegetative stage. In this regard, DNA barcoding could become a good alternative. In this study, 242 samples belonging to 26 species from 10 genera of aquatic plants were assessed using the chloroplast loci trnH-psbA, matK and rbcL. Despite testing a large number of primer sets and several PCR protocols, the matK locus could not be amplified or sequenced reliably and therefore was left out of the analysis. Using the other two loci, eight invasive species could be distinguished from their respective related species, a ninth one failed to produce sequences of sufficient quality. Based on the criteria of universal application, high sequence divergence and level of species discrimination, the trnH-psbA noncoding spacer was the best performing barcode in the aquatic plant species studied. Thus, DNA barcoding may be helpful with enforcing a ban on trade of such invasive species, such as is already in place in the Netherlands. This will become even more so once DNA barcoding would be turned into machinery routinely operable by a nonspecialist in botany and molecular genetics.

  9. Targeted deletion of one or two copies of the G protein β subunit Gβ5 gene has distinct effects on body weight and behavior in mice

    PubMed Central

    Wang, Qiang; Levay, Konstantin; Chanturiya, Tatyana; Dvoriantchikova, Galina; Anderson, Karen L.; Bianco, Suzy D. C.; Ueta, Cintia B.; Molano, R. Damaris; Pileggi, Antonello; Gurevich, Eugenia V.; Gavrilova, Oksana; Slepak, Vladlen Z.

    2011-01-01

    We investigated the physiological role of Gβ5, a unique G protein β subunit that dimerizes with regulators of G protein signaling (RGS) proteins of the R7 family instead of Gγ. Gβ5 is essential for stability of these complexes, so that its knockout (KO)causes degradation of the entire Gβ5-R7 family. We report that the Gβ5-KO mice remain leaner than the wild type (WT) throughout their lifetime and are resistant to a high-fat diet. They have a 5-fold increase in locomotor activity, increased thermogenesis, and lower serum insulin, all of which correlate with a higher level of secreted epinephrine. Heterozygous (HET) mice are 2-fold more active than WT mice. Surprisingly, with respect to body weight, the HET mice display a phenotype opposite to that of the KO mice: by the age of 6 mo, they are ≥15% heavier than the WT and have increased adiposity, insulin resistance, and liver steatosis. These changes occur in HET mice fed a normal diet and without apparent hyperphagia, mimicking basic characteristics of human metabolic syndrome. We conclude that even a partial reduction in Gβ5-R7 level can perturb normal animal metabolism and behavior. Our data on Gβ5 haploinsufficient mice may explain earlier observations of genetic linkage between R7 family mutations and obesity in humans.—Wang, Q., Levay, K., Chanturiya, T., Dvoriantchikova, G., Anderson, K. L., Bianco, S. D. C., Ueta, C. B., Molano, R. D., Pileggi, A., Gurevich, E. V., Gavrilova, O., and Slepak, V. Z. Targeted deletion of one or two copies of the G protein β subunit Gβ5 gene has distinct effects on body weight and behavior in mice. PMID:21804131

  10. Differential localization of ion transporters suggests distinct cellular mechanisms for calcification and photosynthesis between two coral species.

    PubMed

    Barott, Katie L; Perez, Sidney O; Linsmayer, Lauren B; Tresguerres, Martin

    2015-08-01

    Ion transport is fundamental for multiple physiological processes, including but not limited to pH regulation, calcification, and photosynthesis. Here, we investigated ion-transporting processes in tissues from the corals Acropora yongei and Stylophora pistillata, representatives of the complex and robust clades that diverged over 250 million years ago. Antibodies against complex IV revealed that mitochondria, an essential source of ATP for energetically costly ion transporters, were abundant throughout the tissues of A. yongei. Additionally, transmission electron microscopy revealed septate junctions in all cell layers of A. yongei, as previously reported for S. pistillata, as well as evidence for transcellular vesicular transport in calicoblastic cells. Antibodies against the alpha subunit of Na(+)/K(+)-ATPase (NKA) and plasma membrane Ca(2+)-ATPase (PMCA) immunolabeled cells in the calicoblastic epithelium of both species, suggesting conserved roles in calcification. However, NKA was abundant in the apical membrane of the oral epithelium in A. yongei but not S. pistillata, while PMCA was abundant in the gastroderm of S. pistillata but not A. yongei. These differences indicate that these two coral species utilize distinct pathways to deliver ions to the sites of calcification and photosynthesis. Finally, antibodies against mammalian sodium bicarbonate cotransporters (NBC; SLC4 family) resulted in strong immunostaining in the apical membrane of oral epithelial cells and in calicoblastic cells in A. yongei, a pattern identical to NKA. Characterization of ion transport mechanisms is an essential step toward understanding the cellular mechanisms of coral physiology and will help predict how different coral species respond to environmental stress.

  11. Genome-wide Evidence Reveals that African and Eurasian Golden Jackals Are Distinct Species.

    PubMed

    Koepfli, Klaus-Peter; Pollinger, John; Godinho, Raquel; Robinson, Jacqueline; Lea, Amanda; Hendricks, Sarah; Schweizer, Rena M; Thalmann, Olaf; Silva, Pedro; Fan, Zhenxin; Yurchenko, Andrey A; Dobrynin, Pavel; Makunin, Alexey; Cahill, James A; Shapiro, Beth; Álvares, Francisco; Brito, José C; Geffen, Eli; Leonard, Jennifer A; Helgen, Kristofer M; Johnson, Warren E; O'Brien, Stephen J; Van Valkenburgh, Blaire; Wayne, Robert K

    2015-08-17

    The golden jackal of Africa (Canis aureus) has long been considered a conspecific of jackals distributed throughout Eurasia, with the nearest source populations in the Middle East. However, two recent reports found that mitochondrial haplotypes of some African golden jackals aligned more closely to gray wolves (Canis lupus), which is surprising given the absence of gray wolves in Africa and the phenotypic divergence between the two species. Moreover, these results imply the existence of a previously unrecognized phylogenetically distinct species despite a long history of taxonomic work on African canids. To test the distinct-species hypothesis and understand the evolutionary history that would account for this puzzling result, we analyzed extensive genomic data including mitochondrial genome sequences, sequences from 20 autosomal loci (17 introns and 3 exon segments), microsatellite loci, X- and Y-linked zinc-finger protein gene (ZFX and ZFY) sequences, and whole-genome nuclear sequences in African and Eurasian golden jackals and gray wolves. Our results provide consistent and robust evidence that populations of golden jackals from Africa and Eurasia represent distinct monophyletic lineages separated for more than one million years, sufficient to merit formal recognition as different species: C. anthus (African golden wolf) and C. aureus (Eurasian golden jackal). Using morphologic data, we demonstrate a striking morphologic similarity between East African and Eurasian golden jackals, suggesting parallelism, which may have misled taxonomists and likely reflects uniquely intense interspecific competition in the East African carnivore guild. Our study shows how ecology can confound taxonomy if interspecific competition constrains size diversification.

  12. Multi-predator effects produced by functionally distinct species vary with prey density.

    PubMed

    Werling, Ben P; Lowenstein, David M; Straub, Cory S; Gratton, Claudio

    2012-01-01

    Determining when multiple predator species provide better pest suppression than single species is a key step towards developing ecologically-informed biological control strategies. Theory and experiments predict that resource partitioning among functionally different predator species can strengthen prey suppression, because as a group they can access more prey types than functionally redundant predators. However, this prediction assumes that competition limits predation by functionally similar predators. Differences in prey density can alter the strength of competition, suggesting that prey abundance may modulate the effect of combining functionally diverse species. The experiment documented here examined the potential for functional differences among predator species to promote suppression of an insect pest, the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae), at different prey densities. Predation was compared at two prey densities between microcosms that contained one predator species or two functionally distinct species: the lady beetle, Coleomegilla maculata De Geer (Coleoptera: Coccinellidae) that kills early L. decemlineata instars, and the soldier bug, Podisus maculiventris Say (Hemiptera: Pentatomidae) that kills late instars. The data show that combining these predators increased predation only when prey densities were low. This suggests that multiple predator species may only provide greater biological control than single species in systems where prey is limiting. PMID:22958369

  13. Chlamydia Infection Across Host Species Boundaries Promotes Distinct Sets of Transcribed Anti-Apoptotic Factors.

    PubMed

    Messinger, Joshua E; Nelton, Emmalin; Feeney, Colleen; Gondek, David C

    2015-01-01

    Chlamydiae, obligate intracellular bacteria, cause significant human and veterinary associated diseases. Having emerged an estimated 700-million years ago, these bacteria have twice adapted to humans as a host species, causing sexually transmitted infection (C. trachomatis) and respiratory associated disease (C. pneumoniae). The principle mechanism of host cell defense against these intracellular bacteria is the induction of cell death via apoptosis. However, in the "arms race" of co-evolution, Chlamydiae have developed mechanisms to promote cell viability and inhibit cell death. Herein we examine the impact of Chlamydiae infection across multiple host species on transcription of anti-apoptotic genes. We found mostly distinct patterns of gene expression (Mcl1 and cIAPs) elicited by each pathogen-host pair indicating Chlamydiae infection across host species boundaries does not induce a universally shared host response. Understanding species specific host-pathogen interactions is paramount to deciphering how potential pathogens become emerging diseases. PMID:26779446

  14. Chlamydia Infection Across Host Species Boundaries Promotes Distinct Sets of Transcribed Anti-Apoptotic Factors

    PubMed Central

    Messinger, Joshua E.; Nelton, Emmalin; Feeney, Colleen; Gondek, David C.

    2015-01-01

    Chlamydiae, obligate intracellular bacteria, cause significant human and veterinary associated diseases. Having emerged an estimated 700-million years ago, these bacteria have twice adapted to humans as a host species, causing sexually transmitted infection (C. trachomatis) and respiratory associated disease (C. pneumoniae). The principle mechanism of host cell defense against these intracellular bacteria is the induction of cell death via apoptosis. However, in the “arms race” of co-evolution, Chlamydiae have developed mechanisms to promote cell viability and inhibit cell death. Herein we examine the impact of Chlamydiae infection across multiple host species on transcription of anti-apoptotic genes. We found mostly distinct patterns of gene expression (Mcl1 and cIAPs) elicited by each pathogen-host pair indicating Chlamydiae infection across host species boundaries does not induce a universally shared host response. Understanding species specific host-pathogen interactions is paramount to deciphering how potential pathogens become emerging diseases. PMID:26779446

  15. Identification of distinct physiochemical properties of toxic prefibrillar species formed by A{beta} peptide variants

    SciTech Connect

    Goeransson, Anna-Lena; Nilsson, K. Peter R.; Kagedal, Katarina; Brorsson, Ann-Christin

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer Identification of toxic prefibrillar A{beta} species. Black-Right-Pointing-Pointer Fluorescence measurements using a combined set of fluorophores. Black-Right-Pointing-Pointer Morphology studies using transmission electron microscopy. -- Abstract: The formation of amyloid-{beta} peptide (A{beta}) aggregates at an early stage during the self-assembly process is an important factor in the development of Alzheimer's disease. The toxic effect is believed to be exerted by prefibrillar species of A{beta}. It is therefore important to identify which prefibrillar species are toxic and characterize their distinct properties. In the present study, we investigated the in vitro aggregation behavior of A{beta}-derived peptides possessing different levels of neurotoxic activity, using fluorescence spectroscopy in combination with transmission electron microscopy. The toxicity of various A{beta} aggregates was assessed by using cultures of human neuroblastoma cells. Through combined use of the fluorescence probe 8-anilino-1-napthalenesulfonate (ANS) and the novel luminescent probe pentamer formyl thiophene acetic acid (p-FTAA), we were able to identify those A{beta} peptide-derived prefibrillar species which exhibited cellular toxicity. In particular, species, which formed early during the aggregation process and showed strong p-FTAA and ANS fluorescence, were the species that possessed toxic activities. Moreover, by manipulating the aggregation conditions, it was possible to change the capacity of the A{beta} peptide to form nontoxic versus toxic species.

  16. Morphological Differences between Larvae of the Ciona intestinalis Species Complex: Hints for a Valid Taxonomic Definition of Distinct Species.

    PubMed

    Pennati, Roberta; Ficetola, Gentile Francesco; Brunetti, Riccardo; Caicci, Federico; Gasparini, Fabio; Griggio, Francesca; Sato, Atsuko; Stach, Thomas; Kaul-Strehlow, Sabrina; Gissi, Carmela; Manni, Lucia

    2015-01-01

    The cosmopolitan ascidian Ciona intestinalis is the most common model species of Tunicata, the sister-group of Vertebrata, and widely used in developmental biology, genomics and evolutionary studies. Recently, molecular studies suggested the presence of cryptic species hidden within the C. intestinalis species, namely C. intestinalis type A and type B. So far, no substantial morphological differences have been identified between individuals belonging to the two types. Here we present morphometric, immunohistochemical, and histological analyses, as well as 3-D reconstructions, of late larvae obtained by cross-fertilization experiments of molecularly determined type A and type B adults, sampled in different seasons and in four different localities. Our data point to quantitative and qualitative differences in the trunk shape of larvae belonging to the two types. In particular, type B larvae exhibit a longer pre-oral lobe, longer and relatively narrower total body length, and a shorter ocellus-tail distance than type A larvae. All these differences were found to be statistically significant in a Discriminant Analysis. Depending on the number of analyzed parameters, the obtained discriminant function was able to correctly classify > 93% of the larvae, with the remaining misclassified larvae attributable to the existence of intra-type seasonal variability. No larval differences were observed at the level of histology and immunohistochemical localization of peripheral sensory neurons. We conclude that type A and type B are two distinct species that can be distinguished on the basis of larval morphology and molecular data. Since the identified larval differences appear to be valid diagnostic characters, we suggest to raise both types to the rank of species and to assign them distinct names.

  17. A new rhabditoid nematode species in Asian sciurids, distinct from Strongyloides robustus in North American sciurids.

    PubMed

    Sato, Hiroshi; Torii, Harumi; Une, Yumi; Ooi, Hong-Kean

    2007-12-01

    Strongyloides callosciureus n. sp. (Nematoda: Rhabditoidea), from Asian sciurids, is described based on morphology, morphometry, and the small and large subunit (SSU/LSU) ribosomal RNA gene (rDNA) sequences. This new species was collected from Pallas's squirrels (Callosciurus erythraeus) in the central part of mainland Japan (Honshu), which were originally introduced from Taiwan some decades ago, and plantain squirrels (Callosciurus notatus) imported from Malaysia as personal pets. For comparison, Strongyloides robustus Chandler, 1942 was collected from American red squirrels (Tamiasciurus hudsonicus) and southern flying squirrels (Glaucomys volans) imported from the United States as personal pets. The parasitic females found in North American and Asian sciurids shared some key morphological features such as the ovary running spirally around the gut, and the shapes of the stoma in the apical view and the tail. However, morphometric features of parasitic females in North American and Asian sciurids differed significantly from each other; the former was larger than the latter, and the relative position of the vulva to the whole body length from the mouth was different. The SSU/LSU rDNA sequences supported the division of sciurid Strongyloides isolates by geographical distribution of the host and morphological features, leading us to propose the erection of new species. PMID:18314696

  18. Genome-wide Evidence Reveals that African and Eurasian Golden Jackals Are Distinct Species.

    PubMed

    Koepfli, Klaus-Peter; Pollinger, John; Godinho, Raquel; Robinson, Jacqueline; Lea, Amanda; Hendricks, Sarah; Schweizer, Rena M; Thalmann, Olaf; Silva, Pedro; Fan, Zhenxin; Yurchenko, Andrey A; Dobrynin, Pavel; Makunin, Alexey; Cahill, James A; Shapiro, Beth; Álvares, Francisco; Brito, José C; Geffen, Eli; Leonard, Jennifer A; Helgen, Kristofer M; Johnson, Warren E; O'Brien, Stephen J; Van Valkenburgh, Blaire; Wayne, Robert K

    2015-08-17

    The golden jackal of Africa (Canis aureus) has long been considered a conspecific of jackals distributed throughout Eurasia, with the nearest source populations in the Middle East. However, two recent reports found that mitochondrial haplotypes of some African golden jackals aligned more closely to gray wolves (Canis lupus), which is surprising given the absence of gray wolves in Africa and the phenotypic divergence between the two species. Moreover, these results imply the existence of a previously unrecognized phylogenetically distinct species despite a long history of taxonomic work on African canids. To test the distinct-species hypothesis and understand the evolutionary history that would account for this puzzling result, we analyzed extensive genomic data including mitochondrial genome sequences, sequences from 20 autosomal loci (17 introns and 3 exon segments), microsatellite loci, X- and Y-linked zinc-finger protein gene (ZFX and ZFY) sequences, and whole-genome nuclear sequences in African and Eurasian golden jackals and gray wolves. Our results provide consistent and robust evidence that populations of golden jackals from Africa and Eurasia represent distinct monophyletic lineages separated for more than one million years, sufficient to merit formal recognition as different species: C. anthus (African golden wolf) and C. aureus (Eurasian golden jackal). Using morphologic data, we demonstrate a striking morphologic similarity between East African and Eurasian golden jackals, suggesting parallelism, which may have misled taxonomists and likely reflects uniquely intense interspecific competition in the East African carnivore guild. Our study shows how ecology can confound taxonomy if interspecific competition constrains size diversification. PMID:26234211

  19. The cane or marine toad, Rhinella marina (Anura, Bufonidae): two genetically and morphologically distinct species.

    PubMed

    Acevedo, Aldemar A; Lampo, Margarita; Cipriani, Roberto

    2016-04-18

    Rhinella marina is a Neotropical toad that has been introduced widely worldwide. Its toxic effects to frog-eating predators threaten the native and domestic fauna of some regions where it has been introduced. Despite previous studies suggesting two genetically distinct cryptic species within R. marina, one east and one west of the Andes, its taxonomic status remained unresolved due to the absence of morphological complementary evidence. For the first time, data from two mitochondrial genes (ND3 and CR) and 23 morphometric landmarks are combined to evaluate the taxonomic status of this species. Our results support the hypothesis of two separate evolutionary lineages within R. marina and demonstrate that these lineages have significantly diverged in skull shape. We identified two distinct morphotypes, one eastern and one Andean western, with no overlapping morphospaces. The geographic pattern of genetic variation was consistent with a stable structured population with no evidence of recent demographic or geographic expansions. The concordance between the observed geographic patterns in morphometric and genic traits calls for the recognition of two species under R. marina name.

  20. Distinct Host Species Correlate with Anaplasma phagocytophilum ankA Gene Clusters▿†

    PubMed Central

    Scharf, Wiebke; Schauer, Sonja; Freyburger, Felix; Petrovec, Miroslav; Schaarschmidt-Kiener, Daniel; Liebisch, Gabriele; Runge, Martin; Ganter, Martin; Kehl, Alexandra; Dumler, J. Stephen; Garcia-Perez, Ana L.; Jensen, Jennifer; Fingerle, Volker; Meli, Marina L.; Ensser, Armin; Stuen, Snorre; von Loewenich, Friederike D.

    2011-01-01

    Anaplasma phagocytophilum is a Gram-negative, tick-transmitted, obligate intracellular bacterium that elicits acute febrile diseases in humans and domestic animals. In contrast to the United States, human granulocytic anaplasmosis seems to be a rare disease in Europe despite the initial recognition of A. phagocytophilum as the causative agent of tick-borne fever in European sheep and cattle. Considerable strain variation has been suggested to occur within this species, because isolates from humans and animals differed in their pathogenicity for heterologous hosts. In order to explain host preference and epidemiological diversity, molecular characterization of A. phagocytophilum strains has been undertaken. Most often the 16S rRNA gene was used, but it might be not informative enough to delineate distinct genotypes of A. phagocytophilum. Previously, we have shown that A. phagocytophilum strains infecting Ixodes ricinus ticks are highly diverse in their ankA genes. Therefore, we sequenced the 16S rRNA and ankA genes of 194 A. phagocytophilum strains from humans and several animal species. Whereas the phylogenetic analysis using 16S rRNA gene sequences was not meaningful, we showed that distinct host species correlate with A. phagocytophilum ankA gene clusters. PMID:21177886

  1. The cane or marine toad, Rhinella marina (Anura, Bufonidae): two genetically and morphologically distinct species.

    PubMed

    Acevedo, Aldemar A; Lampo, Margarita; Cipriani, Roberto

    2016-01-01

    Rhinella marina is a Neotropical toad that has been introduced widely worldwide. Its toxic effects to frog-eating predators threaten the native and domestic fauna of some regions where it has been introduced. Despite previous studies suggesting two genetically distinct cryptic species within R. marina, one east and one west of the Andes, its taxonomic status remained unresolved due to the absence of morphological complementary evidence. For the first time, data from two mitochondrial genes (ND3 and CR) and 23 morphometric landmarks are combined to evaluate the taxonomic status of this species. Our results support the hypothesis of two separate evolutionary lineages within R. marina and demonstrate that these lineages have significantly diverged in skull shape. We identified two distinct morphotypes, one eastern and one Andean western, with no overlapping morphospaces. The geographic pattern of genetic variation was consistent with a stable structured population with no evidence of recent demographic or geographic expansions. The concordance between the observed geographic patterns in morphometric and genic traits calls for the recognition of two species under R. marina name. PMID:27394759

  2. Two Rumex species from contrasting hydrological niches regulate flooding tolerance through distinct mechanisms.

    PubMed

    van Veen, Hans; Mustroph, Angelika; Barding, Gregory A; Vergeer-van Eijk, Marleen; Welschen-Evertman, Rob A M; Pedersen, Ole; Visser, Eric J W; Larive, Cynthia K; Pierik, Ronald; Bailey-Serres, Julia; Voesenek, Laurentius A C J; Sasidharan, Rashmi

    2013-11-01

    Global climate change has increased flooding events, which affect both natural vegetation dynamics and crop productivity. The flooded environment is lethal for most plant species because it restricts gas exchange and induces an energy and carbon crisis. Flooding survival strategies have been studied in Oryza sativa, a cultivated monocot. However, our understanding of plant adaptation to natural flood-prone environments remains scant, even though wild plants represent a valuable resource of tolerance mechanisms that could be used to generate stress-tolerant crops. Here we identify mechanisms that mediate the distinct flooding survival strategies of two related wild dicot species: Rumex palustris and Rumex acetosa. Whole transcriptome sequencing and metabolite profiling reveal flooding-induced metabolic reprogramming specific to R. acetosa. By contrast, R. palustris uses the early flooding signal ethylene to increase survival by regulating shade avoidance and photomorphogenesis genes to outgrow submergence and by priming submerged plants for future low oxygen stress. These results provide molecular resolution of flooding survival strategies of two species occupying distinct hydrological niches. Learning how these contrasting flood adaptive strategies evolved in nature will be instrumental for the development of stress-tolerant crop varieties that deliver enhanced yields in a changing climate.

  3. Enhanced malignant transformation induced by expression of a distinct protein domain of ribonucleotide reductase large subunit from herpes simplex virus type 2.

    PubMed Central

    Ali, M A; McWeeney, D; Milosavljevic, A; Jurka, J; Jariwalla, R J

    1991-01-01

    The 1.3-kilobase (kb) Pst I DNA fragment C (Pst I-C) of herpes simplex virus type 2 (HSV-2) morphological transforming region III (mtrIII; map unit 0.562-0.570) encodes part of the N-terminal half of the large subunit of ribonucleotide reductase (RR1; amino acid residues 71-502) and induces the neoplastic transformation of immortalized cell lines. To assess directly the role of these RR1 protein sequences in cell transformation, the Pst I-C fragment was cloned in an expression vector (p91023) containing an adenovirus-simian virus 40 promoter-enhancer to generate recombinant plasmid p9-C. Expression of a protein domain (approximately 65 kDa) was observed in p9-C-transfected COS-7 and Rat2 cells but not in those transfected with plasmid pHC-14 (Pst I-C in a promoterless vector). In Rat2 cells, p9-C induced highly transformed foci at an elevated frequency compared with that of pHC-14. Introduction of translation termination (TAG) condons within the RR1 coding sequence and within all three reading frames inactivated RR1 protein expression from p9-C and reduced its transforming activity to the level seen with the standard pHC-14 construct. Wild-type p9-C specified a protein kinase capable of autophosphorylation. Computer-assisted analysis further revealed significant similarity between regions of mtrIII-specific RR1 and amino acid patterns conserved within the proinsulin precursor family and DNA transposition proteins. These results identify a distinct domain of the HSV-2 RR1 protein involved in the induction of enhanced malignant transformation. In addition, the data indicate that the mtrIII DNA itself can induce basal-level transformation in the absence of protein expression. Images PMID:1654564

  4. Coexisting orchid species have distinct mycorrhizal communities and display strong spatial segregation.

    PubMed

    Jacquemyn, Hans; Brys, Rein; Merckx, Vincent S F T; Waud, Michael; Lievens, Bart; Wiegand, Thorsten

    2014-04-01

    Because orchids are dependent on mycorrhizal fungi for germination and establishment of seedlings, differences in the mycorrhizal communities associating with orchids can be expected to mediate the abundance, spatial distribution and coexistence of terrestrial orchids in natural communities. We assessed the small-scale spatial distribution of seven orchid species co-occurring in 25 × 25 m plots in two Mediterranean grasslands. In order to characterize the mycorrhizal community associating with each orchid species, 454 pyrosequencing was used. The extent of spatial clustering was assessed using techniques of spatial point pattern analysis. The community of mycorrhizal fungi consisted mainly of members of the Tulasnellaceae, Thelephoraceae and Ceratobasidiaceae, although sporadically members of the Sebacinaceae, Russulaceae and Cortinariaceae were observed. Pronounced differences in mycorrhizal communities were observed between species, whereas strong clustering and significant segregation characterized the spatial distribution of orchid species. However, spatial segregation was not significantly related to phylogenetic dissimilarity of fungal communities. Our results indicate that co-occurring orchid species have distinctive mycorrhizal communities and show strong spatial segregation, suggesting that mycorrhizal fungi are important factors driving niche partitioning in terrestrial orchids and may therefore contribute to orchid coexistence.

  5. Testing the utility of mitochondrial cytochrome oxidase subunit 1 sequences for phylogenetic estimates of relationships between crane (Grus) species.

    PubMed

    Yu, D B; Chen, R; Kaleri, H A; Jiang, B C; Xu, H X; Du, W-X

    2011-12-21

    Morphology and biogeography are widely used in animal taxonomy. Recent study has suggested that a DNA-based identification system, using a 648-bp portion of the mitochondrial gene cytochrome oxidase subunit 1 (CO1), also known as the barcoding gene, can aid in the resolution of inferences concerning phylogenetic relationships and for identification of species. However, the effectiveness of DNA barcoding for identifying crane species is unknown. We amplified and sequenced 894-bp DNA fragments of CO1 from Grus japonensis (Japanese crane), G. grus (Eurasian crane), G. monacha (hooded crane), G. canadensis (sandhill crane), G. leucogeranus (Siberian crane), and Balearica pavonina (crowned crane), along with those of 15 species obtained from GenBank and DNA barcoding, to construct four algorithms using Tringa stagnatilis, Scolopax rusticola, and T. erythropus as outgroups. The four phylum profiles showed good resolution of the major taxonomic groups. We concluded that reconstruction of the molecular phylogenetic tree can be helpful for classification and that CO1 sequences are suitable for studying the molecular evolution of cranes. Although support for several deeper branches was limited, CO1 data gave remarkably good separations, especially considering that our analysis was based on just a fragment of the gene and that CO1 has generally been viewed as useful only for resolving shallow divergences.

  6. Testing the utility of mitochondrial cytochrome oxidase subunit 1 sequences for phylogenetic estimates of relationships between crane (Grus) species.

    PubMed

    Yu, D B; Chen, R; Kaleri, H A; Jiang, B C; Xu, H X; Du, W-X

    2011-01-01

    Morphology and biogeography are widely used in animal taxonomy. Recent study has suggested that a DNA-based identification system, using a 648-bp portion of the mitochondrial gene cytochrome oxidase subunit 1 (CO1), also known as the barcoding gene, can aid in the resolution of inferences concerning phylogenetic relationships and for identification of species. However, the effectiveness of DNA barcoding for identifying crane species is unknown. We amplified and sequenced 894-bp DNA fragments of CO1 from Grus japonensis (Japanese crane), G. grus (Eurasian crane), G. monacha (hooded crane), G. canadensis (sandhill crane), G. leucogeranus (Siberian crane), and Balearica pavonina (crowned crane), along with those of 15 species obtained from GenBank and DNA barcoding, to construct four algorithms using Tringa stagnatilis, Scolopax rusticola, and T. erythropus as outgroups. The four phylum profiles showed good resolution of the major taxonomic groups. We concluded that reconstruction of the molecular phylogenetic tree can be helpful for classification and that CO1 sequences are suitable for studying the molecular evolution of cranes. Although support for several deeper branches was limited, CO1 data gave remarkably good separations, especially considering that our analysis was based on just a fragment of the gene and that CO1 has generally been viewed as useful only for resolving shallow divergences. PMID:22194213

  7. Visual stimuli that elicit appetitive behaviors in three morphologically distinct species of praying mantis.

    PubMed

    Prete, Frederick R; Komito, Justin L; Dominguez, Salina; Svenson, Gavin; López, LeoLin Y; Guillen, Alex; Bogdanivich, Nicole

    2011-09-01

    We assessed the differences in appetitive responses to visual stimuli by three species of praying mantis (Insecta: Mantodea), Tenodera aridifolia sinensis, Mantis religiosa, and Cilnia humeralis. Tethered, adult females watched computer generated stimuli (erratically moving disks or linearly moving rectangles) that varied along predetermined parameters. Three responses were scored: tracking, approaching, and striking. Threshold stimulus size (diameter) for tracking and striking at disks ranged from 3.5 deg (C. humeralis) to 7.8 deg (M. religiosa), and from 3.3 deg (C. humeralis) to 11.7 deg (M. religiosa), respectively. Unlike the other species which struck at disks as large as 44 deg, T. a. sinensis displayed a preference for 14 deg disks. Disks moving at 143 deg/s were preferred by all species. M. religiosa exhibited the most approaching behavior, and with T. a. sinensis distinguished between rectangular stimuli moving parallel versus perpendicular to their long axes. C. humeralis did not make this distinction. Stimulus sizes that elicited the target behaviors were not related to mantis size. However, differences in compound eye morphology may be related to species differences: C. humeralis' eyes are farthest apart, and it has an apparently narrower binocular visual field which may affect retinal inputs to movement-sensitive visual interneurons. PMID:21553126

  8. Phenotypic Convergence in Genetically Distinct Lineages of a Rhinolophus Species Complex (Mammalia, Chiroptera)

    PubMed Central

    Jacobs, David S.; Babiker, Hassan; Bastian, Anna; Kearney, Teresa; van Eeden, Rowen; Bishop, Jacqueline M.

    2013-01-01

    Phenotypes of distantly related species may converge through adaptation to similar habitats and/or because they share biological constraints that limit the phenotypic variants produced. A common theme in bats is the sympatric occurrence of cryptic species that are convergent in morphology but divergent in echolocation frequency, suggesting that echolocation may facilitate niche partitioning, reducing competition. If so, allopatric populations freed from competition, could converge in both morphology and echolocation provided they occupy similar niches or share biological constraints. We investigated the evolutionary history of a widely distributed African horseshoe bat, Rhinolophus darlingi, in the context of phenotypic convergence. We used phylogenetic inference to identify and date lineage divergence together with phenotypic comparisons and ecological niche modelling to identify morphological and geographical correlates of those lineages. Our results indicate that R. darlingi is paraphyletic, the eastern and western parts of its distribution forming two distinct non-sister lineages that diverged ~9.7 Mya. We retain R. darlingi for the eastern lineage and argue that the western lineage, currently the sub-species R. d. damarensis, should be elevated to full species status. R. damarensis comprises two lineages that diverged ~5 Mya. Our findings concur with patterns of divergence of other co-distributed taxa which are associated with increased regional aridification between 7-5 Mya suggesting possible vicariant evolution. The morphology and echolocation calls of R. darlingi and R. damarensis are convergent despite occupying different biomes. This suggests that adaptation to similar habitats is not responsible for the convergence. Furthermore, R. darlingi forms part of a clade comprising species that are bigger and echolocate at lower frequencies than R. darlingi, suggesting that biological constraints are unlikely to have influenced the convergence. Instead, the

  9. DNA-dependent RNA polymerase III from the fungus Podospora comata. Purification, subunit structure and comparison with the homologous enzyme of a related species.

    PubMed

    Barreau, C; Begueret, J

    1982-12-15

    DNA-dependent RNA polymerase III has been purified to homogeneity from the filamentous fungus Podospora comata. The enzyme was extracted at low ionic strength, separated from the polymerases I and II by DEAE-Sephadex chromatography and purified by heparin-Sepharose and phosphocellulose chromatography; 0.1-0.2 mg highly purified homogeneous enzyme with a specific activity of 220 units/mg could be obtained from 2 kg wet mycelium. The subunit composition of the enzyme was determined after sodium dodecyl sulphate/polyacrylamide gel electrophoresis; thirteen putative subunits of molecular weight 174000 (a), 129000 b), 87000 (c), 50000 (d), 39000 (e), 23500 (f), 21000 (g), 19000 (h), 17000 (i), 16500 (j), 13500 (k), 11000 (l) and 10000 (m) were identified. All of the polypeptide components of the enzyme are present in about integral stoichiometric amounts as judged by dye binding. The presence of subunit Mr = 87000 in a molar ratio 1:1 is necessary to obtain very active enzyme. Thirteen homologous subunits were observed in a preparation of RNA polymerase III from Podospora anserina, which is a related species. Only subunit i is different in the two species. PMID:7151805

  10. Beyond the EDGE with EDAM: Prioritising British Plant Species According to Evolutionary Distinctiveness, and Accuracy and Magnitude of Decline

    PubMed Central

    Pearse, William D.; Chase, Mark W.; Crawley, Michael J.; Dolphin, Konrad; Fay, Michael F.; Joseph, Jeffrey A.; Powney, Gary; Preston, Chris D.; Rapacciuolo, Giovanni; Roy, David B.; Purvis, Andy

    2015-01-01

    Conservation biologists have only finite resources, and so must prioritise some species over others. The EDGE-listing approach ranks species according to their combined evolutionary distinctiveness and degree of threat, but ignores the uncertainty surrounding both threat and evolutionary distinctiveness. We develop a new family of measures for species, which we name EDAM, that incorporates evolutionary distinctiveness, the magnitude of decline, and the accuracy with which decline can be predicted. Further, we show how the method can be extended to explore phyogenetic uncertainty. Using the vascular plants of Britain as a case study, we find that the various EDAM measures emphasise different species and parts of Britain, and that phylogenetic uncertainty can strongly affect the prioritisation scores of some species. PMID:26018568

  11. Subunit Composition and Substrate Specificity of a MOF-containing Histone Acetyltransferase Distinct from the Male-specific Lethal (MSL) Complex*

    PubMed Central

    Cai, Yong; Jin, Jingji; Swanson, Selene K.; Cole, Michael D.; Choi, Seung Hyuk; Florens, Laurence; Washburn, Michael P.; Conaway, Joan W.; Conaway, Ronald C.

    2010-01-01

    Human MOF (MYST1), a member of the MYST (Moz-Ybf2/Sas3-Sas2-Tip60) family of histone acetyltransferases (HATs), is the human ortholog of the Drosophila males absent on the first (MOF) protein. MOF is the catalytic subunit of the male-specific lethal (MSL) HAT complex, which plays a key role in dosage compensation in the fly and is responsible for a large fraction of histone H4 lysine 16 (H4K16) acetylation in vivo. MOF was recently reported to be a component of a second HAT complex, designated the non-specific lethal (NSL) complex (Mendjan, S., Taipale, M., Kind, J., Holz, H., Gebhardt, P., Schelder, M., Vermeulen, M., Buscaino, A., Duncan, K., Mueller, J., Wilm, M., Stunnenberg, H. G., Saumweber, H., and Akhtar, A. (2006) Mol. Cell 21, 811–823). Here we report an analysis of the subunit composition and substrate specificity of the NSL complex. Proteomic analyses of complexes purified through multiple candidate subunits reveal that NSL is composed of nine subunits. Two of its subunits, WD repeat domain 5 (WDR5) and host cell factor 1 (HCF1), are shared with members of the MLL/SET family of histone H3 lysine 4 (H3K4) methyltransferase complexes, and a third subunit, MCRS1, is shared with the human INO80 chromatin-remodeling complex. In addition, we show that assembly of the MOF HAT into MSL or NSL complexes controls its substrate specificity. Although MSL-associated MOF acetylates nucleosomal histone H4 almost exclusively on lysine 16, NSL-associated MOF exhibits a relaxed specificity and also acetylates nucleosomal histone H4 on lysines 5 and 8. PMID:20018852

  12. Achromobacter Species Isolated from Cystic Fibrosis Patients Reveal Distinctly Different Biofilm Morphotypes.

    PubMed

    Nielsen, Signe M; Nørskov-Lauritsen, Niels; Bjarnsholt, Thomas; Meyer, Rikke L

    2016-01-01

    Achromobacter species have attracted attention as emerging pathogens in cystic fibrosis. The clinical significance of Achromobacter infection is not yet fully elucidated; however, their intrinsic resistance to antimicrobials and ability to form biofilms renders them capable of establishing long-term chronic infections. Still, many aspects of Achromobacter biofilm formation remain uncharacterized. In this study, we characterized biofilm formation in clinical isolates of Achromobacter and investigated the effect of challenging the biofilm with antimicrobials and/or enzymes targeting the extracellular matrix. In vitro biofilm growth and subsequent visualization by confocal microscopy revealed distinctly different biofilm morphotypes: a surface-attached biofilm morphotype of small aggregates and an unattached biofilm morphotype of large suspended aggregates. Aggregates consistent with our in vitro findings were visualized in sputum samples from cystic fibrosis patients using an Achromobacter specific peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) probe, confirming the presence of Achromobacter biofilms in the CF lung. High antibiotic tolerance was associated with the biofilm phenotype, and biocidal antibiotic concentrations were up to 1000 fold higher than for planktonic cultures. Treatment with DNase or subtilisin partially dispersed the biofilm and reduced the tolerance to specific antimicrobials, paving the way for further research into using dispersal mechanisms to improve treatment strategies. PMID:27681927

  13. Genome Sequencing and Analysis of Catopsilia pomona nucleopolyhedrovirus: A Distinct Species in Group I Alphabaculovirus

    PubMed Central

    Wang, Jun; Zhu, Zheng; Zhang, Lei; Hou, Dianhai; Wang, Manli; Arif, Basil; Kou, Zheng; Wang, Hualin; Deng, Fei; Hu, Zhihong

    2016-01-01

    The genome sequence of Catopsilia pomona nucleopolyhedrovirus (CapoNPV) was determined by the Roche 454 sequencing system. The genome consisted of 128,058 bp and had an overall G+C content of 40%. There were 130 hypothetical open reading frames (ORFs) potentially encoding proteins of more than 50 amino acids and covering 92% of the genome. Among all the hypothetical ORFs, 37 baculovirus core genes, 23 lepidopteran baculovirus conserved genes and 10 genes conserved in Group I alphabaculoviruses were identified. In addition, the genome included regions of 8 typical baculoviral homologous repeat sequences (hrs). Phylogenic analysis showed that CapoNPV was in a distinct branch of clade “a” in Group I alphabaculoviruses. Gene parity plot analysis and overall similarity of ORFs indicated that CapoNPV is more closely related to the Group I alphabaculoviruses than to other baculoviruses. Interesting, CapoNPV lacks the genes encoding the fibroblast growth factor (fgf) and ac30, which are conserved in most lepidopteran and Group I baculoviruses, respectively. Sequence analysis of the F-like protein of CapoNPV showed that some amino acids were inserted into the fusion peptide region and the pre-transmembrane region of the protein. All these unique features imply that CapoNPV represents a member of a new baculovirus species. PMID:27166956

  14. Achromobacter Species Isolated from Cystic Fibrosis Patients Reveal Distinctly Different Biofilm Morphotypes

    PubMed Central

    Nielsen, Signe M.; Nørskov-Lauritsen, Niels; Bjarnsholt, Thomas; Meyer, Rikke L.

    2016-01-01

    Achromobacter species have attracted attention as emerging pathogens in cystic fibrosis. The clinical significance of Achromobacter infection is not yet fully elucidated; however, their intrinsic resistance to antimicrobials and ability to form biofilms renders them capable of establishing long-term chronic infections. Still, many aspects of Achromobacter biofilm formation remain uncharacterized. In this study, we characterized biofilm formation in clinical isolates of Achromobacter and investigated the effect of challenging the biofilm with antimicrobials and/or enzymes targeting the extracellular matrix. In vitro biofilm growth and subsequent visualization by confocal microscopy revealed distinctly different biofilm morphotypes: a surface-attached biofilm morphotype of small aggregates and an unattached biofilm morphotype of large suspended aggregates. Aggregates consistent with our in vitro findings were visualized in sputum samples from cystic fibrosis patients using an Achromobacter specific peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) probe, confirming the presence of Achromobacter biofilms in the CF lung. High antibiotic tolerance was associated with the biofilm phenotype, and biocidal antibiotic concentrations were up to 1000 fold higher than for planktonic cultures. Treatment with DNase or subtilisin partially dispersed the biofilm and reduced the tolerance to specific antimicrobials, paving the way for further research into using dispersal mechanisms to improve treatment strategies. PMID:27681927

  15. Differential production of reactive oxygen species in distinct brain regions of hypoglycemic mice.

    PubMed

    Amador-Alvarado, Leticia; Montiel, Teresa; Massieu, Lourdes

    2014-09-01

    Hypoglycemia is a serious complication of insulin therapy in patients suffering from type 1 Diabetes Mellitus. Severe hypoglycemia leading to coma (isoelectricity) induces massive neuronal death in vulnerable brain regions such as the hippocampus, the striatum and the cerebral cortex. It has been suggested that the production of reactive oxygen species (ROS) and oxidative stress is involved in hypoglycemic brain damage, and that ROS generation is stimulated by glucose reintroduction (GR) after the hypoglycemic coma. However, the distribution of ROS in discrete brain regions has not been studied in detail. Using the oxidation sensitive marker dihydroethidium (DHE) we have investigated the distribution of ROS in different regions of the mouse brain during prolonged severe hypoglycemia without isoelectricity, as well as the effect of GR on ROS levels. Results show that ROS generation increases in the hippocampus, the cerebral cortex and the striatum after prolonged severe hypoglycemia before the coma. The hippocampus showed the largest increases in ROS levels. GR further stimulated ROS production in the hippocampus and the striatum while in the cerebral cortex, only the somatosensory and parietal areas were significantly affected by GR. Results suggest that ROS are differentially produced during the hypoglycemic insult and that a different response to GR is present among distinct brain regions.

  16. Evidence for a Common Origin of Homomorphic and Heteromorphic Sex Chromosomes in Distinct Spinacia Species

    PubMed Central

    Fujito, Satoshi; Takahata, Satoshi; Suzuki, Reimi; Hoshino, Yoichiro; Ohmido, Nobuko; Onodera, Yasuyuki

    2015-01-01

    The dioecious genus Spinacia is thought to include two wild relatives (S. turkestanica Ilj. and S. tetrandra Stev.) of cultivated spinach (S. oleracea L.). In this study, nuclear and chloroplast sequences from 21 accessions of Spinacia germplasm and six spinach cultivars or lines were subjected to phylogenetic analysis to define the relationships among the three species. Maximum-likelihood sequence analysis suggested that the Spinacia plant samples could be classified into two monophyletic groups (Group 1 and Group 2): Group 1 consisted of all accessions, cultivars, and lines of S. oleracea L. and S. turkestanica Ilj. and two of five S. tetrandra Stev. accessions, whereas Group 2 was composed of the three remaining S. tetrandra Stev. accessions. By using flow cytometry, we detected a distinct difference in nuclear genome size between the groups. Group 2 also was characterized by a sexual dimorphism in inflorescence structure, which was not observed in Group 1. Interspecific crosses between the groups produced hybrids with drastically reduced pollen fertility and showed that the male is the heterogametic sex (XY) in Group 2, as is the case in S. oleracea L. (Group 1). Cytogenetic and DNA marker analyses suggested that Group 1 and Group 2 have homomorphic and heteromorphic sex chromosome pairs (XY), respectively, and that the sex chromosome pairs of the two groups evolved from a common ancestral pair. Our data suggest that the Spinacia genus may serve as a good model for investigation of evolutionary mechanisms underlying the emergence of heteromorphic sex chromosome pairs from ancestral homomorphic pairs. PMID:26048564

  17. Achromobacter Species Isolated from Cystic Fibrosis Patients Reveal Distinctly Different Biofilm Morphotypes

    PubMed Central

    Nielsen, Signe M.; Nørskov-Lauritsen, Niels; Bjarnsholt, Thomas; Meyer, Rikke L.

    2016-01-01

    Achromobacter species have attracted attention as emerging pathogens in cystic fibrosis. The clinical significance of Achromobacter infection is not yet fully elucidated; however, their intrinsic resistance to antimicrobials and ability to form biofilms renders them capable of establishing long-term chronic infections. Still, many aspects of Achromobacter biofilm formation remain uncharacterized. In this study, we characterized biofilm formation in clinical isolates of Achromobacter and investigated the effect of challenging the biofilm with antimicrobials and/or enzymes targeting the extracellular matrix. In vitro biofilm growth and subsequent visualization by confocal microscopy revealed distinctly different biofilm morphotypes: a surface-attached biofilm morphotype of small aggregates and an unattached biofilm morphotype of large suspended aggregates. Aggregates consistent with our in vitro findings were visualized in sputum samples from cystic fibrosis patients using an Achromobacter specific peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) probe, confirming the presence of Achromobacter biofilms in the CF lung. High antibiotic tolerance was associated with the biofilm phenotype, and biocidal antibiotic concentrations were up to 1000 fold higher than for planktonic cultures. Treatment with DNase or subtilisin partially dispersed the biofilm and reduced the tolerance to specific antimicrobials, paving the way for further research into using dispersal mechanisms to improve treatment strategies.

  18. Distinct Origin of the Y and St Genome in Elymus Species: Evidence from the Analysis of a Large Sample of St Genome Species Using Two Nuclear Genes

    PubMed Central

    Yan, Chi; Sun, Genlou; Sun, Dongfa

    2011-01-01

    Background Previous cytological and single copy nuclear genes data suggested the St and Y genome in the StY-genomic Elymus species originated from different donors: the St from a diploid species in Pseudoroegneria and the Y from an unknown diploid species, which are now extinct or undiscovered. However, ITS data suggested that the Y and St genome shared the same progenitor although rather few St genome species were studied. In a recent analysis of many samples of St genome species Pseudoroegneria spicata (Pursh) À. Löve suggested that one accession of P. spicata species was the most likely donor of the Y genome. The present study tested whether intraspecific variation during sampling could affect the outcome of analyses to determining the origin of Y genome in allotetraploid StY species. We also explored the evolutionary dynamics of these species. Methodology/Principal Findings Two single copy nuclear genes, the second largest subunit of RNA polymerase II (RPB2) and the translation elongation factor G (EF-G) sequences from 58 accessions of Pseudoroegneria and Elymus species, together with those from Hordeum (H), Agropyron (P), Australopyrum (W), Lophopyrum (Ee), Thinopyrum (Ea), Thinopyrum (Eb), and Dasypyrum (V) were analyzed using maximum parsimony, maximum likelihood and Bayesian methods. Sequence comparisons among all these genomes revealed that the St and Y genomes are relatively dissimilar. Extensive sequence variations have been detected not only between the sequences from St and Y genome, but also among the sequences from diploid St genome species. Phylogenetic analyses separated the Y sequences from the St sequences. Conclusions/Significance Our results confirmed that St and Y genome in Elymus species have originated from different donors, and demonstrated that intraspecific variation does not affect the identification of genome origin in polyploids. Moreover, sequence data showed evidence to support the suggestion of the genome convergent evolution in

  19. DNA barcoding, MALDI-TOF, and AFLP data support Fusarium ficicrescens as a distinct species within the Fusarium fujikuroi species complex.

    PubMed

    Al-Hatmi, Abdullah M S; Mirabolfathy, Mansoureh; Hagen, Ferry; Normand, Anne-Cécile; Stielow, J Benjamin; Karami-Osbo, Rouhollah; van Diepeningen, Anne D; Meis, Jacques F; de Hoog, G Sybren

    2016-02-01

    The Fusarium fujikuroi species complex (FFSC) is one of the most common groups of fusaria associated with plant diseases, mycotoxin production and traumatic and disseminated human infections. Here we present the description and taxonomy of a new taxon, Fusarium ficicrescens sp. nov., collected from contaminated fig fruits in Iran. Initially this species was identified as Fusarium andiyazi by morphology. In the present study the species was studied by multilocus sequence analysis, amplified fragment length polymorphism (AFLP), matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and phenotypic characters. Multilocus analyses were based on translation elongation factor 1α (TEF1), RNA polymerase subunit (RPB2) and beta-tubulin (BT2) and proved F. ficicrescens as a member of the FFSC. Phylogenetic analysis showed that the fungus is closely related to Fusarium lactis, Fusarium ramigenum, and Fusarium napiforme; known plant pathogens, mycotoxin producers, and occasionally occurring multidrug resistant opportunists. The new species differed by being able to grow at 37 °C and by the absence of mycotoxin production. TEF1 was confirmed as an essential barcode for identifying Fusarium species. In addition to TEF1, we evaluated BT2 and RPB2 in order to provide sufficient genetic and species boundaries information for recognition of the novel species.

  20. 75 FR 30769 - Endangered and Threatened Species; Proposed Listing of Nine Distinct Population Segments of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-02

    ... Distinct Population Segments of Loggerhead Sea Turtles as Endangered or Threatened; Extension of Comment... proposed listing of nine distinct population segments of loggerhead sea turtles as endangered or threatened... . Mail: NMFS National Sea Turtle Coordinator, Attn: Loggerhead Proposed Listing Rule, Office of...

  1. 77 FR 64959 - Endangered and Threatened Species; Initiation of 5-Year Review for the Southern Distinct...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-24

    ... update, if necessary, the Northern DPS' status in five years time (70 FR 17386; April 6, 2005). Therefore...; Initiation of 5-Year Review for the Southern Distinct Population Segment of North American Green Sturgeon... 5-year review of the Southern Distinct Population Segment (DPS) of North American green...

  2. First comparative characterization of three distinct ferritin subunits from a teleost: Evidence for immune-responsive mRNA expression and iron depriving activity of seahorse (Hippocampus abdominalis) ferritins.

    PubMed

    Oh, Minyoung; Umasuthan, Navaneethaiyer; Elvitigala, Don Anushka Sandaruwan; Wan, Qiang; Jo, Eunyoung; Ko, Jiyeon; Noh, Gyeong Eon; Shin, Sangok; Rho, Sum; Lee, Jehee

    2016-02-01

    Ferritins play an indispensable role in iron homeostasis through their iron-withholding function in living beings. In the current study, cDNA sequences of three distinct ferritin subunits, including a ferritin H, a ferritin M, and a ferritin L, were identified from big belly seahorse, Hippocampus abdominalis, and molecularly characterized. Complete coding sequences (CDS) of seahorse ferritin H (HaFerH), ferritin M (HaFerM), and ferritin L (HaFerL) subunits were comprised of 531, 528, and 522 base pairs (bp), respectively, which encode polypeptides of 177, 176, and 174 amino acids, respectively, with molecular masses of ∼20-21 kDa. Our in silico analyses demonstrate that these three ferritin subunits exhibit the typical characteristics of ferritin superfamily members including iron regulatory elements, domain signatures, and reactive centers. The coding sequences of HaFerH, M, and L were cloned and the corresponding proteins were overexpressed in a bacterial system. Recombinantly expressed HaFer proteins demonstrated detectable in vivo iron sequestrating (ferroxidase) activity, consistent with their putative iron binding capability. Quantification of the basal expression of these three HaFer sequences in selected tissues demonstrated a gene-specific ubiquitous spatial distribution pattern, with abundance of mRNA in HaFerM in the liver and predominant expression of HaFerH and HaFerL in blood. Interestingly, the basal expression of all three ferritin genes was found to be significantly modulated against pathogenic stress mounted by lipopolysaccharides (LPS), poly I:C, Streptococcus iniae, and Edwardsiella tarda. Collectively, our findings suggest that the three HaFer subunits may be involved in iron (II) homeostasis in big belly seahorse and that they are important in its host defense mechanisms.

  3. First comparative characterization of three distinct ferritin subunits from a teleost: Evidence for immune-responsive mRNA expression and iron depriving activity of seahorse (Hippocampus abdominalis) ferritins.

    PubMed

    Oh, Minyoung; Umasuthan, Navaneethaiyer; Elvitigala, Don Anushka Sandaruwan; Wan, Qiang; Jo, Eunyoung; Ko, Jiyeon; Noh, Gyeong Eon; Shin, Sangok; Rho, Sum; Lee, Jehee

    2016-02-01

    Ferritins play an indispensable role in iron homeostasis through their iron-withholding function in living beings. In the current study, cDNA sequences of three distinct ferritin subunits, including a ferritin H, a ferritin M, and a ferritin L, were identified from big belly seahorse, Hippocampus abdominalis, and molecularly characterized. Complete coding sequences (CDS) of seahorse ferritin H (HaFerH), ferritin M (HaFerM), and ferritin L (HaFerL) subunits were comprised of 531, 528, and 522 base pairs (bp), respectively, which encode polypeptides of 177, 176, and 174 amino acids, respectively, with molecular masses of ∼20-21 kDa. Our in silico analyses demonstrate that these three ferritin subunits exhibit the typical characteristics of ferritin superfamily members including iron regulatory elements, domain signatures, and reactive centers. The coding sequences of HaFerH, M, and L were cloned and the corresponding proteins were overexpressed in a bacterial system. Recombinantly expressed HaFer proteins demonstrated detectable in vivo iron sequestrating (ferroxidase) activity, consistent with their putative iron binding capability. Quantification of the basal expression of these three HaFer sequences in selected tissues demonstrated a gene-specific ubiquitous spatial distribution pattern, with abundance of mRNA in HaFerM in the liver and predominant expression of HaFerH and HaFerL in blood. Interestingly, the basal expression of all three ferritin genes was found to be significantly modulated against pathogenic stress mounted by lipopolysaccharides (LPS), poly I:C, Streptococcus iniae, and Edwardsiella tarda. Collectively, our findings suggest that the three HaFer subunits may be involved in iron (II) homeostasis in big belly seahorse and that they are important in its host defense mechanisms. PMID:26747640

  4. Identification of Species and Sources of Cryptosporidium Oocysts in Storm Waters with a Small-Subunit rRNA-Based Diagnostic and Genotyping Tool

    PubMed Central

    Xiao, Lihua; Alderisio, Kerri; Limor, Josef; Royer, Michael; Lal, Altaf A.

    2000-01-01

    The identification of Cryptosporidium oocysts in environmental samples is largely made by the use of an immunofluorescent assay. In this study, we have used a small-subunit rRNA-based PCR-restriction fragment length polymorphism technique to identify species and sources of Cryptosporidium oocysts present in 29 storm water samples collected from a stream in New York. A total of 12 genotypes were found in 27 positive samples; for 4 the species and probable origins were identified by sequence analysis, whereas the rest represent new genotypes from wildlife. Thus, this technique provides an alternative method for the detection and differentiation of Cryptosporidium parasites in environmental samples. PMID:11097935

  5. Diversity of heterotrimeric G-protein γ subunits in plants

    PubMed Central

    2012-01-01

    Background Heterotrimeric G-proteins, consisting of three subunits Gα, Gβ and Gγ are present in most eukaryotes and mediate signaling in numerous biological processes. In plants, Gγ subunits were shown to provide functional selectivity to G-proteins. Three unconventional Gγ subunits were recently reported in Arabidopsis, rice and soybean but no structural analysis has been reported so far. Their relationship with conventional Gγ subunits and taxonomical distribution has not been yet demonstrated. Results After an extensive similarity search through plant genomes, transcriptomes and proteomes we assembled over 200 non-redundant proteins related to the known Gγ subunits. Structural analysis of these sequences revealed that most of them lack the obligatory C-terminal prenylation motif (CaaX). According to their C-terminal structures we classified the plant Gγ subunits into three distinct types. Type A consists of Gγ subunits with a putative prenylation motif. Type B subunits lack a prenylation motif and do not have any cysteine residues in the C-terminal region, while type C subunits contain an extended C-terminal domain highly enriched with cysteines. Comparative analysis of C-terminal domains of the proteins, intron-exon arrangement of the corresponding genes and phylogenetic studies suggested a common origin of all plant Gγ subunits. Conclusion Phylogenetic analyses suggest that types C and B most probably originated independently from type A ancestors. We speculate on a potential mechanism used by those Gγ subunits lacking isoprenylation motifs to anchor the Gβγ dimer to the plasma membrane and propose a new flexible nomenclature for plant Gγ subunits. Finally, in the light of our new classification, we give a word of caution about the interpretation of Gγ research in Arabidopsis and its generalization to other plant species. PMID:23113884

  6. Poles apart: the "bipolar" pteropod species Limacina helicina is genetically distinct between the Arctic and Antarctic oceans.

    PubMed

    Hunt, Brian; Strugnell, Jan; Bednarsek, Nina; Linse, Katrin; Nelson, R John; Pakhomov, Evgeny; Seibel, Brad; Steinke, Dirk; Würzberg, Laura

    2010-03-23

    The shelled pteropod (sea butterfly) Limacina helicina is currently recognised as a species complex comprising two sub-species and at least five "forma". However, at the species level it is considered to be bipolar, occurring in both the Arctic and Antarctic oceans. Due to its aragonite shell and polar distribution L. helicina is particularly vulnerable to ocean acidification. As a key indicator of the acidification process, and a major component of polar ecosystems, L. helicina has become a focus for acidification research. New observations that taxonomic groups may respond quite differently to acidification prompted us to reassess the taxonomic status of this important species. We found a 33.56% (+/-0.09) difference in cytochrome c oxidase subunit I (COI) gene sequences between L. helicina collected from the Arctic and Antarctic oceans. This degree of separation is sufficient for ordinal level taxonomic separation in other organisms and provides strong evidence for the Arctic and Antarctic populations of L. helicina differing at least at the species level. Recent research has highlighted substantial physiological differences between the poles for another supposedly bipolar pteropod species, Clione limacina. Given the large genetic divergence between Arctic and Antarctic L. helicina populations shown here, similarly large physiological differences may exist between the poles for the L. helicina species group. Therefore, in addition to indicating that L. helicina is in fact not bipolar, our study demonstrates the need for acidification research to take into account the possibility that the L. helicina species group may not respond in the same way to ocean acidification in Arctic and Antarctic ecosystems.

  7. Molecular Identification of Necrophagous Muscidae and Sarcophagidae Fly Species Collected in Korea by Mitochondrial Cytochrome c Oxidase Subunit I Nucleotide Sequences

    PubMed Central

    Ham, Chan Seon; Kim, Seong Yoon; Ko, Kwang Soo; Jo, Tae-Ho; Son, Gi Hoon

    2014-01-01

    Identification of insect species is an important task in forensic entomology. For more convenient species identification, the nucleotide sequences of cytochrome c oxidase subunit I (COI) gene have been widely utilized. We analyzed full-length COI nucleotide sequences of 10 Muscidae and 6 Sarcophagidae fly species collected in Korea. After DNA extraction from collected flies, PCR amplification and automatic sequencing of the whole COI sequence were performed. Obtained sequences were analyzed for a phylogenetic tree and a distance matrix. Our data showed very low intraspecific sequence distances and species-level monophylies. However, sequence comparison with previously reported sequences revealed a few inconsistencies or paraphylies requiring further investigation. To the best of our knowledge, this study is the first report of COI nucleotide sequences from Hydrotaea occulta, Muscina angustifrons, Muscina pascuorum, Ophyra leucostoma, Sarcophaga haemorrhoidalis, Sarcophaga harpax, and Phaonia aureola. PMID:24982938

  8. Targeted deletion of one or two copies of the G protein β subunit Gβ5 gene has distinct effects on body weight and behavior in mice.

    PubMed

    Wang, Qiang; Levay, Konstantin; Chanturiya, Tatyana; Dvoriantchikova, Galina; Anderson, Karen L; Bianco, Suzy D C; Ueta, Cintia B; Molano, R Damaris; Pileggi, Antonello; Gurevich, Eugenia V; Gavrilova, Oksana; Slepak, Vladlen Z

    2011-11-01

    We investigated the physiological role of Gβ5, a unique G protein β subunit that dimerizes with regulators of G protein signaling (RGS) proteins of the R7 family instead of Gγ. Gβ5 is essential for stability of these complexes, so that its knockout (KO)causes degradation of the entire Gβ5-R7 family. We report that the Gβ5-KO mice remain leaner than the wild type (WT) throughout their lifetime and are resistant to a high-fat diet. They have a 5-fold increase in locomotor activity, increased thermogenesis, and lower serum insulin, all of which correlate with a higher level of secreted epinephrine. Heterozygous (HET) mice are 2-fold more active than WT mice. Surprisingly, with respect to body weight, the HET mice display a phenotype opposite to that of the KO mice: by the age of 6 mo, they are ≥ 15% heavier than the WT and have increased adiposity, insulin resistance, and liver steatosis. These changes occur in HET mice fed a normal diet and without apparent hyperphagia, mimicking basic characteristics of human metabolic syndrome. We conclude that even a partial reduction in Gβ5-R7 level can perturb normal animal metabolism and behavior. Our data on Gβ5 haploinsufficient mice may explain earlier observations of genetic linkage between R7 family mutations and obesity in humans.

  9. Single-Cell DNA barcoding using sequences from the small subunit rRNA and internal transcribed spacer region identifies new species of Trichonympha and Trichomitopsis from the hindgut of the termite Zootermopsis angusticollis.

    PubMed

    Tai, Vera; James, Erick R; Perlman, Steve J; Keeling, Patrick J

    2013-01-01

    To aid in their digestion of wood, lower termites are known to harbour a diverse community of prokaryotes as well as parabasalid and oxymonad protist symbionts. One of the best-studied lower termite gut communities is that of Zootermopsis angusticollis which has been known for almost 100 years to possess 3 species of Trichonympha (T. campanula, T. collaris, and T. sphaerica), 1 species of Trichomitopsis (T. termopsidis), as well as smaller flagellates. We have re-assessed this community by sequencing the small subunit (SSU) rRNA gene and the internal transcribed spacer (ITS) region from a large number of single Trichonympha and Trichomitopsis cells for which morphology was also documented. Based on phylogenetic clustering and sequence divergence, we identify 3 new species: Trichonympha postcylindrica, Trichomitopsis minor, and Trichomitopsis parvus spp. nov. Once identified by sequencing, the morphology of the isolated cells for all 3 new species was re-examined and found to be distinct from the previously described species: Trichonympha postcylindrica can be morphologically distinguished from the other Trichonympha species by an extension on its posterior end, whereas Trichomitopsis minor and T. parvus are smaller than T. termopsidis but similar in size to each other and cannot be distinguished based on morphology using light microscopy. Given that Z. angusticollis has one of the best characterized hindgut communities, the near doubling of the number of the largest and most easily identifiable symbiont species suggests that the diversity of hindgut symbionts is substantially underestimated in other termites as well. Accurate descriptions of the diversity of these microbial communities are essential for understanding hindgut ecology and disentangling the interactions among the symbionts, and molecular barcoding should be a priority for these systems.

  10. 76 FR 9734 - Endangered and Threatened Species; Proposed Threatened Status for Distinct Population Segments of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-22

    ..., 2010 (75 FR 77476), we published a proposed rule to list the Beringia and Okhotsk Distinct Population... (February 8, 2011) was extended to March 25, 2011 (76 FR 6754; February 8, 2011). Public Hearings Joint... both the proposed rule for bearded seals and the proposed rule for ringed seals (75 FR 77476;...

  11. 78 FR 69310 - Endangered and Threatened Species; Protective Regulations for the Gulf of Maine Distinct...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-19

    ... Gulf of Maine Distinct Population Segment of Atlantic Sturgeon (76 FR 34023; June 10, 2011) contain a... two Federal Register notices published February 6, 2012 (77 FR 5880 and 77 FR 5914), we determined... listing the Gulf of Maine DPS as threatened and all of the other DPSs as endangered was warranted (77...

  12. Wild plant species growing closely connected in a subalpine meadow host distinct root-associated bacterial communities

    PubMed Central

    Aleklett, Kristin; Leff, Jonathan W.; Fierer, Noah

    2015-01-01

    Plant roots are known to harbor large and diverse communities of bacteria. It has been suggested that plant identity can structure these root-associated communities, but few studies have specifically assessed how the composition of root microbiota varies within and between plant species growing under natural conditions. We assessed the community composition of endophytic and epiphytic bacteria through high throughput sequencing using 16S rDNA derived from root tissues collected from a population of a wild, clonal plant (Orange hawkweed–Pilosella aurantiaca) as well as two neighboring plant species (Oxeye daisy–Leucanthemum vulgare and Alsike clover–Trifolium hybridum). Our first goal was to determine if plant species growing in close proximity, under similar environmental conditions, still hosted unique root microbiota. Our results showed that plants of different species host distinct bacterial communities in their roots. In terms of community composition, Betaproteobacteria (especially the family Oxalobacteraceae) were found to dominate in the root microbiota of L. vulgare and T. hybridum samples, whereas the root microbiota of P. aurantiaca had a more heterogeneous distribution of bacterial abundances where Gammaproteobacteria and Acidobacteria occupied a larger portion of the community. We also explored the extent of individual variance within each plant species investigated, and found that in the plant species thought to have the least genetic variance among individuals (P. aurantiaca) still hosted just as diverse microbial communities. Whether all plant species host their own distinct root microbiota and plants more closely related to each other share more similar bacterial communities still remains to be fully explored, but among the plants examined in this experiment there was no trend that the two species belonging to the same family shared more similarities in terms of bacterial community composition. PMID:25755932

  13. Gene Expression Variation in Duplicate Lactate dehydrogenase Genes: Do Ecological Species Show Distinct Responses?

    PubMed Central

    Cristescu, Melania E.; Demiri, Bora; Altshuler, Ianina; Crease, Teresa J.

    2014-01-01

    Lactate dehydrogenase (LDH) has been shown to play an important role in adaptation of several aquatic species to different habitats. The genomes of Daphnia pulex, a pond species, and Daphnia pulicaria, a lake inhabitant, encode two L-LDH enzymes, LDHA and LDHB. We estimated relative levels of Ldh gene expression in these two closely related species and their hybrids in four environmental settings, each characterized by one of two temperatures (10°C or 20°C), and one of two concentrations of dissolved oxygen (DO; 6.5–7 mg/l or 2–3 mg/l). We found that levels of LdhA expression were 4 to 48 times higher than LdhB expression (p<0.005) in all three groups (the two parental species and hybrids). Moreover, levels of LdhB expression differed significantly (p<0.05) between D. pulex and D. pulicaria, but neither species differed from the hybrid. Consistently higher expression of LdhA relative to LdhB in both species and the hybrid suggests that the two isozymes could be performing different functions. No significant differences in levels of gene expression were observed among the four combinations of temperature and dissolved oxygen (p>0.1). Given that Daphnia dwell in environments characterized by fluctuating conditions with long periods of low dissolved oxygen concentration, we suggest that these species could employ regulated metabolic depression to survive in such environments. PMID:25080082

  14. Distinct genetic subdivision in sympatric and sibling species of the genus Littorina (Gastropoda: Littorinidae).

    PubMed

    Rolán-Alvarez, E; Zapata, C; Alvarez, G

    1995-01-01

    The genetic structure of two sibling and sympatric species of the genus Littorina was compared using allozymic loci. The two species are biologically and ecologically well-known and mostly show similar life history characteristics. Three populations of L. mariae Sacchi & Rastelli and L. obstusata (L.) were studied in the Muros-Noya Ria (Galicia, NW Spain). In addition, four microgeographical subsamples taken from one of the populations were analysed for each species. Age, sex and genotypes for nine polymorphic loci were studied in 1250 snails of both species. L. mariae showed larger genetic population subdivision and lower heterozygosity levels for the loci studied than did L. obtusata. Heterozygote deficiencies were found in only a few cases in natural populations of both species, usually affecting the Lap-1 locus. No significant genetic differences among age or sex classes were found. These results may be explained by the lower effective population size in L. mariae than in L. obtusata. Known differences between these species in generation interval and population density during the winter can cause the different effective population sizes suggested. These life history characteristics appear to provide the most likely explanations for the differences in genetic differentiation and heterozygosity between the two species. A previously unknown L. mariae morph from exposed shores is tentatively suggested to be conspecific. PMID:7852095

  15. Crustose coralline algal species host distinct bacterial assemblages on their surfaces.

    PubMed

    Sneed, Jennifer M; Ritson-Williams, Raphael; Paul, Valerie J

    2015-11-01

    Crustose coralline algae (CCA) are important components of many marine ecosystems. They aid in reef accretion and stabilization, create habitat for other organisms, contribute to carbon sequestration and are important settlement substrata for a number of marine invertebrates. Despite their ecological importance, little is known about the bacterial communities associated with CCA or whether differences in bacterial assemblages may have ecological implications. This study examined the bacterial communities on four different species of CCA collected in Belize using bacterial tag-encoded FLX amplicon pyrosequencing of the V1-V3 region of the 16S rDNA. CCA were dominated by Alphaproteobacteria, Gammaproteobacteria and Actinomycetes. At the operational taxonomic unit (OTU) level, each CCA species had a unique bacterial community that was significantly different from all other CCA species. Hydrolithon boergesenii and Titanoderma prototypum, CCA species that facilitate larval settlement in multiple corals, had higher abundances of OTUs related to bacteria that inhibit the growth and/or biofilm formation of coral pathogens. Fewer coral larvae settle on the surfaces of Paragoniolithon solubile and Porolithon pachydermum. These CCA species had higher abundances of OTUs related to known coral pathogens and cyanobacteria. Coral larvae may be able to use the observed differences in bacterial community composition on CCA species to assess the suitability of these substrata for settlement and selectively settle on CCA species that contain beneficial bacteria. PMID:25918832

  16. Phylogenetic analysis of Primula section Primula reveals rampant non-monophyly among morphologically distinct species.

    PubMed

    Schmidt-Lebuhn, Alexander N; de Vos, Jurriaan M; Keller, Barbara; Conti, Elena

    2012-10-01

    The type section of Primula (Primulaceae), here considered to include seven species, is phylogenetically quite isolated in its genus. Although its species are popular ornamentals, traditional medicinal plants and model organisms for the study of heterostyly, the section has not yet been studied from a phylogenetic or evolutionary perspective. Using phylogenetic analysis of nuclear ITS and plastid data from all species and subspecies, we find that widespread Primula elatior is genetically heterogeneous and non-monophyletic to most if not all of the other ingroup taxa. The Genealogical Sorting Index (GSI) indicates that the assumption of all currently accepted species being independent lineages is consistent with the data. It is possible that P. elatior in its current circumscription may represents the disjointed remnant of an ancestral species from which the other recognized species diverged. However, based on available data, the alternative possibility of introgression explaining the non-monophyly of this species cannot be excluded. Species trees show P. elatior and P. veris as sister species. Primula vulgaris and P. juliae are closely related, while, in contrast to previous assumptions, P. renifolia does not appear to be a close relative of P. megaseifolia. With the section's isolation from the rest of the genus and very short internal branches, our dataset also presents a case study of the confounding effects of different branch length priors on the Bayesian estimation of resulting branch length estimates. Experimental runs using different priors confirm the problem of resulting estimates varying by orders of magnitude, while topology and relative branch lengths seem unaffected.

  17. Distinct Leishmania Species Infecting Wild Caviomorph Rodents (Rodentia: Hystricognathi) from Brazil

    PubMed Central

    Cássia-Pires, Renata; Boité, Mariana C.; D'Andrea, Paulo S.; Herrera, Heitor M.; Cupolillo, Elisa; Jansen, Ana Maria; Roque, André Luiz R.

    2014-01-01

    Background Caviomorph rodents, some of the oldest Leishmania spp. hosts, are widely dispersed in Brazil. Despite both experimental and field studies having suggested that these rodents are potential reservoirs of Leishmania parasites, not more than 88 specimens were analyzed in the few studies of natural infection. Our hypothesis was that caviomorph rodents are inserted in the transmission cycles of Leishmania in different regions, more so than is currently recognized. Methodology We investigated the Leishmania infection in spleen fragments of 373 caviomorph rodents from 20 different species collected in five Brazilian biomes in a period of 13 years. PCR reactions targeting kDNA of Leishmania sp. were used to diagnose infection, while Leishmania species identification was performed by DNA sequencing of the amplified products obtained in the HSP70 (234) targeting. Serology by IFAT was performed on the available serum of these rodents. Principal findings In 13 caviomorph rodents, DNA sequencing analyses allowed the identification of 4 species of the subgenus L. (Viannia): L. shawi, L. guyanensis, L. naiffi, and L. braziliensis; and 1 species of the subgenus L. (Leishmania): L. infantum. These include the description of parasite species in areas not previously included in their known distribution: L. shawi in Thrichomys inermis from Northeastern Brazil and L. naiffi in T. fosteri from Western Brazil. From the four other positive rodents, two were positive for HSP70 (234) targeting but did not generate sequences that enabled the species identification, and another two were positive only in kDNA targeting. Conclusions/Significance The infection rate demonstrated by the serology (51.3%) points out that the natural Leishmania infection in caviomorph rodents is much higher than that observed in the molecular diagnosis (4.6%), highlighting that, in terms of the host species responsible for maintaining Leishmania species in the wild, our current knowledge represents only the

  18. Analyses of mitochondrial amino acid sequence datasets support the proposal that specimens of Hypodontus macropi from three species of macropodid hosts represent distinct species

    PubMed Central

    2013-01-01

    Background Hypodontus macropi is a common intestinal nematode of a range of kangaroos and wallabies (macropodid marsupials). Based on previous multilocus enzyme electrophoresis (MEE) and nuclear ribosomal DNA sequence data sets, H. macropi has been proposed to be complex of species. To test this proposal using independent molecular data, we sequenced the whole mitochondrial (mt) genomes of individuals of H. macropi from three different species of hosts (Macropus robustus robustus, Thylogale billardierii and Macropus [Wallabia] bicolor) as well as that of Macropicola ocydromi (a related nematode), and undertook a comparative analysis of the amino acid sequence datasets derived from these genomes. Results The mt genomes sequenced by next-generation (454) technology from H. macropi from the three host species varied from 13,634 bp to 13,699 bp in size. Pairwise comparisons of the amino acid sequences predicted from these three mt genomes revealed differences of 5.8% to 18%. Phylogenetic analysis of the amino acid sequence data sets using Bayesian Inference (BI) showed that H. macropi from the three different host species formed distinct, well-supported clades. In addition, sliding window analysis of the mt genomes defined variable regions for future population genetic studies of H. macropi in different macropodid hosts and geographical regions around Australia. Conclusions The present analyses of inferred mt protein sequence datasets clearly supported the hypothesis that H. macropi from M. robustus robustus, M. bicolor and T. billardierii represent distinct species. PMID:24261823

  19. Subunit sequences of the 4 x 6-mer hemocyanin from the golden orb-web spider, Nephila inaurata.

    PubMed

    Averdam, Anne; Markl, Jürgen; Burmester, Thorsten

    2003-08-01

    The transport of oxygen in the hemolymph of many arthropod and mollusc species is mediated by large copper-proteins that are referred to as hemocyanins. Arthropod hemocyanins are composed of hexamers and oligomers of hexamers. Arachnid hemocyanins usually form 4 x 6-mers consisting of seven distinct subunit types (termed a-g), although in some spider taxa deviations from this standard scheme have been observed. Applying immunological and electrophoretic methods, six distinct hemocyanin subunits were identified in the red-legged golden orb-web spider Nephila inaurata madagascariensis (Araneae: Tetragnathidae). The complete cDNA sequences of six subunits were obtained that corresponded to a-, b-, d-, e-, f- and g-type subunits. No evidence for a c-type subunit was found in this species. The inclusion of the N. inaurata hemocyanins in a multiple alignment of the arthropod hemocyanins and the application of the Bayesian method of phylogenetic inference allow, for the first time, a solid reconstruction of the intramolecular evolution of the chelicerate hemocyanin subunits. The branch leading to subunit a diverged first, followed by the common branch of the dimer-forming b and c subunits, while subunits d and f, as well as subunits e and g form common branches. Assuming a clock-like evolution of the chelicerate hemocyanins, a timescale for the evolution of the Chelicerata was obtained that agrees with the fossil record.

  20. A distinct alleles and genetic recombination of pmrCAB operon in species of Acinetobacter baumannii complex isolates.

    PubMed

    Kim, Dae Hun; Ko, Kwan Soo

    2015-07-01

    To investigate pmrCAB sequence divergence in 5 species of Acinetobacter baumannii complex, a total of 80 isolates from a Korean hospital were explored. We evaluated nucleotide and amino acid polymorphisms of pmrCAB operon, and phylogenetic trees were constructed for each gene of prmCAB operon. Colistin and polymyxin B susceptibility was determined for all isolates, and multilocus sequence typing was also performed for A. baumannii isolates. Our results showed that each species of A. baumannii complex has divergent pmrCAB operon sequences. We identified a distinct pmrCAB allele allied with Acinetobacter nosocomialis in gene trees. Different grouping in each gene tree suggests sporadic recombination or emergence of pmrCAB genes among Acinetobacter species. Sequence polymorphisms among Acinetobacter species might not be associated with colistin resistance. We revealed that a distinct pmrCAB allele may be widespread across the continents such as North America and Asia and that sporadic genetic recombination or emergence of pmrCAB genes might occur.

  1. The discovery of the two types of small subunit ribosomal RNA gene in Eimeria mitis contests the existence of E. mivati as an independent species.

    PubMed

    Vrba, Vladimir; Poplstein, Martin; Pakandl, Michal

    2011-12-29

    Although the validity of the coccidian species, Eimeria mivati, has been questioned by many researchers for a long time there has not been any molecular analysis that would help resolve this issue. Here we report on the discovery of the two types of small ribosomal subunit (18S) gene within the Eimeria mitis genome that correspond to the known 18S sequences of E. mitis and E. mivati, and this is in conflict with the existence of E. mivati as an independent species. We have carried out five single oocyst isolations to obtain five single-oocyst-derived strains of E. mitis and these were analyzed by the sequencing of 18S and mitochondrial cytochrome c oxidase subunit I genes. The two types of 18S gene were found to be present in each strain in roughly equal ratios. This indicates that if the strains carrying only one or the other 18S type exist, they will likely cross-breed and still represent a single species. However, the more probable explanation is that all strains of E. mitis contain two types of 18S gene and that the occasional detection of only one or the other type by sequencing might be caused by insufficient sampling. This is also the first report of the two types of 18S gene in Eimeria, which has already been described in some other apicomplexan species, most notably Plasmodium. We also found that these two types of ribosomal RNA differ significantly in their secondary structure. The biological significance of the two 18S gene variants in E. mitis is not known, however, we hypothesize that these variants might be used in different stages of the parasite's life-cycle as it is in other apicomplexan species investigated so far.

  2. Are Phenacoccus solani Ferris and P. defectus Ferris (Hemiptera: Pseudococcidae) distinct species?

    PubMed

    Chatzidimitriou, Evangelia; Simonato, Mauro; Watson, Gillian W; Martinez-Sañudo, Isabel; Tanaka, Hirotaka; Zhao, Jing; Pellizzari, Giuseppina

    2016-01-01

    Among the Nearctic species of Phenacoccus (Hemiptera: Pseudococcidae), Phenacoccus solani Ferris and P. defectus Ferris are morphologically similar and it can be difficult to separate them on the basis of microscopic morphological characters of the adult female alone. In order to resolve their identity, a canonical variates morphological analysis of 199 specimens from different geographical origins and host plants and a molecular analysis of the COI and 28S genes were performed. The morphological analysis supported synonymy of the two species, as although the type specimens of the "species" are widely separated from each other in the canonical variates plot, they are all part of a continuous range of variation. The molecular analysis showed that P. solani and P. defectus are grouped in the same clade. On the basis of the morphological and molecular analyses, P. defectus is synonymized under the senior name P. solani, syn. n. PMID:27394512

  3. Are Phenacoccus solani Ferris and P. defectus Ferris (Hemiptera: Pseudococcidae) distinct species?

    PubMed

    Chatzidimitriou, Evangelia; Simonato, Mauro; Watson, Gillian W; Martinez-Sañudo, Isabel; Tanaka, Hirotaka; Zhao, Jing; Pellizzari, Giuseppina

    2016-03-24

    Among the Nearctic species of Phenacoccus (Hemiptera: Pseudococcidae), Phenacoccus solani Ferris and P. defectus Ferris are morphologically similar and it can be difficult to separate them on the basis of microscopic morphological characters of the adult female alone. In order to resolve their identity, a canonical variates morphological analysis of 199 specimens from different geographical origins and host plants and a molecular analysis of the COI and 28S genes were performed. The morphological analysis supported synonymy of the two species, as although the type specimens of the "species" are widely separated from each other in the canonical variates plot, they are all part of a continuous range of variation. The molecular analysis showed that P. solani and P. defectus are grouped in the same clade. On the basis of the morphological and molecular analyses, P. defectus is synonymized under the senior name P. solani, syn. n.

  4. Mercury's Exosphere During MESSENGER's Second Flyby: Detection of Magnesium and Distinct Distributions of Neutral Species

    NASA Technical Reports Server (NTRS)

    McClintock, William E.; Vervack, Ronald J., Jr.; Bradley, E. Todd; Killen, Rosemary M.; Mouawad, Nelly; Sprague, Ann L.; Burger, Matthew H.; Solomon, Sean C.; Izenberg, Noam R.

    2009-01-01

    During MESSENGER's second Mercury flyby, the Mercury Atmospheric and Surface Composition Spectrometer observed emission from Mercury's neutral exosphere. These observations include the first detection of emission from magnesium. Differing spatial distributions for sodium, calcium, and magnesium were revealed by observations beginning in Mercury's tail region, approximately 8 Mercury radii anti-sunward of the planet, continuing past the nightside, and ending near the dawn terminator. Analysis of these observations, supplemented by observations during the first Mercury flyby as well as those by other MESSENGER instruments, suggests that the distinct spatial distributions arise from a combination of differences in source, transfer, and loss processes.

  5. A Hepatozoon species genetically distinct from H. canis infecting spotted hyenas in the Serengeti ecosystem, Tanzania.

    PubMed

    East, Marion L; Wibbelt, Gudrun; Lieckfeldt, Dietmar; Ludwig, Arne; Goller, Katja; Wilhelm, Kerstin; Schares, Gereon; Thierer, Dagmar; Hofer, Heribert

    2008-01-01

    Health monitoring of spotted hyenas (Crocuta crocuta) in the Serengeti ecosystem, Tanzania, revealed Hepatozoon infection in all of 11 immature individuals examined following death from natural causes. Hepatozoon infection was probably an important factor contributing to mortality in two cases that exhibited clinical signs of ataxia, lethargy, ocular discharge, retching, and labored breathing before death. Whether Hepatozoon infection contributed to six deaths from fire, probable lion predation and unknown causes could not be determined. Four deaths from infanticide and starvation were unlikely to be associated with Hepatozoon infection. Histologic examination revealed lung tissue infected with cyst-like structures containing protozoan stages in all eight cases examined and interstitial pneumonia in most cases. Systemic spread of infection to several organs was found in three cases. Alignment of a 426 bp sequence from the parasite's 18s rRNA gene revealed a Hepatozoon species identical to that recently described from two domestic cats in Spain and only 7 bp substitutions when a 853 bp sequence was aligned to this cat Hepatozoon species. Previous reports of infection of wild carnivores in eastern and southern Africa with an unspecified Hepatozoon species similar in appearance to H. canis may have involved the species described in this study.

  6. Characterization of the Uukuniemi virus group (Phlebovirus: Bunyaviridae): evidence for seven distinct species.

    PubMed

    Palacios, Gustavo; Savji, Nazir; Travassos da Rosa, Amelia; Guzman, Hilda; Yu, Xuejie; Desai, Aaloki; Rosen, Gail Emilia; Hutchison, Stephen; Lipkin, W Ian; Tesh, Robert

    2013-03-01

    Evolutionary insights into the phleboviruses are limited because of an imprecise classification scheme based on partial nucleotide sequences and scattered antigenic relationships. In this report, the serologic and phylogenetic relationships of the Uukuniemi group viruses and their relationships with other recently characterized tick-borne phleboviruses are described using full-length genome sequences. We propose that the viruses currently included in the Uukuniemi virus group be assigned to five different species as follows: Uukuniemi virus, EgAn 1825-61 virus, Fin V707 virus, Chizé virus, and Zaliv Terpenia virus would be classified into the Uukuniemi species; Murre virus, RML-105-105355 virus, and Sunday Canyon virus would be classified into a Murre virus species; and Grand Arbaud virus, Precarious Point virus, and Manawa virus would each be given individual species status. Although limited sequence similarity was detected between current members of the Uukuniemi group and Severe fever with thrombocytopenia syndrome virus (SFTSV) and Heartland virus, a clear serological reaction was observed between some of them, indicating that SFTSV and Heartland virus should be considered part of the Uukuniemi virus group. Moreover, based on the genomic diversity of the phleboviruses and given the low correlation observed between complement fixation titers and genetic distance, we propose a system for classification of the Bunyaviridae based on genetic as well as serological data. Finally, the recent descriptions of SFTSV and Heartland virus also indicate that the public health importance of the Uukuniemi group viruses must be reevaluated. PMID:23283959

  7. Characterization of the Uukuniemi Virus Group (Phlebovirus: Bunyaviridae): Evidence for Seven Distinct Species

    PubMed Central

    Savji, Nazir; Travassos da Rosa, Amelia; Guzman, Hilda; Yu, Xuejie; Desai, Aaloki; Rosen, Gail Emilia; Hutchison, Stephen; Lipkin, W. Ian; Tesh, Robert

    2013-01-01

    Evolutionary insights into the phleboviruses are limited because of an imprecise classification scheme based on partial nucleotide sequences and scattered antigenic relationships. In this report, the serologic and phylogenetic relationships of the Uukuniemi group viruses and their relationships with other recently characterized tick-borne phleboviruses are described using full-length genome sequences. We propose that the viruses currently included in the Uukuniemi virus group be assigned to five different species as follows: Uukuniemi virus, EgAn 1825-61 virus, Fin V707 virus, Chizé virus, and Zaliv Terpenia virus would be classified into the Uukuniemi species; Murre virus, RML-105-105355 virus, and Sunday Canyon virus would be classified into a Murre virus species; and Grand Arbaud virus, Precarious Point virus, and Manawa virus would each be given individual species status. Although limited sequence similarity was detected between current members of the Uukuniemi group and Severe fever with thrombocytopenia syndrome virus (SFTSV) and Heartland virus, a clear serological reaction was observed between some of them, indicating that SFTSV and Heartland virus should be considered part of the Uukuniemi virus group. Moreover, based on the genomic diversity of the phleboviruses and given the low correlation observed between complement fixation titers and genetic distance, we propose a system for classification of the Bunyaviridae based on genetic as well as serological data. Finally, the recent descriptions of SFTSV and Heartland virus also indicate that the public health importance of the Uukuniemi group viruses must be reevaluated. PMID:23283959

  8. 75 FR 12597 - Endangered and Threatened Species; Proposed Listing of Nine Distinct Population Segments of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-16

    ... loggerhead sea turtle as threatened throughout its worldwide range on July 28, 1978 (43 FR 32800). On July 12... endangered species under the ESA. NMFS published a notice in the Federal Register on November 16, 2007 (72 FR... Federal Register on March 5, 2008 (73 FR 11849), concluding that the petitioners (Center for...

  9. Use of Cytochrome c Oxidase Subunit I (COI) Nucleotide Sequences for Identification of the Korean Luciliinae Fly Species (Diptera: Calliphoridae) in Forensic Investigations

    PubMed Central

    Park, Seong Hwan; Zhang, Yong; Piao, Huguo; Yu, Dong Ha; Jeong, Hyun Ju; Yoo, Ga Young; Chung, Ukhee; Jo, Tae-Ho

    2009-01-01

    Blowflies, especially species belonging to the subfamily Luciliinae, are the first insects to lay eggs on corpses in Korea. Fast and accurate species identification has been a key task for forensic entomologists. Because conventional morphologic identification methods have many limitations with respect to forensic practice, molecular methods have been proposed to identify fly species of forensic importance. To this end, the authors amplified and sequenced the full length of the cytochrome c oxidase subunit I (COI) gene of the Luciliinae fly species collected in Korea. The results showed the COI sequences are instrumental in identifying Luciliinae fly species. However, when compared with previously reported data, considerable inconsistencies were noted. Hemipyrellia ligurriens data in this study differed significantly from two of the five pre-existing data. Two closely related species, Lucilia illustris and Lucilia caesar, showed an overlap of COI haplotypes due to four European sequences. The results suggest that more individuals from various geographic regions and additive nuclear DNA markers should be analyzed, and morphologic identification keys must be reconfirmed to overcome these inconsistencies. PMID:19949660

  10. Use of cytochrome c oxidase subunit i (COI) nucleotide sequences for identification of the Korean Luciliinae fly species (Diptera: Calliphoridae) in forensic investigations.

    PubMed

    Park, Seong Hwan; Zhang, Yong; Piao, Huguo; Yu, Dong Ha; Jeong, Hyun Ju; Yoo, Ga Young; Chung, Ukhee; Jo, Tae-Ho; Hwang, Juck-Joon

    2009-12-01

    Blowflies, especially species belonging to the subfamily Luciliinae, are the first insects to lay eggs on corpses in Korea. Fast and accurate species identification has been a key task for forensic entomologists. Because conventional morphologic identification methods have many limitations with respect to forensic practice, molecular methods have been proposed to identify fly species of forensic importance. To this end, the authors amplified and sequenced the full length of the cytochrome c oxidase subunit I (COI) gene of the Luciliinae fly species collected in Korea. The results showed the COI sequences are instrumental in identifying Luciliinae fly species. However, when compared with previously reported data, considerable inconsistencies were noted. Hemipyrellia ligurriens data in this study differed significantly from two of the five pre-existing data. Two closely related species, Lucilia illustris and Lucilia caesar, showed an overlap of COI haplotypes due to four European sequences. The results suggest that more individuals from various geographic regions and additive nuclear DNA markers should be analyzed, and morphologic identification keys must be reconfirmed to overcome these inconsistencies.

  11. Phylogenetic distinction of iNOS and IDO function in mesenchymal stem cell-mediated immunosuppression in mammalian species.

    PubMed

    Su, J; Chen, X; Huang, Y; Li, W; Li, J; Cao, K; Cao, G; Zhang, L; Li, F; Roberts, A I; Kang, H; Yu, P; Ren, G; Ji, W; Wang, Y; Shi, Y

    2014-03-01

    Mammalian mesenchymal stem cells (MSCs) have been shown to be strongly immunosuppressive in both animal disease models and human clinical trials. We have reported that the key molecule mediating immunosuppression by MSCs is species dependent: indoleamine 2,3-dioxygenase (IDO) in human and inducible nitric oxide synthase (iNOS) in mouse. In the present study, we isolated MSCs from several mammalian species, each of a different genus, and investigated the involvement of IDO and iNOS during MSC-mediated immunosuppression. The characterization of MSCs from different species was by adherence to tissue culture plastic, morphology, specific marker expression, and differentiation potential. On the basis of the inducibility of IDO and iNOS by inflammatory cytokines in MSCs, the tested mammalian species fall into two distinct groups: IDO utilizers and iNOS utilizers. MSCs from monkey, pig, and human employ IDO to suppress immune responses, whereas MSCs from mouse, rat, rabbit, and hamster utilize iNOS. Interestingly, based on the limited number of species tested, the iNOS-utilizing species all belong to the phylogenetic clade, Glires. Although the evolutionary significance of this divergence is not known, we believe that this study provides critical guidance for choosing appropriate animal models for preclinical studies of MSCs.

  12. Sink-source characteristics of two distinctly different forest species as affected by elevated carbon dioxide

    SciTech Connect

    Pushnik, J.C.; Florv, W.B.; Demaree, R.S. ); Anderson, P.D.; Houpis J.L.J. )

    1993-05-01

    The basic physiology and biochemistry of photosynthesis is being correlated with the leaf level processes and morphology of the Sierra Nevada varieties of Taxus brevifolia and Pinus ponderosa in an attempt to identify control mechanisms of carbohydrate partitioning. We are evaluating sink/source relationships in terms of carbon assimilation (gas-exchange (A[ci] curves and temperature effects); RuBPCase activity, chloroplast structure, integrity, and distributions, stomatal densities, internal leaf organization); transport functions (sucrose-phosphate synthetase (SPS) activity); long-term sink (immunoelectron microscopic detection of taxol). The results of these investigations suggest carbon acquisition characteristics are similar among the conifers, but with distinct differences in carboxylation efficiencies, SPS activity, needle starch content/chloroplast, and vascular tissue areas. These baseline characteristics are currently being evaluated in response to elevated CO[sub 2].

  13. Evidence for a species of nuclear actin distinct from cytoplasmic and muscles actins.

    PubMed

    Bremer, J W; Busch, H; Yeoman, L C

    1981-03-31

    Nuclear actin (protein BJ) has been isolated from the chromatin of Novikoff hepatoma ascites cells and purified to homogeneity by selective extraction, Sepharose CL-6B chromatography, and preparative polyacrylamide gel electrophoresis. A comparison of nuclear and cytoplasmic actins from Novikoff hepatoma cells and rabbit muscle actin was made by amino acid analysis, isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and two-dimensional peptide mapping procedures. By these criteria, all of the proteins compared are actins, but each is chemically distinct. It was concluded, therefore, that nuclear actin is similar to, but not identical with, cytoplasmic actin isolated from Novikoff hepatoma cells. A striking similarity in peptide charge and migration as shown by peptide map analysis was observed for nuclear and rabbit skeletal muscle actins. This may indicate that nuclear actin has the capacity for contractile function. In addition, the actins synthesized in Novikoff hepatoma cells may results from more than two structural genes.

  14. Lignin extraction distinctively enhances biomass enzymatic saccharification in hemicelluloses-rich Miscanthus species under various alkali and acid pretreatments.

    PubMed

    Si, Shengli; Chen, Yan; Fan, Chunfen; Hu, Huizhen; Li, Ying; Huang, Jiangfeng; Liao, Haofeng; Hao, Bo; Li, Qing; Peng, Liangcai; Tu, Yuanyuan

    2015-05-01

    In this study, one- and two-step pretreatments with alkali and acid were performed in the three Miscanthus species that exhibit distinct hemicelluloses levels. As a result, one-step with 4% NaOH or two-step with 2% NaOH and 1% H2SO4 was examined to be optimal for high biomass saccharification, indicating that alkali was the main effecter of pretreatments. Notably, both one- and two-step pretreatments largely enhanced biomass digestibility distinctive in hemicelluloses-rich samples by effectively co-extracting hemicelluloses and lignin. However, correlation analysis further indicated that the effective lignin extraction, other than the hemicelluloses removals, predominately determined biomass saccharification under various alkali and acid pretreatments, leading to a significant alteration of cellulose crystallinity. Hence, this study has suggested the potential approaches in bioenergy crop breeding and biomass process technology.

  15. Nucleotide sequence and genome organization of a member of a new and distinct Caulimovirus species from dahlia.

    PubMed

    Pappu, H R; Druffel, K L; Miglino, R; van Schadewijk, A R

    2008-01-01

    A distinct caulimovirus, associated with dahlia mosaic, was cloned and sequenced. The caulimovirus, tentatively designated as dahlia common mosaic virus (DCMV), had a double-stranded DNA genome of ca. 8 kb. The genome organization of DCMV was found to be typical of members of the genus Caulimovirus and consisted of six major open reading frames (ORFs), ORFs I-VI, and one minor ORF, ORF VII. Sequence comparisons with the DNA genomes of two known caulimoviruses isolated from dahlia, Dahlia mosaic virus (DMV) and an endogenous caulimovirus, DMV-D10, showed that DCMV is a member of a distinct caulimovirus species, with sequence identities among various ORFs ranging from 25 to 80%. PMID:18974923

  16. Simultaneous identification and DNA barcoding of six Eimeria species infecting turkeys using PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus.

    PubMed

    Hafeez, Mian A; Shivaramaiah, Srichaitanya; Dorsey, Kristi Moore; Ogedengbe, Mosun E; El-Sherry, Shiem; Whale, Julia; Cobean, Julie; Barta, John R

    2015-05-01

    Species-specific PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus were generated that allow for the specific identification of the most common Eimeria species infecting turkeys (i.e., Eimeria adenoeides, Eimeria meleagrimitis, Eimeria gallopavonis, Eimeria meleagridis, Eimeria dispersa, and Eimeria innocua). PCR reaction chemistries were optimized with respect to divalent cation (MgCl2) and dNTP concentrations, as well as PCR cycling conditions (particularly anneal temperature for primers). Genomic DNA samples from single oocyst-derived lines of six Eimeria species were tested to establish specificity and sensitivity of these newly designed primer pairs. A mixed 60-ng total DNA sample containing 10 ng of each of the six Eimeria species was used as DNA template to demonstrate specific amplification of the correct product using each of the species-specific primer pairs. Ten nanograms of each of the five non-target Eimeria species was pooled to provide a non-target, control DNA sample suitable to test the specificity of each primer pair. The amplifications of the COI region with species-specific primer pairs from pooled samples yielded products of expected sizes (209 to 1,012 bp) and no amplification of non-target Eimeria sp. DNA was detected using the non-target, control DNA samples. These primer pairs specific for Eimeria spp. of turkeys did not amplify any of the seven Eimeria species infecting chickens. The newly developed PCR primers can be used as a diagnostic tool capable of specifically identifying six turkey Eimeria species; additionally, sequencing of the PCR amplification products yields sequence-based genotyping data suitable for identification and molecular phylogenetics.

  17. Two Rumex Species from Contrasting Hydrological Niches Regulate Flooding Tolerance through Distinct Mechanisms[C][W][OPEN

    PubMed Central

    van Veen, Hans; Mustroph, Angelika; Barding, Gregory A.; Vergeer-van Eijk, Marleen; Welschen-Evertman, Rob A.M.; Pedersen, Ole; Visser, Eric J.W.; Larive, Cynthia K.; Pierik, Ronald; Bailey-Serres, Julia; Voesenek, Laurentius A.C.J.; Sasidharan, Rashmi

    2013-01-01

    Global climate change has increased flooding events, which affect both natural vegetation dynamics and crop productivity. The flooded environment is lethal for most plant species because it restricts gas exchange and induces an energy and carbon crisis. Flooding survival strategies have been studied in Oryza sativa, a cultivated monocot. However, our understanding of plant adaptation to natural flood-prone environments remains scant, even though wild plants represent a valuable resource of tolerance mechanisms that could be used to generate stress-tolerant crops. Here we identify mechanisms that mediate the distinct flooding survival strategies of two related wild dicot species: Rumex palustris and Rumex acetosa. Whole transcriptome sequencing and metabolite profiling reveal flooding-induced metabolic reprogramming specific to R. acetosa. By contrast, R. palustris uses the early flooding signal ethylene to increase survival by regulating shade avoidance and photomorphogenesis genes to outgrow submergence and by priming submerged plants for future low oxygen stress. These results provide molecular resolution of flooding survival strategies of two species occupying distinct hydrological niches. Learning how these contrasting flood adaptive strategies evolved in nature will be instrumental for the development of stress-tolerant crop varieties that deliver enhanced yields in a changing climate. PMID:24285788

  18. Differential Costs of Two Distinct Resistance Mechanisms Induced by Different Herbivore Species in Arabidopsis.

    PubMed

    Onkokesung, Nawaporn; Reichelt, Michael; van Doorn, Arjen; Schuurink, Robert C; Dicke, Marcel

    2016-02-01

    Plants respond to herbivory with the induction of resistance, mediated by distinct phytohormonal signaling pathways and their interactions. Phloem feeders are known to induce plant resistance via the salicylic acid pathway, whereas biting-chewing herbivores induce plant resistance mainly via the jasmonate pathway. Here, we show that a specialist caterpillar (biting-chewing herbivore) and a specialist aphid (phloem feeder) differentially induce resistance against Pieris brassicae caterpillars in Arabidopsis (Arabidopsis thaliana) plants. Caterpillar feeding induces resistance through the jasmonate signaling pathway that is associated with the induction of kaempferol 3,7-dirhamnoside, whereas aphid feeding induces resistance via a novel mechanism involving sinapoyl malate. The role of sinapoyl malate is confirmed through the use of a mutant compromised in the biosynthesis of this compound. Caterpillar-induced resistance is associated with a lower cost in terms of plant growth reduction than aphid-induced resistance. A strong constitutive resistance against P. brassicae caterpillars in combination with a strong growth attenuation in plants of a transfer DNA (T-DNA) insertion mutant of WRKY70 (wrky70) suggest that the WRKY70 transcription factor, a regulator of downstream responses mediated by jasmonate-salicylic acid signaling cross talk, is involved in the negative regulation of caterpillar resistance and in the tradeoff between growth and defense. In conclusion, different mechanisms of herbivore-induced resistance come with different costs, and a functional WRKY70 transcription factor is required for the induction of low-cost resistance. PMID:26603653

  19. Distinctive features of dinoflagellate chromatin. Absence of nucleosomes in a primitive species Prorocentrum micans E.

    PubMed

    Herzog, M; Soyer, M O

    1981-02-01

    The absence of nucleosome-like structures from purified nuclei of the primitive dinoflagellate Prorocentrum micans was demonstrated by three means. i) Electron microscopy revealed mostly thin, smooth 6-nm nucleofilaments in chromatin incubated at various ionic strengths and either fixed in 0.1% glutaraldehyde or unfixed. No "beads-on-a-string" structure was found. ii) Analysis of nuclear proteins showed that low amounts of basic proteins were present (basis proteins: DNA less than 0.1), the two major one with molecular weights 12 000 and 13 000 and that histones characteristic of eucaryotes were absent. iii) Digestion of the nuclei with micrococcal endonuclease of DNase I did not result in partially digested DNA fragment repeats. Only about 10% of the bulk of the nuclear DNA was digested by micrococcal endonuclease. The high molecular weight of the remainder suggests particular protection against this type of nuclease. In the light of these distinctive nuclear features, we discuss the evolutionary position of the dinoflagellate protists with respect to the procaryotes and eucaryotes.

  20. Thioredoxin redox regulates ATPase activity of magnesium chelatase CHLI subunit and modulates redox-mediated signaling in tetrapyrrole biosynthesis and homeostasis of reactive oxygen species in pea plants.

    PubMed

    Luo, Tao; Fan, Tingting; Liu, Yinan; Rothbart, Maxi; Yu, Jing; Zhou, Shuaixiang; Grimm, Bernhard; Luo, Meizhong

    2012-05-01

    The chloroplast thioredoxins (TRXs) function as messengers of redox signals from ferredoxin to target enzymes. In this work, we studied the regulatory impact of pea (Pisum sativum) TRX-F on the magnesium (Mg) chelatase CHLI subunit and the enzymatic activation of Mg chelatase in vitro and in vivo. In vitro, reduced TRX-F activated the ATPase activity of pea CHLI and enhanced the activity of Mg chelatase reconstituted from the three recombinant subunits CHLI, CHLD, and CHLH in combination with the regulator protein GENOMES UNCOUPLED4 (GUN4). Yeast two-hybrid and bimolecular fluorescence complementation assays demonstrated that TRX-F physically interacts with CHLI but not with either of the other two subunits or GUN4. In vivo, virus-induced TRX-F gene silencing (VIGS-TRX-F) in pea plants did not result in an altered redox state of CHLI. However, simultaneous silencing of the pea TRX-F and TRX-M genes (VIGS-TRX-F/TRX-M) resulted in partially and fully oxidized CHLI in vivo. VIGS-TRX-F/TRX-M plants demonstrated a significant reduction in Mg chelatase activity and 5-aminolevulinic acid synthesizing capacity as well as reduced pigment content and lower photosynthetic capacity. These results suggest that, in vivo, TRX-M can compensate for a lack of TRX-F and that both TRXs act as important redox regulators of Mg chelatase. Furthermore, the silencing of TRX-F and TRX-M expression also affects gene expression in the tetrapyrrole biosynthesis pathway and leads to the accumulation of reactive oxygen species, which may also serve as an additional signal for the transcriptional regulation of photosynthesis-associated nuclear genes.

  1. Phylogenetic Reassessment of Antarctic Tetillidae (Demospongiae, Tetractinellida) Reveals New Genera and Genetic Similarity among Morphologically Distinct Species.

    PubMed

    Carella, Mirco; Agell, Gemma; Cárdenas, Paco; Uriz, Maria J

    2016-01-01

    Species of Tetillidae are distributed worldwide. However, some genera are unresolved and only a few genera and species of this family have been described from the Antarctic. The incorporation of 25 new COI and 18S sequences of Antarctic Tetillidae to those used recently for assessing the genera phylogeny, has allowed us to improve the resolution of some poorly resolved nodes and to confirm the monophyly of previously identified clades. Classical genera such as Craniella recovered their traditional diagnosis by moving the Antarctic Tetilla from Craniella, where they were placed in the previous family phylogeny, to Antarctotetilla gen. nov. The morphological re-examination of specimens used in the previous phylogeny and their comparison to the type material revealed misidentifications. The proposed monotypic new genus Levantinella had uncertain phylogenetic relationships depending on the gene partition used. Two more clades would require the inclusion of additional species to be formally established as new genera. The parsimony tree based on morphological characters and the secondary structure of the 18S (V4 region) almost completely matched the COI M1-M6 and the COI+18S concatenated phylogenies. Morphological synapomorphies have been identified for the genera proposed. New 15 28S (D3-D5) and 11 COI I3-M11 partitions were exclusively sequenced for the Antarctic species subset. Remarkably, species within the Antarctic genera Cinachyra (C. barbata and C. antarctica) and Antarctotetilla (A. leptoderma, A. grandis, and A. sagitta), which are clearly distinguishable morphologically, were not genetically differentiated with any of the markers assayed. Thus, as it has been reported for other Antarctic sponges, both the mitochondrial and nuclear partitions used did not differentiate species that were well characterized morphologically. Antarctic Tetillidae offers a rare example of genetically cryptic (with the traditional markers used for sponges), morphologically distinct

  2. Phylogenetic Reassessment of Antarctic Tetillidae (Demospongiae, Tetractinellida) Reveals New Genera and Genetic Similarity among Morphologically Distinct Species

    PubMed Central

    Carella, Mirco; Agell, Gemma; Cárdenas, Paco; Uriz, Maria J.

    2016-01-01

    Species of Tetillidae are distributed worldwide. However, some genera are unresolved and only a few genera and species of this family have been described from the Antarctic. The incorporation of 25 new COI and 18S sequences of Antarctic Tetillidae to those used recently for assessing the genera phylogeny, has allowed us to improve the resolution of some poorly resolved nodes and to confirm the monophyly of previously identified clades. Classical genera such as Craniella recovered their traditional diagnosis by moving the Antarctic Tetilla from Craniella, where they were placed in the previous family phylogeny, to Antarctotetilla gen. nov. The morphological re-examination of specimens used in the previous phylogeny and their comparison to the type material revealed misidentifications. The proposed monotypic new genus Levantinella had uncertain phylogenetic relationships depending on the gene partition used. Two more clades would require the inclusion of additional species to be formally established as new genera. The parsimony tree based on morphological characters and the secondary structure of the 18S (V4 region) almost completely matched the COI M1-M6 and the COI+18S concatenated phylogenies. Morphological synapomorphies have been identified for the genera proposed. New 15 28S (D3-D5) and 11 COI I3-M11 partitions were exclusively sequenced for the Antarctic species subset. Remarkably, species within the Antarctic genera Cinachyra (C. barbata and C. antarctica) and Antarctotetilla (A. leptoderma, A. grandis, and A. sagitta), which are clearly distinguishable morphologically, were not genetically differentiated with any of the markers assayed. Thus, as it has been reported for other Antarctic sponges, both the mitochondrial and nuclear partitions used did not differentiate species that were well characterized morphologically. Antarctic Tetillidae offers a rare example of genetically cryptic (with the traditional markers used for sponges), morphologically distinct

  3. Phylogenetic Reassessment of Antarctic Tetillidae (Demospongiae, Tetractinellida) Reveals New Genera and Genetic Similarity among Morphologically Distinct Species.

    PubMed

    Carella, Mirco; Agell, Gemma; Cárdenas, Paco; Uriz, Maria J

    2016-01-01

    Species of Tetillidae are distributed worldwide. However, some genera are unresolved and only a few genera and species of this family have been described from the Antarctic. The incorporation of 25 new COI and 18S sequences of Antarctic Tetillidae to those used recently for assessing the genera phylogeny, has allowed us to improve the resolution of some poorly resolved nodes and to confirm the monophyly of previously identified clades. Classical genera such as Craniella recovered their traditional diagnosis by moving the Antarctic Tetilla from Craniella, where they were placed in the previous family phylogeny, to Antarctotetilla gen. nov. The morphological re-examination of specimens used in the previous phylogeny and their comparison to the type material revealed misidentifications. The proposed monotypic new genus Levantinella had uncertain phylogenetic relationships depending on the gene partition used. Two more clades would require the inclusion of additional species to be formally established as new genera. The parsimony tree based on morphological characters and the secondary structure of the 18S (V4 region) almost completely matched the COI M1-M6 and the COI+18S concatenated phylogenies. Morphological synapomorphies have been identified for the genera proposed. New 15 28S (D3-D5) and 11 COI I3-M11 partitions were exclusively sequenced for the Antarctic species subset. Remarkably, species within the Antarctic genera Cinachyra (C. barbata and C. antarctica) and Antarctotetilla (A. leptoderma, A. grandis, and A. sagitta), which are clearly distinguishable morphologically, were not genetically differentiated with any of the markers assayed. Thus, as it has been reported for other Antarctic sponges, both the mitochondrial and nuclear partitions used did not differentiate species that were well characterized morphologically. Antarctic Tetillidae offers a rare example of genetically cryptic (with the traditional markers used for sponges), morphologically distinct

  4. The genome of African yam (Dioscorea cayenensis-rotundata complex) hosts endogenous sequences from four distinct Badnavirus species.

    PubMed

    Umber, Marie; Filloux, Denis; Muller, Emmanuelle; Laboureau, Nathalie; Galzi, Serge; Roumagnac, Philippe; Iskra-Caruana, Marie-Line; Pavis, Claudie; Teycheney, Pierre-Yves; Seal, Susan E

    2014-10-01

    Several endogenous viral elements (EVEs) have been identified in plant genomes, including endogenous pararetroviruses (EPRVs). Here, we report the first characterization of EPRV sequences in the genome of African yam of the Dioscorea cayenensis-rotundata complex. We propose that these sequences should be termed 'endogenous Dioscorea bacilliform viruses' (eDBVs). Molecular characterization of eDBVs shows that they constitute sequences originating from various parts of badnavirus genomes, resulting in a mosaic structure that is typical of most EPRVs characterized to date. Using complementary molecular approaches, we show that eDBVs belong to at least four distinct Badnavirus species, indicating multiple, independent, endogenization events. Phylogenetic analyses of eDBVs support and enrich the current taxonomy of yam badnaviruses and lead to the characterization of a new Badnavirus species in yam. The impact of eDBVs on diagnosis, yam germplasm conservation and movement, and breeding is discussed.

  5. Distinct and Conserved Prominin-1/CD133–Positive Retinal Cell Populations Identified across Species

    PubMed Central

    Jászai, József; Fargeas, Christine A.; Graupner, Sylvi; Tanaka, Elly M.; Brand, Michael; Huttner, Wieland B.; Corbeil, Denis

    2011-01-01

    Besides being a marker of various somatic stem cells in mammals, prominin-1 (CD133) plays a role in maintaining the photoreceptor integrity since mutations in the PROM1 gene are linked with retinal degeneration. In spite of that, little information is available regarding its distribution in eyes of non-mammalian vertebrates endowed with high regenerative abilities. To address this subject, prominin-1 cognates were isolated from axolotl, zebrafish and chicken, and their retinal compartmentalization was investigated and compared to that of their mammalian orthologue. Interestingly, prominin-1 transcripts—except for the axolotl—were not strictly restricted to the outer nuclear layer (i.e., photoreceptor cells), but they also marked distinct subdivisions of the inner nuclear layer (INL). In zebrafish, where the prominin-1 gene is duplicated (i.e., prominin-1a and prominin-1b), a differential expression was noted for both paralogues within the INL being localized either to its vitreal or scleral subdivision, respectively. Interestingly, expression of prominin-1a within the former domain coincided with Pax-6–positive cells that are known to act as progenitors upon injury-induced retino-neurogenesis. A similar, but minute population of prominin-1–positive cells located at the vitreal side of the INL was also detected in developing and adult mice. In chicken, however, prominin-1–positive cells appeared to be aligned along the scleral side of the INL reminiscent of zebrafish prominin-1b. Taken together our data indicate that in addition to conserved expression of prominin-1 in photoreceptors, significant prominin-1–expressing non-photoreceptor retinal cell populations are present in the vertebrate eye that might represent potential sources of stem/progenitor cells for regenerative therapies. PMID:21407811

  6. Meteorological influences on stemflow generation across diameter size classes of two morphologically distinct deciduous species.

    PubMed

    Van Stan, John T; Van Stan, Jarrad H; Levia, Delphis F

    2014-12-01

    Many tree species have been shown to funnel substantial rainfall to their stem base as stemflow flux, given a favorable stand structure and storm conditions. As stemflow is a spatially concentrated flux, prior studies have shown its impact on ecohydrological and biogeochemical processes can be significant. Less work has been performed examining stemflow variability from meteorological conditions compared to canopy structural traits. As such, this study performs multiple regressions: (1) to examine stemflow variability due to event-based rainfall amount, intensity, mean wind speeds, and vapor pressure deficit; (2) across three diameter size classes (10-20, 21-40, and >41 cm DBH); and (3) for two common tree species in the northeastern USA of contrasting canopy morphology--Liriodendron tulipifera L. (yellow poplar) versus Fagus grandifolia Ehrh. (American beech). On the whole, multiple regression results yielded significant positive correlations with stemflow for rainfall amount, intensity, and mean wind speed and a significant negative correlation for vapor pressure deficit (VPD). Tree size altered stemflow-meteorological condition relationships, where larger trees strengthened indirect stemflow-VPD and direct stemflow-rainfall and stemflow-intensity associations. Canopies of rougher bark and lower branch angle (represented by L. tulipifera) enhanced correlations for nearly all meteorological conditions via greater stemflow residence time (and longer exposure to meteorological conditions). Multiple regressions performed on leafless canopy stemflow resulted in an inverse relationship with wind speeds, likely decoupling stemflow sheltered solely on bark surfaces from VPD influences. Leaf presence generally increased direct stemflow associations with rainfall intensity, yet diminished stemflow-rainfall relationships. F. grandifolia canopies (exemplifying structures of smoother bark and greater branch angle) strengthened differences in stemflow associations with

  7. Evidence of Distinct Populations and Specific Subpopulations within the Species Oenococcus oeni▿

    PubMed Central

    Bridier, Julen; Claisse, Olivier; Coton, Monika; Coton, Emmanuel; Lonvaud-Funel, Aline

    2010-01-01

    Among the lactic acid bacteria (LAB) present in the oenological microbial ecosystem, Oenococcus oeni, an acidophilic lactic acid bacterium, is essential during winemaking. It outclasses all other bacterial species during malolactic fermentation (MLF). Oenological performances, such as malic acid degradation rate and sensorial impact, vary significantly according to the strain. The genetic diversity of the O. oeni species was evaluated using a multilocus sequence typing (MLST) scheme. Seven housekeeping genes were sequenced for a collection of 258 strains that had been isolated all over the world (particularly Burgundy, Champagne, and Aquitaine, France, Chile, South Africa, and Italy) and in several wine types (red wines, white wines, and champagne) and cider. The allelic diversity was high, with an average of 20.7 alleles per locus, many of them being rare alleles. The collection comprised 127 sequence types, suggesting an important genotypic diversity. The neighbor-joining phylogenetic tree constructed from the concatenated sequence of the seven housekeeping genes showed two major phylogenetic groups, named A and B. One unique strain isolated from cider composed a third group, rooting the phylogenetic tree. However, all other strains isolated from cider were in group B. Eight phylogenetic subgroups were statistically differentiated and could be delineated by the analysis of only 32 mutations instead of the 600 mutations observed in the concatenated sequence of the seven housekeeping genes. Interestingly, in group A, several phylogenetic subgroups were composed mostly of strains coming from a precise geographic origin. Three subgroups were identified, composed of strains from Chile, South Africa, and eastern France. PMID:20935119

  8. Meteorological influences on stemflow generation across diameter size classes of two morphologically distinct deciduous species

    NASA Astrophysics Data System (ADS)

    Van Stan, John T.; Van Stan, Jarrad H.; Levia, Delphis F.

    2014-12-01

    Many tree species have been shown to funnel substantial rainfall to their stem base as stemflow flux, given a favorable stand structure and storm conditions. As stemflow is a spatially concentrated flux, prior studies have shown its impact on ecohydrological and biogeochemical processes can be significant. Less work has been performed examining stemflow variability from meteorological conditions compared to canopy structural traits. As such, this study performs multiple regressions: (1) to examine stemflow variability due to event-based rainfall amount, intensity, mean wind speeds, and vapor pressure deficit; (2) across three diameter size classes (10-20, 21-40, and >41 cm DBH); and (3) for two common tree species in the northeastern USA of contrasting canopy morphology— Liriodendron tulipifera L. (yellow poplar) versus Fagus grandifolia Ehrh. (American beech). On the whole, multiple regression results yielded significant positive correlations with stemflow for rainfall amount, intensity, and mean wind speed and a significant negative correlation for vapor pressure deficit (VPD). Tree size altered stemflow-meteorological condition relationships, where larger trees strengthened indirect stemflow-VPD and direct stemflow-rainfall and stemflow-intensity associations. Canopies of rougher bark and lower branch angle (represented by L. tulipifera) enhanced correlations for nearly all meteorological conditions via greater stemflow residence time (and longer exposure to meteorological conditions). Multiple regressions performed on leafless canopy stemflow resulted in an inverse relationship with wind speeds, likely decoupling stemflow sheltered solely on bark surfaces from VPD influences. Leaf presence generally increased direct stemflow associations with rainfall intensity, yet diminished stemflow-rainfall relationships. F. grandifolia canopies (exemplifying structures of smoother bark and greater branch angle) strengthened differences in stemflow associations with

  9. Subtractive genomics approach to identify putative drug targets and identification of drug-like molecules for beta subunit of DNA polymerase III in Streptococcus species.

    PubMed

    Georrge, John J; Umrania, V V

    2012-07-01

    The prolonged use of the antibiotics over the years has transformed many organisms resistant to multiple drugs. This has made the field of drug discovery of vital importance in curing various infections and diseases. The drugs act by binding to a specific target protein of prime importance for the cell's survival. Streptococcus agalactiae, Streptococcus pneumoniae, and Streptococcus pyogenes are the few gram positive organisms that have developed resistance to drugs. It causes pneumonia, meningitis, pharyngitis, otitis media, sinusitis, bacteremia, pericarditis, and arthritis infections. The present study was carried out to identify potential drug targets and inhibitors for beta subunit of DNA polymerase III in these three Streptococcus species that might facilitate the discovery of novel drugs in near future. Various steps were adopted to find out novel drug targets. And finally 3D structure of DNA polymerase III subunit beta was modeled. The ligand library was generated from various databases to find the most suitable ligands. All the ligands were docked using Molegro Virtual Docker and the lead molecules were investigated for ADME and toxicity. PMID:22415782

  10. A Cross-Species Analysis in Pancreatic Neuroendocrine Tumors Reveals Molecular Subtypes with Distinctive Clinical, Metastatic, Developmental, and Metabolic Characteristics

    PubMed Central

    Sadanandam, Anguraj; Wullschleger, Stephan; Lyssiotis, Costas A.; Grötzinger, Carsten; Barbi, Stefano; Bersani, Samantha; Körner, Jan; Wafy, Ismael; Mafficini, Andrea; Lawlor, Rita T.; Simbolo, Michele; Asara, John M.; Bläker, Hendrik; Cantley, Lewis C.; Wiedenmann, Bertram; Scarpa, Aldo; Hanahan, Douglas

    2016-01-01

    Seeking to assess the representative and instructive value of an engineered mouse model of pancreatic neuroendocrine tumors (PanNET) for its cognate human cancer, we profiled and compared mRNA and miRNA transcriptomes of tumors from both. Mouse PanNET tumors could be classified into two distinctive subtypes, well-differentiated islet/insulinoma tumors (IT) and poorly differentiated tumors associated with liver metastases, dubbed metastasis-like primary (MLP). Human PanNETs were independently classified into these same two subtypes, along with a third, specific gene mutation–enriched subtype. The MLP subtypes in human and mouse were similar to liver metastases in terms of miRNA and mRNA transcriptome profiles and signature genes. The human/mouse MLP subtypes also similarly expressed genes known to regulate early pancreas development, whereas the IT subtypes expressed genes characteristic of mature islet cells, suggesting different tumorigenesis pathways. In addition, these subtypes exhibit distinct metabolic profiles marked by differential pyruvate metabolism, substantiating the significance of their separate identities. SIGNIFICANCE This study involves a comprehensive cross-species integrated analysis of multi-omics profiles and histology to stratify PanNETs into subtypes with distinctive characteristics. We provide support for the RIP1-TAG2 mouse model as representative of its cognate human cancer with prospects to better understand PanNET heterogeneity and consider future applications of personalized cancer therapy. PMID:26446169

  11. The Mitochondrial Cytochrome Oxidase Subunit I Gene Occurs on a Minichromosome with Extensive Heteroplasmy in Two Species of Chewing Lice, Geomydoecus aurei and Thomomydoecus minor.

    PubMed

    Pietan, Lucas L; Spradling, Theresa A; Demastes, James W

    2016-01-01

    In animals, mitochondrial DNA (mtDNA) typically occurs as a single circular chromosome with 13 protein-coding genes and 22 tRNA genes. The various species of lice examined previously, however, have shown mitochondrial genome rearrangements with a range of chromosome sizes and numbers. Our research demonstrates that the mitochondrial genomes of two species of chewing lice found on pocket gophers, Geomydoecus aurei and Thomomydoecus minor, are fragmented with the 1,536 base-pair (bp) cytochrome-oxidase subunit I (cox1) gene occurring as the only protein-coding gene on a 1,916-1,964 bp minicircular chromosome in the two species, respectively. The cox1 gene of T. minor begins with an atypical start codon, while that of G. aurei does not. Components of the non-protein coding sequence of G. aurei and T. minor include a tRNA (isoleucine) gene, inverted repeat sequences consistent with origins of replication, and an additional non-coding region that is smaller than the non-coding sequence of other lice with such fragmented mitochondrial genomes. Sequences of cox1 minichromosome clones for each species reveal extensive length and sequence heteroplasmy in both coding and noncoding regions. The highly variable non-gene regions of G. aurei and T. minor have little sequence similarity with one another except for a 19-bp region of phylogenetically conserved sequence with unknown function. PMID:27589589

  12. The Mitochondrial Cytochrome Oxidase Subunit I Gene Occurs on a Minichromosome with Extensive Heteroplasmy in Two Species of Chewing Lice, Geomydoecus aurei and Thomomydoecus minor

    PubMed Central

    Pietan, Lucas L.; Spradling, Theresa A.

    2016-01-01

    In animals, mitochondrial DNA (mtDNA) typically occurs as a single circular chromosome with 13 protein-coding genes and 22 tRNA genes. The various species of lice examined previously, however, have shown mitochondrial genome rearrangements with a range of chromosome sizes and numbers. Our research demonstrates that the mitochondrial genomes of two species of chewing lice found on pocket gophers, Geomydoecus aurei and Thomomydoecus minor, are fragmented with the 1,536 base-pair (bp) cytochrome-oxidase subunit I (cox1) gene occurring as the only protein-coding gene on a 1,916–1,964 bp minicircular chromosome in the two species, respectively. The cox1 gene of T. minor begins with an atypical start codon, while that of G. aurei does not. Components of the non-protein coding sequence of G. aurei and T. minor include a tRNA (isoleucine) gene, inverted repeat sequences consistent with origins of replication, and an additional non-coding region that is smaller than the non-coding sequence of other lice with such fragmented mitochondrial genomes. Sequences of cox1 minichromosome clones for each species reveal extensive length and sequence heteroplasmy in both coding and noncoding regions. The highly variable non-gene regions of G. aurei and T. minor have little sequence similarity with one another except for a 19-bp region of phylogenetically conserved sequence with unknown function. PMID:27589589

  13. IDENTIFICATION OF CRYPTOSPORIDIUM SPECIES AND SOURCES IN RAW WASTEWATER USING A SMALL SUBUNIT RRNA-BASED PCR-RFLP TOOL

    EPA Science Inventory

    The species composition and source of Cryptosporidium oocysts in wastewater have never been determined, even though it is widely assumed that these oocysts are from human sewage. Recent molecular characterizations of Cryptosporidium parasites make it possible to differentiate hum...

  14. Cryptosporidiosis caused by two distinct species in Russian tortoises and a pancake tortoise.

    PubMed

    Griffin, Chris; Reavill, Drury R; Stacy, Brian A; Childress, April L; Wellehan, James F X

    2010-05-28

    Cryptosporidiosis in squamates is well documented, but there is very limited information available on cryptosporidiosis in testudines. We describe three cases of cryptosporidiosis in tortoises with associated pathology. Two Russian tortoises (Agrionemys [Testudo] horsfieldii) and a pancake tortoise (Malacochersus tornieri), all from separate collections, were found dead. At necropsy, two had histological evidence of intestinal cryptosporidiosis and one had gastric cryptosporidiosis. Consensus Cryptosporidium sp. PCR and sequencing was used to identify the Cryptosporidium sp. present in these three tortoises. In the juvenile Russian tortoise with gastric cryptosporidiosis, the organism had 98% homology with a previously reported sequence from an Indian star tortoise isolate. A second chelonian Cryptosporidium sp. was identified in the pancake tortoise and the second Russian tortoise. This sequence was 100% identical to a shorter gene sequence previously reported in a marginated tortoise. This is the first report coordinating pathology with Cryptosporidium characterization in chelonians. The two Cryptosporidium sp. found in tortoises segregate according to site of infection, and there may be further differences in pathology, host range, and transmission. These Cryptosporidium sp. appear to be able to infect diverse tortoise host species. This may be an under-recognized problem in tortoises.

  15. Remarkable conservation of distinct nonclassical MHC class I lineages in divergent amphibian species.

    PubMed

    Goyos, Ana; Sowa, Jessica; Ohta, Yuko; Robert, Jacques

    2011-01-01

    Nonclassical MHC class Ib (class Ib) genes are heterogeneous genes encoding molecules that are structurally similar to classical MHC class Ia molecules but with limited tissue distribution and polymorphism. Mammalian class Ib genes have diverse and often uncharacterized functions, and because of their rapid rate of evolution, class Ib phylogeny is difficult to establish. We have conducted an extensive genomic, molecular, and phylogenetic characterization of class Ib genes in two Xenopodinae amphibian species of different genera that diverged from a common ancestor as long ago as primates and rodents (∼65 million years). In contrast with the unsteadiness of mammalian class Ib genes, our results reveal an unusual degree of conservation of most Xenopodinae class Ib gene lineages, including a novel monogenic lineage represented by the divergent Xenopus laevis XNC10 gene and its unequivocal Silurana (Xenopus) tropicalis orthologue, SNC10. The preferential expression of this gene lineage by thymocytes themselves from the onset of thymic organogenesis is consistent with a specialized role of class Ib in early T cell development and suggests such a function is conserved in all tetrapods. PMID:21115732

  16. The trigonal bipyramidal MN3M species: a new kind of aromatic complex containing a multiple-fold aromatic N3 subunit.

    PubMed

    Li, Ying; Li, Zhi-Ru; Wu, Di; Chen, Wei; Sun, Chia-Chung

    2005-12-01

    A new kind of aromatic trigonal bipyramidal MN3M (M=Be, B, Mg, Al, and Ca) species, with all real frequencies, is obtained at the MP2/6-311+G(3d) level. The nucleus-independent chemical shift values are -102.16 ppm for the N3 (3-) ring, and -74.09, -79.39, -65.06, -74.44, and -62.33 ppm (at the geometrical center of the trigonal bipyramid) for BeN3Be, BN3B, MgN3Mg, AlN3Al, and CaN3Ca, respectively. Molecular orbital analysis indicates that the regular triangular N3 (3-) ring and each MN3M species have three aromatic six-electron systems (pi, sigma(p), and sigma(s)) and exhibit threefold aromaticity. The CaN3Ca species has a very low vertical ionization energy of 3.64 eV at the CCSD(T)/6-311+G(3d) level, which is even lower than the ionization energy (3.9 eV) of the Cs atom. Therefore, CaN3Ca can be considered as a new superalkali species. A further study on the CaN3CaCl molecule confirms the superalkali characteristics of CaN3Ca. Two interesting phenomena are explored in the MN3M species: the delocalized electron cloud of the N3 subunit is elongated by two M cations, and the electron clouds of two M cations are distended by the N3 (3-) ring.

  17. Novel sequences encoding venom C-type lectins are conserved in phylogenetically and geographically distinct Echis and Bitis viper species.

    PubMed

    Harrison, R A; Oliver, J; Hasson, S S; Bharati, K; Theakston, R D G

    2003-10-01

    Envenoming by Echis saw scaled vipers and Bitis arietans puff adders is the leading cause of death and morbidity in Africa due to snake bite. Despite their medical importance, the composition and constituent functionality of venoms from these vipers remains poorly understood. Here, we report the cloning of cDNA sequences encoding seven clusters or isoforms of the haemostasis-disruptive C-type lectin (CTL) proteins from the venom glands of Echis ocellatus, E. pyramidum leakeyi, E. carinatus sochureki and B. arietans. All these CTL sequences encoded the cysteine scaffold that defines the carbohydrate-recognition domain of mammalian CTLs. All but one of the Echis and Bitis CTL sequences showed greater sequence similarity to the beta than alpha CTL subunits in venoms of related Asian and American vipers. Four of the new CTL clusters showed marked inter-cluster sequence conservation across all four viper species which were significantly different from that of previously published viper CTLs. The other three Echis and Bitis CTL clusters showed varying degrees of sequence similarity to published viper venom CTLs. Because viper venom CTLs exhibit a high degree of sequence similarity and yet exert profoundly different effects on the mammalian haemostatic system, no attempt was made to assign functionality to the new Echis and Bitis CTLs on the basis of sequence alone. The extraordinary level of inter-specific and inter-generic sequence conservation exhibited by the Echis and Bitis CTLs leads us to speculate that antibodies to representative molecules should neutralise the biological function of this important group of venom toxins in vipers that are distributed throughout Africa, the Middle East and the Indian subcontinent. PMID:14557069

  18. Molecular characterization of a distinct bipartite Begomovirus species infecting ivy gourd (Coccinia grandis L.) in Tamil Nadu, India.

    PubMed

    Nagendran, K; Satya, V K; Mohankumar, S; Karthikeyan, G

    2016-02-01

    A distinct bipartite begomovirus was found to be associated with the mosaic disease on ivy gourd (Coccinia grandis L.) in Tamil Nadu, India. The complete DNA A and DNA B components were cloned by rolling circle amplification. Genome organization of this virus is found to be typical of Old World bipartite begomovirus. The association of betasatellite component with this virus is absent. The closest nucleotide identity of 73.4 % was seen with the Loofa yellow mosaic virus (LYMV-[VN]-AF509739) suggesting that it is a new virus species Coccinia mosaic virus (CoMoV-Ivy gourd [TN TDV Coc1]) and distantly related to the other known begomoviruses. The DNA B component shared a maximum identity of 55 % with that of Tomato leaf curl New Delhi virus (ToLCNDV). In the phylogenetic analysis, CoMoV-Ivy gourd form cluster separate from other begomoviruses. Recombination analysis showed that there was no recombination event in the genome. This is the distinct begomovirus infecting ivy gourd. PMID:26739457

  19. Deep mitochondrial DNA lineage divergences within Alberta populations of Dermacentor albipictus (Acari: Ixodidae) do not indicate distinct species.

    PubMed

    Leo, Sarah S T; Pybus, Margo J; Sperling, Felix A H

    2010-07-01

    The winter tick Dermacentor albipictus (Packard) has a single-host life cycle that allows it to reach severe infestation levels on ungulates, particularly moose. Genotypic variation within these and related ticks has been a source of taxonomic confusion, although the continuity in their morphology and life history has generally been interpreted as indicating the existence of a single species. To further investigate this variation, we sequenced regions of two mitochondrial DNA (mtDNA) genes (COI and 16S rDNA),two nuclear genes (lysozyme and ITS-2), and two bacterial markers from Francisella-like endosymbionts found in these ticks (eubacterial mtDNA 16S rRNA and a homolog of Francisella tularensis [Dorofe'ev] 17-kDa lipoprotein). We sampled 42 D. albipictus individuals from whitetail and mule deer culled from three populations in east-central Alberta, as well as four D. albipictus and two Dermacentor variabilis (Say) from other locations. We then compared DNA sequence variation between the genes and related this to variation in the morphology of spiracle plates. Both mtDNA regions indicated two deeply diverged lineages (mean difference of 7.1% for COI and 4.5% for 16S) that would normally be considered diagnostic of distinct species in DNA barcoding studies. However, very little divergence was revealed by nuclear gene sequences, bacterial endosymbionts, and morphometric analyses, and any variation that did occur in these markers was not congruent with mtDNA divergences. We conclude that the sampled populations in Alberta represent a single species, D. albipictus, and reiterate the importance of integrative approaches in species delimitation. PMID:20695271

  20. Single and mixed-species trypanosome and microsporidia infections elicit distinct, ephemeral cellular and humoral immune responses in honey bees.

    PubMed

    Schwarz, Ryan S; Evans, Jay D

    2013-01-01

    Frequently encountered parasite species impart strong selective pressures on host immune system evolution and are more apt to concurrently infect the same host, yet molecular impacts in light of this are often overlooked. We have contrasted immune responses in honey bees to two common eukaryotic endoparasites by establishing single and mixed-species infections using the long-associated parasite Crithidia mellificae and the emergent parasite Nosema ceranae. Quantitative polymerase chain reaction was used to screen host immune gene expression at 9 time points post inoculation. Systemic responses in abdomens during early stages of parasite establishment revealed conserved receptor (Down syndrome cell adhesion molecule, Dscam and nimrod C1, nimC1), signaling (MyD88 and Imd) and antimicrobial peptide (AMP) effector (Defensin 2) responses. Late, established infections were distinct with a refined 2 AMP response to C. mellificae that contrasted starkly with a 5 AMP response to N. ceranae. Mixed species infections induced a moderate 3 AMPs. Transcription in gut tissues highlighted important local roles for Dscam toward both parasites and Imd signaling toward N. ceranae. At both systemic and local levels Dscam, MyD88 and Imd transcription was consistently correlated based on clustering analysis. Significant gene suppression occurred in two cases from midgut to ileum tissue: Dscam was lowered during mixed infections compared to N. ceranae infections and both C. mellificae and mixed infections had reduced nimC1 transcription compared to uninfected controls. We show that honey bees rapidly mount complex immune responses to both Nosema and Crithidia that are dynamic over time and that mixed-species infections significantly alter local and systemic immune gene transcription.

  1. Development and application of small-subunit rRNA probes for assessment of selected Thiobacillus species and members of the genus Acidiphilium.

    PubMed

    Peccia, J; Marchand, E A; Silverstein, J; Hernandez, M

    2000-07-01

    Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers. Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of the Thiobacillus and Acidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria: Thiobacillus ferrooxidans and T. thiooxidans (probe Thio820) and members of the genus Acidiphilium (probe Acdp821). Using (32)P radiolabels, probe specificity was characterized by hybridization dissociation temperature (T(d)) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determined T(d)s. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris.

  2. Development and application of small-subunit rRNA probes for assessment of selected Thiobacillus species and members of the genus Acidiphilium.

    PubMed

    Peccia, J; Marchand, E A; Silverstein, J; Hernandez, M

    2000-07-01

    Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers. Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of the Thiobacillus and Acidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria: Thiobacillus ferrooxidans and T. thiooxidans (probe Thio820) and members of the genus Acidiphilium (probe Acdp821). Using (32)P radiolabels, probe specificity was characterized by hybridization dissociation temperature (T(d)) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determined T(d)s. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris. PMID:10877807

  3. Development and Application of Small-Subunit rRNA Probes for Assessment of Selected Thiobacillus Species and Members of the Genus Acidiphilium

    PubMed Central

    Peccia, Jordan; Marchand, Eric A.; Silverstein, Joann; Hernandez, Mark

    2000-01-01

    Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers. Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of the Thiobacillus and Acidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria: Thiobacillus ferrooxidans and T. thiooxidans (probe Thio820) and members of the genus Acidiphilium (probe Acdp821). Using 32P radiolabels, probe specificity was characterized by hybridization dissociation temperature (Td) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determined Tds. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris. PMID:10877807

  4. Subterranean, Herbivore-Induced Plant Volatile Increases Biological Control Activity of Multiple Beneficial Nematode Species in Distinct Habitats

    PubMed Central

    Ali, Jared G.; Alborn, Hans T.; Campos-Herrera, Raquel; Kaplan, Fatma; Duncan, Larry W.; Rodriguez-Saona, Cesar; Koppenhöfer, Albrecht M.; Stelinski, Lukasz L.

    2012-01-01

    While the role of herbivore-induced volatiles in plant-herbivore-natural enemy interactions is well documented aboveground, new evidence suggests that belowground volatile emissions can protect plants by attracting entomopathogenic nematodes (EPNs). However, due to methodological limitations, no study has previously detected belowground herbivore-induced volatiles in the field or quantified their impact on attraction of diverse EPN species. Here we show how a belowground herbivore-induced volatile can enhance mortality of agriculturally significant root pests. First, in real time, we identified pregeijerene (1,5-dimethylcyclodeca-1,5,7-triene) from citrus roots 9–12 hours after initiation of larval Diaprepes abbreviatus feeding. This compound was also detected in the root zone of mature citrus trees in the field. Application of collected volatiles from weevil-damaged citrus roots attracted native EPNs and increased mortality of beetle larvae (D. abbreviatus) compared to controls in a citrus orchard. In addition, field applications of isolated pregeijerene caused similar results. Quantitative real-time PCR revealed that pregeijerene increased pest mortality by attracting four species of naturally occurring EPNs in the field. Finally, we tested the generality of this root-zone signal by application of pregeijerene in blueberry fields; mortality of larvae (Galleria mellonella and Anomala orientalis) again increased by attracting naturally occurring populations of an EPN. Thus, this specific belowground signal attracts natural enemies of widespread root pests in distinct agricultural systems and may have broad potential in biological control of root pests. PMID:22761668

  5. Molecular evidence for genetic distinctions between Chlamydiaceae detected in Siamese crocodiles (Crocodylus siamensis) and known Chlamydiaceae species.

    PubMed

    Sariya, Ladawan; Kladmanee, Kan; Bhusri, Benjaporn; Thaijongrak, Prawporn; Tonchiangsai, Kanittha; Chaichoun, Kridsada; Ratanakorn, Parntep

    2015-02-01

    Chlamydiosis, caused by Chlamydiaceae, is a zoonotic disease found in humans and several species of animals, including reptiles and amphibians. Although chlamydiosis in saltwater crocodiles has been previously reported in South Africa and Papua New Guinea, the reported strains have not been identified or confirmed. Therefore, the main aim of this study was to sequence and characterize Chamydiaceae isolated from Siamese crocodiles. Results showed the 16S ribosomal (r) RNA and the 16S/23S rRNA gene of the crocodile isolates were closely related to the genus Chlamydophila with matched identity greater than 98%. The phylogenetic tree constructed from the 16S/23S rRNA gene showed the crocodile cluster diverges far from Cp. caviae with a 100% bootstrap value. The tree based on the ompA gene loci distinguished the crocodile strains into genotypes I, II, and III. The present study is the first report on Chlamydophila detected in Siamese crocodiles that is genetically distinct from the known species of Chlamydiaceae.

  6. Molecular evidence for genetic distinctions between Chlamydiaceae detected in Siamese crocodiles (Crocodylus siamensis) and known Chlamydiaceae species.

    PubMed

    Sariya, Ladawan; Kladmanee, Kan; Bhusri, Benjaporn; Thaijongrak, Prawporn; Tonchiangsai, Kanittha; Chaichoun, Kridsada; Ratanakorn, Parntep

    2015-02-01

    Chlamydiosis, caused by Chlamydiaceae, is a zoonotic disease found in humans and several species of animals, including reptiles and amphibians. Although chlamydiosis in saltwater crocodiles has been previously reported in South Africa and Papua New Guinea, the reported strains have not been identified or confirmed. Therefore, the main aim of this study was to sequence and characterize Chamydiaceae isolated from Siamese crocodiles. Results showed the 16S ribosomal (r) RNA and the 16S/23S rRNA gene of the crocodile isolates were closely related to the genus Chlamydophila with matched identity greater than 98%. The phylogenetic tree constructed from the 16S/23S rRNA gene showed the crocodile cluster diverges far from Cp. caviae with a 100% bootstrap value. The tree based on the ompA gene loci distinguished the crocodile strains into genotypes I, II, and III. The present study is the first report on Chlamydophila detected in Siamese crocodiles that is genetically distinct from the known species of Chlamydiaceae. PMID:25854083

  7. Nine novel DNA components associated with the foorkey disease of large cardamom: evidence of a distinct babuvirus species in Nanoviridae.

    PubMed

    Mandal, Bikash; Shilpi, S; Barman, Ashis Roy; Mandal, Seema; Varma, Anupam

    2013-12-26

    Foorkey disease is a serious constraint to the production of large cardamom (Amomum subulatum, family Zingiberaceae). The disease is characterized by profuse proliferation of excessive stunted shoots, which makes the clump totally unproductive. The disease has been known in India since 1936 but the complete genome of the virus had not yet been characterized. In a preliminary study, an associated virus tentatively named as Cardamom bushy dwarf virus (CBDV) was identified based on the partial sequence of a single DNA component (DNA-R). In the present study, a high incidence (37.2-39.3%) of foorkey was recorded in certain plantations in the Darjeeling hills located at lower altitudes (300-1380 m) and CBDV was detected in several field samples by PCR. Nine novel DNA components were isolated and characterized from foorkey affected plants. CBDV contained six major DNA components (DNA-R, -S, -M, -C, -N and -U3) similar to the integral genome components known for the members of the genus Babuvirus in the family Nanoviridae. Additional components, satellite Rep (DNA-sRep1) and unknown components (DNA-Uf1 and -Uf2) were also identified. The size of the genome components ranged from 1028 to 1127. The sequence identity and phylogeny based on the individual components as well as overall genome (59.8-62% identity) distinguished CBDV from the two existing babuvirus species, Banana bunchy top virus and Abaca bunchy top virus. CBDV is the first distinct babuvirus species that affects plant species outside family Musaceae. This study shows further diversity in the genus Babuvirus. PMID:24091364

  8. Nine novel DNA components associated with the foorkey disease of large cardamom: evidence of a distinct babuvirus species in Nanoviridae.

    PubMed

    Mandal, Bikash; Shilpi, S; Barman, Ashis Roy; Mandal, Seema; Varma, Anupam

    2013-12-26

    Foorkey disease is a serious constraint to the production of large cardamom (Amomum subulatum, family Zingiberaceae). The disease is characterized by profuse proliferation of excessive stunted shoots, which makes the clump totally unproductive. The disease has been known in India since 1936 but the complete genome of the virus had not yet been characterized. In a preliminary study, an associated virus tentatively named as Cardamom bushy dwarf virus (CBDV) was identified based on the partial sequence of a single DNA component (DNA-R). In the present study, a high incidence (37.2-39.3%) of foorkey was recorded in certain plantations in the Darjeeling hills located at lower altitudes (300-1380 m) and CBDV was detected in several field samples by PCR. Nine novel DNA components were isolated and characterized from foorkey affected plants. CBDV contained six major DNA components (DNA-R, -S, -M, -C, -N and -U3) similar to the integral genome components known for the members of the genus Babuvirus in the family Nanoviridae. Additional components, satellite Rep (DNA-sRep1) and unknown components (DNA-Uf1 and -Uf2) were also identified. The size of the genome components ranged from 1028 to 1127. The sequence identity and phylogeny based on the individual components as well as overall genome (59.8-62% identity) distinguished CBDV from the two existing babuvirus species, Banana bunchy top virus and Abaca bunchy top virus. CBDV is the first distinct babuvirus species that affects plant species outside family Musaceae. This study shows further diversity in the genus Babuvirus.

  9. Characterization of a seventh different subunit of beef heart cytochrome c oxidase. Similarities between the beef heart enzyme and that from other species.

    PubMed

    Downer, N W; Robinson, N C

    1976-06-29

    Beef heart cytochrome c oxidase has been resolved into seven subunits by electrophoresis in highly cross-linked gels containing urea and sodium dodecyl sulfate. The molecular weights of the polypeptides are estimated to be I, 35 400; II, 24 100; III, 21 000; IV, 16 800; V, 12 400; VI, 8200; and VII, 4400. It has been shown that subunits II and III can coelectrophorese on standard sodium dodecyl sulfate-polyacrylamide gels and appear as a single component with an apparent molecular weight of 22 500. This accounts for previous reports that the beef heart enzyme contains only six subunits. Amino acid analysis of the isolated subunits I, II, and III revealed that they have polarities of 35.5, 44.7, and 39.9%, respectively. All three subunits have an extremely high leucine content and a low percentage of basic amino acids relative to subunits IV-VII. The size, number, and properties of subunits in the beef heart cytochrome c oxidase complex suggest that it has essentially the same subunit structure as the complexes isolated from Saccharomyces cerevisiae and Neurospora crassa.

  10. 75 FR 38979 - Endangered and Threatened Species; Initiation of a 5-Year Review of the Eastern Distinct...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-07-07

    ... for the eastern Distinct Population Segment of the Steller sea lion (75 FR 37385). NMFS inadvertently... of a 5-Year Review of the Eastern Distinct Population Segment of the Steller Sea Lion AGENCY... of a 5-year review of the eastern Distinct Population Segment (DPS) of the Steller Sea...

  11. Comparison of tick-borne microorganism communities in Ixodes spp. of the Ixodes ricinus species complex at distinct geographical regions.

    PubMed

    Movila, Alexandru; Dubinina, Helen V; Sitnicova, Natalia; Bespyatova, Liubov; Uspenskaia, Inga; Efremova, Galina; Toderas, Ion; Alekseev, Andrey N

    2014-05-01

    Characterizing the tick-borne microorganism communities of Ixodes ricinus (sheep tick) and Ixodes persulcatus (taiga tick) from the I. ricinus species complex in distinct geographical regions of Eastern Europe and European Russia, we demonstrated differences between the two ticks. Taiga ticks were more frequently mono- and co-infected than sheep ticks: 24.4 % (45/184 tested ticks) versus 17.5 % (52/297) and 4.3 % (8/184) versus 3.4 % (10/297), respectively. Ginsberg co-infection index values were significant at the various sites. Diversity of the tick-borne microorganism communities was estimated by the Shannon index, reaching values of 1.71 ± 0.46 and 1.20 ± 0.15 at the sheep-tick and the taiga-tick harbored sites, respectively. Richness of the tick-borne microorganism community in the sheep tick collection sites was about twice the value of the taiga tick collection sites. Future investigations are warranted to further characterize the peculiarities of the tick-borne microorganism communities among the ticks of the Ixodes ricinus complex.

  12. Simultaneous Observation of an Intraband Transition and Distinct Transient Species in the Infrared Region for Perovskite Solar Cells.

    PubMed

    Narra, Sudhakar; Chung, Chih-Chun; Diau, Eric Wei-Guang; Shigeto, Shinsuke

    2016-07-01

    Solar cells based on organometal-halide perovskites such as CH3NH3PbI3 have emerged as a promising next-generation photovoltaic system, but the underlying photophysics and photochemistry remain to be established because of the limited availability of methods to implement the simultaneous and direct measurement of various charge carriers and ions that play a crucial role in the operating device. We used nanosecond time-resolved infrared (IR) spectroscopy to investigate, with high molecular specificity, distinct transient species that are formed in perovskite solar cells after photoexcitation. In CH3NH3PbI3 planar-heterojuction solar cells, we simultaneously observed infrared spectral signatures that are associated with an intraband transition of conduction-band electrons, Fano resonance, and the spiro-OMeTAD cation having an exceptionally short lifetime of 1.0 μs (at ∼1485 cm(-1)). The present results show that the time-resolved IR method offers a unique capability to elucidate these important transients in perovskite solar cells and their dynamic interplay in a comprehensive manner.

  13. Simultaneous Observation of an Intraband Transition and Distinct Transient Species in the Infrared Region for Perovskite Solar Cells.

    PubMed

    Narra, Sudhakar; Chung, Chih-Chun; Diau, Eric Wei-Guang; Shigeto, Shinsuke

    2016-07-01

    Solar cells based on organometal-halide perovskites such as CH3NH3PbI3 have emerged as a promising next-generation photovoltaic system, but the underlying photophysics and photochemistry remain to be established because of the limited availability of methods to implement the simultaneous and direct measurement of various charge carriers and ions that play a crucial role in the operating device. We used nanosecond time-resolved infrared (IR) spectroscopy to investigate, with high molecular specificity, distinct transient species that are formed in perovskite solar cells after photoexcitation. In CH3NH3PbI3 planar-heterojuction solar cells, we simultaneously observed infrared spectral signatures that are associated with an intraband transition of conduction-band electrons, Fano resonance, and the spiro-OMeTAD cation having an exceptionally short lifetime of 1.0 μs (at ∼1485 cm(-1)). The present results show that the time-resolved IR method offers a unique capability to elucidate these important transients in perovskite solar cells and their dynamic interplay in a comprehensive manner. PMID:27302315

  14. Molecular characterization of Iranian wheat stripe virus shows its taxonomic position as a distinct species in the genus Tenuivirus.

    PubMed

    Heydarnejad, J; Barclay, W S; Izadpanah, K; Hunter, F R; Gooding, M J

    2006-02-01

    The full lengths of three genome segments of Iranian wheat stripe virus (IWSV) were amplified by reverse transcription (RT) followed by polymerase chain reaction (PCR) using a primer complementary to tenuivirus conserved terminal sequences. The segments were sequenced and found to comprise 3469, 2337, and 1831 nt, respectively. The gene organization of these segments is similar to that of other known tenuiviruses, each displaying an ambisense coding strategy. IWSV segments, however, are different from those of other viruses with respect to the number of nucleotides and deduced amino acid sequence for each ORF. Depending on the segment, the first 16-22 nt at the 5' end and the first 16 nt at the 3' end are highly conserved among IWSV and rice hoja blanca virus (RHBV), rice stripe virus (RSV) and maize stripe virus (MStV). In addition, the first 15-18 nt at the 5' end are complementary to the first 16-18 nt at the 3' end. Phylogenetic analyses showed close similarity and a common ancestor for IWSV, RHBV, and Echinochloa hoja blanca virus (EHBV). These findings confirm the position of IWSV as a distinct species in the genus Tenuivirus. PMID:16328148

  15. Discovery of a Novel Bottlenose Dolphin Coronavirus Reveals a Distinct Species of Marine Mammal Coronavirus in Gammacoronavirus

    PubMed Central

    Woo, Patrick C. Y.; Lau, Susanna K. P.; Lam, Carol S. F.; Tsang, Alan K. L.; Hui, Suk-Wai; Fan, Rachel Y. Y.; Martelli, Paolo

    2014-01-01

    While gammacoronaviruses mainly comprise infectious bronchitis virus (IBV) and its closely related bird coronaviruses (CoVs), the only mammalian gammacoronavirus was discovered from a white beluga whale (beluga whale CoV [BWCoV] SW1) in 2008. In this study, we discovered a novel gammacoronavirus from fecal samples from three Indo-Pacific bottlenose dolphins (Tursiops aduncus), which we named bottlenose dolphin CoV (BdCoV) HKU22. All the three BdCoV HKU22-positive samples were collected on the same date, suggesting a cluster of infection, with viral loads of 1 × 103 to 1 × 105 copies per ml. Clearance of virus was associated with a specific antibody response against the nucleocapsid of BdCoV HKU22. Complete genome sequencing and comparative genome analysis showed that BdCoV HKU22 and BWCoV SW1 have similar genome characteristics and structures. Their genome size is about 32,000 nucleotides, the largest among all CoVs, as a result of multiple unique open reading frames (NS5a, NS5b, NS5c, NS6, NS7, NS8, NS9, and NS10) between their membrane (M) and nucleocapsid (N) protein genes. Although comparative genome analysis showed that BdCoV HKU22 and BWCoV SW1 should belong to the same species, a major difference was observed in the proteins encoded by their spike (S) genes, which showed only 74.3 to 74.7% amino acid identities. The high ratios of the number of synonymous substitutions per synonymous site (Ks) to the number of nonsynonymous substitutions per nonsynonymous site (Ka) in multiple regions of the genome, especially the S gene (Ka/Ks ratio, 2.5), indicated that BdCoV HKU22 may be evolving rapidly, supporting a recent transmission event to the bottlenose dolphins. We propose a distinct species, Cetacean coronavirus, in Gammacoronavirus, to include BdCoV HKU22 and BWCoV SW1, whereas IBV and its closely related bird CoVs represent another species, Avian coronavirus, in Gammacoronavirus. PMID:24227844

  16. Discovery of a novel bottlenose dolphin coronavirus reveals a distinct species of marine mammal coronavirus in Gammacoronavirus.

    PubMed

    Woo, Patrick C Y; Lau, Susanna K P; Lam, Carol S F; Tsang, Alan K L; Hui, Suk-Wai; Fan, Rachel Y Y; Martelli, Paolo; Yuen, Kwok-Yung

    2014-01-01

    While gammacoronaviruses mainly comprise infectious bronchitis virus (IBV) and its closely related bird coronaviruses (CoVs), the only mammalian gammacoronavirus was discovered from a white beluga whale (beluga whale CoV [BWCoV] SW1) in 2008. In this study, we discovered a novel gammacoronavirus from fecal samples from three Indo-Pacific bottlenose dolphins (Tursiops aduncus), which we named bottlenose dolphin CoV (BdCoV) HKU22. All the three BdCoV HKU22-positive samples were collected on the same date, suggesting a cluster of infection, with viral loads of 1 × 10(3) to 1 × 10(5) copies per ml. Clearance of virus was associated with a specific antibody response against the nucleocapsid of BdCoV HKU22. Complete genome sequencing and comparative genome analysis showed that BdCoV HKU22 and BWCoV SW1 have similar genome characteristics and structures. Their genome size is about 32,000 nucleotides, the largest among all CoVs, as a result of multiple unique open reading frames (NS5a, NS5b, NS5c, NS6, NS7, NS8, NS9, and NS10) between their membrane (M) and nucleocapsid (N) protein genes. Although comparative genome analysis showed that BdCoV HKU22 and BWCoV SW1 should belong to the same species, a major difference was observed in the proteins encoded by their spike (S) genes, which showed only 74.3 to 74.7% amino acid identities. The high ratios of the number of synonymous substitutions per synonymous site (Ks) to the number of nonsynonymous substitutions per nonsynonymous site (Ka) in multiple regions of the genome, especially the S gene (Ka/Ks ratio, 2.5), indicated that BdCoV HKU22 may be evolving rapidly, supporting a recent transmission event to the bottlenose dolphins. We propose a distinct species, Cetacean coronavirus, in Gammacoronavirus, to include BdCoV HKU22 and BWCoV SW1, whereas IBV and its closely related bird CoVs represent another species, Avian coronavirus, in Gammacoronavirus. PMID:24227844

  17. Experimental Crossing of Two Distinct Species of Leopard Geckos, Eublepharis angramainyu and E. macularius: Viability, Fertility and Phenotypic Variation of the Hybrids

    PubMed Central

    Jančúchová-Lásková, Jitka; Landová, Eva; Frynta, Daniel

    2015-01-01

    Hybridization between distinct species of animals and subsequent genetic introgression plays a considerable role in the speciation process and the emergence of adaptive characters. Fitness of between-species hybrids usually sharply decreases with the divergence time of the concerned species and the divergence depth, which still allows for a successful crossing differs among principal clades of vertebrates. Recently, a review of hybridization events among distinct lizard species revealed that lizards belong to vertebrates with a highly developed ability to hybridize. In spite of this, reliable reports of experimental hybridizations between genetically fairly divergent species are only exceptional. Here, we show the results of the crossing of two distinct allopatric species of eyelid geckos possessing temperature sex determination and lacking sex chromosomes: Eublepharis macularius distributed in Pakistan/Afghanistan area and E. angramainyu, which inhabits Mesopotamia and adjacent areas. We demonstrated that F1 hybrids were viable and fertile, and the introgression of E. angramainyu genes into the E. macularius genome can be enabled via a backcrossing. The examined hybrids (except those of the F2 generation) displayed neither malformations nor a reduced survival. Analyses of morphometric and coloration traits confirmed phenotypic distinctness of both parental species and their F1 hybrids. These findings contrast with long-term geographic and an evolutionary separation of the studied species. Thus, the occurrence of fertile hybrids of comparably divergent species, such as E. angramainyu and E. macularius, may also be expected in other taxa of squamates. This would violate the current estimates of species diversity in lizards. PMID:26633648

  18. Experimental Crossing of Two Distinct Species of Leopard Geckos, Eublepharis angramainyu and E. macularius: Viability, Fertility and Phenotypic Variation of the Hybrids.

    PubMed

    Jančúchová-Lásková, Jitka; Landová, Eva; Frynta, Daniel

    2015-01-01

    Hybridization between distinct species of animals and subsequent genetic introgression plays a considerable role in the speciation process and the emergence of adaptive characters. Fitness of between-species hybrids usually sharply decreases with the divergence time of the concerned species and the divergence depth, which still allows for a successful crossing differs among principal clades of vertebrates. Recently, a review of hybridization events among distinct lizard species revealed that lizards belong to vertebrates with a highly developed ability to hybridize. In spite of this, reliable reports of experimental hybridizations between genetically fairly divergent species are only exceptional. Here, we show the results of the crossing of two distinct allopatric species of eyelid geckos possessing temperature sex determination and lacking sex chromosomes: Eublepharis macularius distributed in Pakistan/Afghanistan area and E. angramainyu, which inhabits Mesopotamia and adjacent areas. We demonstrated that F1 hybrids were viable and fertile, and the introgression of E. angramainyu genes into the E. macularius genome can be enabled via a backcrossing. The examined hybrids (except those of the F2 generation) displayed neither malformations nor a reduced survival. Analyses of morphometric and coloration traits confirmed phenotypic distinctness of both parental species and their F1 hybrids. These findings contrast with long-term geographic and an evolutionary separation of the studied species. Thus, the occurrence of fertile hybrids of comparably divergent species, such as E. angramainyu and E. macularius, may also be expected in other taxa of squamates. This would violate the current estimates of species diversity in lizards. PMID:26633648

  19. A non-toxigenic but morphologically and phylogenetically distinct new species of Pseudo-nitzschia, P. sabit sp. nov. (Bacillariophyceae).

    PubMed

    Teng, Sing Tung; Lim, Po Teen; Lim, Hong Chang; Rivera-Vilarelle, María; Quijano-Scheggia, Sonia; Takata, Yoshinobu; Quilliam, Michael A; Wolf, Matthias; Bates, Stephen S; Leaw, Chui Pin

    2015-08-01

    A new species of Pseudo-nitzschia (Bacillariophyceae) is described from plankton samples collected from Port Dickson (Malacca Strait, Malaysia) and Manzanillo Bay (Colima, Mexico). The species possesses a distinctive falcate cell valve, from which they form sickle-like colonies in both environmental samples and cultured strains. Detailed observation of frustules under TEM revealed ultrastructure that closely resembles P. decipiens, yet the new species differs by the valve shape and greater ranges of striae and poroid densities. The species is readily distinguished from the curve-shaped P. subcurvata by the presence of a central interspace. The morphological distinction is further supported by phylogenetic discrimination. We sequenced and analyzed the nuclear ribosomal RNA genes in the LSU and the second internal transcribed spacer, including its secondary structure, to infer the phylogenetic relationship of the new species with its closest relatives. The results revealed a distinct lineage of the new species, forming a sister cluster with its related species, P. decipiens and P. galaxiae, but not with P. subcurvata. We examined the domoic acid (DA) production of five cultured strains from Malaysia by Liquid chromatography-mass spectrometry (LC-MS), but they showed no detectable DA. Here, we present the taxonomic description of the vegetative cells, document the sexual reproduction, and detail the molecular phylogenetics of Pseudo-nitzschia sabit sp. nov.

  20. Phylogenetic study of clonal complex (CC)198 capsule null locus (cnl) genomes: A distinctive group within the species Neisseria meningitidis.

    PubMed

    Xu, Zheng; Du, Pengcheng; Zhu, Bingqing; Xu, Li; Wang, Haiyin; Gao, Yuan; Zhou, Haijian; Zhang, Wen; Chen, Chen; Shao, Zhujun

    2015-08-01

    Capsule null locus (cnl) strains, one type of specific unencapsulated Neisseria spp., only have regions D and E of the capsule gene cluster which encodes the genes for capsule biosynthesis, modification, and transportation. Compared with encapsulated strains, regions A and C of cnl strains have been replaced by 113 or 114 bp conserved non-coding sequences. Cnl strains include multiple clonal complexes (CC). According to previous studies, CC198 is the major clonal lineage in both cnl patients and healthy cnl carriers. We hypothesized that CC198 possesses different genome characteristics compared with other cnl strains. In this study, we obtained the draft genomes of two CC198 strains from healthy carriers. Using 75071 single nucleotide polymorphisms located in 1163 core genes, we constructed the phylogenetic relationships between a batch of representative Neisseria meningitidis genomes. CC198 and CC1136 clustered together, but apart from other N. meningitidis strains including CC53. We also aligned the sequences of genes located in regions D and E of the capsule gene locus from encapsulated and unencapsulated strains. A number of possible recombination events were identified in the galE and tex genes between different serogroups of encapsulated N. meningitidis and CC53 strains, especially in tex. In contrast, there is almost no recombination in N. meningitidis CC198 strains. These results showed that CC198 belongs to a phylogenetically distinct group within the species N. meningitidis, which may be directly derived from the cnl-type ancestor of N. meningitidis. The encapsulated strains may acquire other necessary genes for capsule formation by horizontal transfer.

  1. The identity of Cintractia carpophila var. kenaica: reclassification of a North American smut on Carex micropoda as a distinct species of Anthracoidea.

    PubMed

    Piątek, Marcin

    2013-07-01

    Cintractia carpophila var. kenaica, a neglected taxon described from Alaska more than half a century ago, is re-described and illustrated. Its nomenclature and taxonomic status are discussed. This smut species is characterised by small spores with a very finely verruculose surface rarely enclosed by a thin, hyaline, mucilaginous sheath, a wall with 2-5 distinct internal swellings, and parasitism on Carex micropoda (Carex sect. Dornera). It is reallocated to the genus Anthracoidea as a distinct species, Anthracoidea kenaica comb. nov., and assigned to Anthracoidea section Leiosporae which includes species having smooth or very finely verruculose spores. Morphological and biological characteristics of the five most similar Anthracoidea species are contrasted and discussed.

  2. Genes encoding the biotin carboxylase subunit of acetyl-CoA carboxylase from Brassica napus and parental species: cloning, expression patterns, and evolution.

    PubMed

    Li, Zhi-Guo; Yin, Wei-Bo; Song, Li-Ying; Chen, Yu-Hong; Guan, Rong-Zhan; Wang, Jing-Qiao; Wang, Richard R-C; Hu, Zan-Min

    2011-03-01

    Comparative genomics is a useful tool to investigate gene and genome evolution. Biotin carboxylase (BC), an important subunit of heteromeric acetyl-CoA carboxylase (ACCase) that is a rate-limiting enzyme in fatty acid biosynthesis in dicots, catalyzes ATP, biotin carboxyl carrier protein, and CO2 to form carboxybiotin carboxyl carrier protein. In this study, we cloned four genes encoding BC from Brassica napus L. (namely BnaC.BC.a, BnaC.BC.b, BnaA.BC.a, and BnaA.BC.b), and two were cloned from each of the two parental species Brassica rapa L. (BraA.BC.a and BraA.BC.b) and Brassica oleracea L. (BolC.BC.a and BolC.BC.b). Sequence analyses revealed that in B. napus the genes BnaC.BC.a and BnaC.BC.b were from the C genome of B. oleracea, whereas BnaA.BC.a and BnaA.BC.b were from the A genome of B. rapa. Comparative and cluster analysis indicated that these genes were divided into two major groups, BnaC.BC.a, BnaA.BC.a, BraA.BC.a, and BolC.BC.a in group-1 and BnaC.BC.b, BnaA.BC.b, BraA.BC.b, and BolC.BC.b in group-2. The divergence of group-1 and group-2 genes occurred in their common ancestor 13-17 million years ago (MYA), soon after the divergence of Arabidopsis and Brassica (15-20 MYA). This time of divergence is identical to the previously reported triplicated time of paralogous subgenomes of diploid Brassica species and the divergence date of group-1 and group-2 genes of α-carboxyltransferase, another subunit of heteromeric ACCase, in Brassica. Reverse transcription PCR revealed that the expression level of group-1 and group-2 genes varied in different organs, and the expression patterns of the two groups of genes were similar in different organs, except in flower. However, two paralogs of group-2 BC genes from B. napus could express differently in mature plants tested by generating BnaA.BC.b and BnaC.BC.b promoter-β-glucuronidase (GUS) fusions. The amino acid sequences of proteins encoded by these genes were highly conserved, except the sequence encoding

  3. 75 FR 37385 - Endangered and Threatened Species; Initiation of a 5-Year Review of the Eastern Distinct...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-29

    ..., population trends, distribution, abundance, demographics, and genetics; (B) habitat conditions, including... the Recognition of Distinct Vertebrate in Population Segments (61 FR 4722). For a population to be... of a 5-Year Review of the Eastern Distinct Population Segment of the Steller Sea Lion...

  4. Binding of EBP50 to Nox organizing subunit p47phox is pivotal to cellular reactive species generation and altered vascular phenotype.

    PubMed

    Al Ghouleh, Imad; Meijles, Daniel N; Mutchler, Stephanie; Zhang, Qiangmin; Sahoo, Sanghamitra; Gorelova, Anastasia; Henrich Amaral, Jefferson; Rodríguez, Andrés I; Mamonova, Tatyana; Song, Gyun Jee; Bisello, Alessandro; Friedman, Peter A; Cifuentes-Pagano, M Eugenia; Pagano, Patrick J

    2016-09-01

    Despite numerous reports implicating NADPH oxidases (Nox) in the pathogenesis of many diseases, precise regulation of this family of professional reactive oxygen species (ROS) producers remains unclear. A unique member of this family, Nox1 oxidase, functions as either a canonical or hybrid system using Nox organizing subunit 1 (NoxO1) or p47(phox), respectively, the latter of which is functional in vascular smooth muscle cells (VSMC). In this manuscript, we identify critical requirement of ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50; aka NHERF1) for Nox1 activation and downstream responses. Superoxide (O2 (•-)) production induced by angiotensin II (AngII) was absent in mouse EBP50 KO VSMC vs. WT. Moreover, ex vivo incubation of aortas with AngII showed a significant increase in O2 (•-) in WT but not EBP50 or Nox1 nulls. Similarly, lipopolysaccharide (LPS)-induced oxidative stress was attenuated in femoral arteries from EBP50 KO vs. WT. In silico analyses confirmed by confocal microscopy, immunoprecipitation, proximity ligation assay, FRET, and gain-/loss-of-function mutagenesis revealed binding of EBP50, via its PDZ domains, to a specific motif in p47(phox) Functional studies revealed AngII-induced hypertrophy was absent in EBP50 KOs, and in VSMC overexpressing EBP50, Nox1 gene silencing abolished VSMC hypertrophy. Finally, ex vivo measurement of lumen diameter in mouse resistance arteries exhibited attenuated AngII-induced vasoconstriction in EBP50 KO vs. WT. Taken together, our data identify EBP50 as a previously unidentified regulator of Nox1 and support that it promotes Nox1 activity by binding p47(phox) This interaction is pivotal for agonist-induced smooth muscle ROS, hypertrophy, and vasoconstriction and has implications for ROS-mediated physiological and pathophysiological processes.

  5. Binding of EBP50 to Nox organizing subunit p47phox is pivotal to cellular reactive species generation and altered vascular phenotype.

    PubMed

    Al Ghouleh, Imad; Meijles, Daniel N; Mutchler, Stephanie; Zhang, Qiangmin; Sahoo, Sanghamitra; Gorelova, Anastasia; Henrich Amaral, Jefferson; Rodríguez, Andrés I; Mamonova, Tatyana; Song, Gyun Jee; Bisello, Alessandro; Friedman, Peter A; Cifuentes-Pagano, M Eugenia; Pagano, Patrick J

    2016-09-01

    Despite numerous reports implicating NADPH oxidases (Nox) in the pathogenesis of many diseases, precise regulation of this family of professional reactive oxygen species (ROS) producers remains unclear. A unique member of this family, Nox1 oxidase, functions as either a canonical or hybrid system using Nox organizing subunit 1 (NoxO1) or p47(phox), respectively, the latter of which is functional in vascular smooth muscle cells (VSMC). In this manuscript, we identify critical requirement of ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50; aka NHERF1) for Nox1 activation and downstream responses. Superoxide (O2 (•-)) production induced by angiotensin II (AngII) was absent in mouse EBP50 KO VSMC vs. WT. Moreover, ex vivo incubation of aortas with AngII showed a significant increase in O2 (•-) in WT but not EBP50 or Nox1 nulls. Similarly, lipopolysaccharide (LPS)-induced oxidative stress was attenuated in femoral arteries from EBP50 KO vs. WT. In silico analyses confirmed by confocal microscopy, immunoprecipitation, proximity ligation assay, FRET, and gain-/loss-of-function mutagenesis revealed binding of EBP50, via its PDZ domains, to a specific motif in p47(phox) Functional studies revealed AngII-induced hypertrophy was absent in EBP50 KOs, and in VSMC overexpressing EBP50, Nox1 gene silencing abolished VSMC hypertrophy. Finally, ex vivo measurement of lumen diameter in mouse resistance arteries exhibited attenuated AngII-induced vasoconstriction in EBP50 KO vs. WT. Taken together, our data identify EBP50 as a previously unidentified regulator of Nox1 and support that it promotes Nox1 activity by binding p47(phox) This interaction is pivotal for agonist-induced smooth muscle ROS, hypertrophy, and vasoconstriction and has implications for ROS-mediated physiological and pathophysiological processes. PMID:27540115

  6. Isotopic 'fingerprinting' of distinct water reservoirs in the critical zone and their exploitation by different tree species

    NASA Astrophysics Data System (ADS)

    Oshun, J.; Dietrich, W. E.; Dawson, T. E.; Rempe, D. M.; Fung, I. Y.

    2013-12-01

    Channel incision, surface erosion, and drainage of landscapes leads to emergent hillslopes underlain by varying degrees of soil development and weathered bedrock. Vertical and lateral differentiation by weathering leads to a structured heterogeneity of soil and rock properties that affect water pathways, moisture retention, and moisture availability to plants. Near surface evaporative losses cause seasonal isotopic enrichment of soil water, but we asked, what is the isotopic pattern of the deeper moisture and how important is it as a water source to the vegetation? Here, we report on the use of stable isotopic data to 'fingerprint' and then characterize the structure of subsurface water reservoirs that lie beneath an old-growth forested hillslope in the Eel River basin of northern California. The site, Rivendell, a 4000 m2 heavily instrumented, unchannled basin draining to Elder Creek 0.5 km upstream from the confluence with the South Fork Eel River, is composed of turbidite sequences of vertically dipping mudstone and minor sandstone. Soils are thin (0-60 cm), and are underlain by a 1-4 m thick saprolite layer, and a 3-20 m thick weathered rock zone. Annual precipitation of 1900 mm, predominantly from October to May, raises the water table from summer lows of 5-28 m below the ground to winter highs of 2.5 m below the ground, with depth to the water table increasing with upslope distance from Elder Creek. Whereas the north-facing slope has a dominant forest canopy composed of Douglas-fir (Pseudotsuga menziesii), and a mid-story of hardwood species (Quercus agrifolia, Umbellularia californica, Notholithocarpus densiflorus, Arbutus menziesii, and the riparian tree, Acer macrophyllum), the south-facing slope, which drains directly to the South Fork Eel River, is primarily hardwood. Intensive sampling over 20 months reveals subsurface moisture reservoirs in the mobile and bulk water of the soil, saprolite, and weathered rock, with persistent and distinct isotopic

  7. Taxonomy and Phylogeny of the Fusarium dimerum Species Group

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The morphospecies Fusarium dimerum, known only from its anamorph, comprises at least 12 phylogenetically distinct species. Analyses of the large subunit ribosomal DNA (LSU rDNA) show they are taxa of the Nectriaceae (Hypocreales) and form a phylogenetically distinct clade within Fusarium. Accordin...

  8. [Molecular cloning of the DNA sequence of activin beta A subunit gene mature peptides from panda and related species and its application in the research of phylogeny and taxonomy].

    PubMed

    Wang, Xiao-Jing; Wang, Xiao-Xing; Wang, Ya-Jun; Wang, Xi-Zhong; He, Guang-Xin; Chen, Hong-Wei; Fei, Li-Song

    2002-09-01

    Activin, which is included in the transforming growth factor-beta (TGF beta) superfamily of proteins and receptors, is known to have broad-ranging effects in the creatures. The mature peptide of beta A subunit of this gene, one of the most highly conserved sequence, can elevate the basal secretion of follicle-stimulating hormone (FSH) in the pituitary and FSH is pivotal to organism's reproduction. Reproduction block is one of the main reasons which cause giant panda to extinct. The sequence of Activin beta A subunit gene mature peptides has been successfully amplified from giant panda, red panda and malayan sun bear's genomic DNA by using polymerase chain reaction (PCR) with a pair of degenerate primers. The PCR products were cloned into the vector pBlueScript+ of Esherichia coli. Sequence analysis of Activin beta A subunit gene mature peptides shows that the length of this gene segment is the same (359 bp) and there is no intron in all three species. The sequence encodes a peptide of 119 amino acid residues. The homology comparison demonstrates 93.9% DNA homology and 99% homology in amino acid among these three species. Both GenBank blast search result and restriction enzyme map reveal that the sequences of Activin beta A subunit gene mature peptides of different species are highly conserved during the evolution process. Phylogeny analysis is performed with PHYLIP software package. A consistent phylogeny tree has been drawn with three different methods. The software analysis outcome accords with the academic view that giant panda has a closer relationship to the malayan sun bear than the red panda. Giant panda should be grouped into the bear family (Uersidae) with the malayan sun bear. As to the red panda, it would be better that this animal be grouped into the unique family (red panda family) because of great difference between the red panda and the bears (Uersidae).

  9. 76 FR 76386 - Endangered and Threatened Species; 5-Year Reviews for 4 Distinct Population Segments of Steelhead...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-07

    ... Pacific salmon ESUs and steelhead DPSs in California, Oregon, Washington, and Idaho (75 FR 13082). Both... Reviews for 4 Distinct Population Segments of Steelhead in California AGENCY: National Marine Fisheries... Viable Salmonid Population framework, which relies on evaluating four key population...

  10. 75 FR 53272 - Endangered and Threatened Species; Initiation of 5-Year Review of the Eastern Distinct Population...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-31

    ... requesting information related to that review (75 FR 37385). A notice correcting the email address and fax number to which comments and information should be sent was published July 7, 2010 (75 FR 38979). Written... of 5-Year Review of the Eastern Distinct Population Segment of the Steller Sea Lion AGENCY:...

  11. Comparison of four species-delimitation methods applied to a DNA barcode data set of insect larvae for use in routine bioassessment for use in routine bioassessment

    EPA Science Inventory

    Species delimitation (grouping individuals into distinct taxonomic groups) is an essential part of evolutionary, conservation, and molecular ecology. Deoxyribonucleic acid (DNA) barcodes, short fragments of the cytochrome c oxidase subunit I (COI) gene, are being used in environm...

  12. Super-resolution fluorescence of huntingtin reveals growth of globular species into short fibers and coexistence of distinct aggregates.

    PubMed

    Duim, Whitney C; Jiang, Yan; Shen, Koning; Frydman, Judith; Moerner, W E

    2014-12-19

    Polyglutamine-expanded huntingtin, the protein encoded by HTT mutations associated with Huntington's disease, forms aggregate species in vitro and in vivo. Elucidation of the mechanism of growth of fibrillar aggregates from soluble monomeric protein is critical to understanding the progression of Huntington's disease and to designing therapeutics for the disease, as well as for aggregates implicated in Alzheimer's and Parkinson's diseases. We used the technique of multicolor single-molecule, super-resolution fluorescence imaging to characterize the growth of huntingtin exon 1 aggregates. The huntingtin exon 1 aggregation followed a pathway from exclusively spherical or globular species of ∼80 nm to fibers ∼1 μm in length that increased in width, but not length, over time with the addition of more huntingtin monomers. The fibers further aggregated with one another into aggregate assemblies of increasing size. Seeds created by sonication, which were comparable in shape and size to the globular species in the pathway, were observed to grow through multidirectional elongation into fibers, suggesting a mechanism for growth of globular species into fibers. The single-molecule sensitivity of our approach made it possible to characterize the aggregation pathway across a large range of size scales, from monomers to fiber assemblies, and revealed the coexistence of different aggregate species (globular species, fibers, fiber assemblies) even at late time points. PMID:25330023

  13. Super-Resolution Fluorescence of Huntingtin Reveals Growth of Globular Species into Short Fibers and Coexistence of Distinct Aggregates

    PubMed Central

    2015-01-01

    Polyglutamine-expanded huntingtin, the protein encoded by HTT mutations associated with Huntington’s disease, forms aggregate species in vitro and in vivo. Elucidation of the mechanism of growth of fibrillar aggregates from soluble monomeric protein is critical to understanding the progression of Huntington’s disease and to designing therapeutics for the disease, as well as for aggregates implicated in Alzheimer’s and Parkinson’s diseases. We used the technique of multicolor single-molecule, super-resolution fluorescence imaging to characterize the growth of huntingtin exon 1 aggregates. The huntingtin exon 1 aggregation followed a pathway from exclusively spherical or globular species of ∼80 nm to fibers ∼1 μm in length that increased in width, but not length, over time with the addition of more huntingtin monomers. The fibers further aggregated with one another into aggregate assemblies of increasing size. Seeds created by sonication, which were comparable in shape and size to the globular species in the pathway, were observed to grow through multidirectional elongation into fibers, suggesting a mechanism for growth of globular species into fibers. The single-molecule sensitivity of our approach made it possible to characterize the aggregation pathway across a large range of size scales, from monomers to fiber assemblies, and revealed the coexistence of different aggregate species (globular species, fibers, fiber assemblies) even at late time points. PMID:25330023

  14. Possible natural hybridization of two morphologically distinct species of Acropora (Cnidaria, Scleractinia) in the Pacific: fertilization and larval survival rates.

    PubMed

    Isomura, Naoko; Iwao, Kenji; Fukami, Hironobu

    2013-01-01

    Natural hybridization of corals in the Indo-Pacific has been considered rather rare. However, field studies have observed many corals with intermediate interspecific or unusual morphologies. Given that the existence of F1 hybrids with intermediate interspecific morphologies has been proven in the Caribbean, hybrids may also inhabit the Indo-Pacific and occur more frequently than expected. In this study, we focused on two morphologically different species, Acropora florida and A. intermedia, and performed crossing experiments at Akajima Island, Japan. Results showed that these species could hybridize in both directions via eggs and sperm, but that fertilization rates significantly differed according to which species provided eggs. These results are similar to those reported from the Caribbean. Although all embryos developed normally to the planular larval stage, the developmental processes of some hybrid embryos were delayed by approximately 1 h compared with conspecific embryos, suggesting that fertilization occurred 1 h later in interspecific crosses than in intraspecific crosses. More successful hybridization could occur under conditions with low numbers of conspecific colonies. Additionally, a comparison of survival rates between hybrid and intraspecific larvae revealed that intra- and interspecific larvae produced from eggs of A. florida survived for significantly longer than those produced from eggs of A. intermedia. Considering these data, under specific conditions, hybrids can be expected to be produced and survive in nature in the Pacific. Furthermore, we identified one colony with intermediate morphology between A. florida and A. intermedia in the field. This colony was fertilized only by eggs of A. florida, with high fertilization rates, suggesting that this colony would be a hybrid of these two species and might be backcrossed.

  15. Involvement of proteasomal subunits zeta and iota in RNA degradation.

    PubMed Central

    Petit, F; Jarrousse, A S; Dahlmann, B; Sobek, A; Hendil, K B; Buri, J; Briand, Y; Schmid, H P

    1997-01-01

    We have identified two distinct subunits of 20 S proteasomes that are associated with RNase activity. Proteasome subunits zeta and iota, eluted from two-dimensional Western blots, hydrolysed tobacco mosaic virus RNA, whereas none of the other subunits degraded this substrate under the same conditions. Additionally, proteasomes were dissociated by 6 M urea, and subunit zeta, containing the highest RNase activity, was isolated by anion-exchange chromatography and gel filtration. Purified subunit zeta migrated as a single spot on two-dimensional PAGE with a molecular mass of approx. 28 kDa. Addition of anti-(subunit zeta) antibodies led to the co-precipitation of this proteasome subunit and nuclease activity. This is the first evidence that proteasomal alpha-type subunits are associated with an enzymic activity, and our results provide further evidence that proteasomes may be involved in cellular RNA metabolism. PMID:9337855

  16. Subunit compositions of Arabidopsis RNA polymerases I and III reveal Pol I- and Pol III-specific forms of the AC40 subunit and alternative forms of the C53 subunit

    SciTech Connect

    Ream, Thomas S.; Haag, Jeremy R.; Pontvianne, Frederic; Nicora, Carrie D.; Norbeck, Angela D.; Pasa-Tolic, Ljiljana; Pikaard, Craig S.

    2015-05-02

    Using affinity purification and mass spectrometry, we identified the subunits of Arabidopsis thaliana multisubunit RNA Polymerases I and III (abbreviated as Pol I and Pol III), providing the first description of their physical compositions in plants. AC40 and AC19 subunits are typically common to Pol I (a.k.a. Pol A) and Pol III (a.k.a. Pol C) and are encoded by single genes whose mutation, in humans, is a cause of the craniofacial disorder, Treacher-Collins Syndrome. Surprisingly, A. thaliana, and related species, express two distinct AC40 paralogs, one of which assembles into Pol I and the other of which assembles into Pol III. Changes at eight amino acid positions correlate with this functional divergence of Pol I and Pol III-specific AC40 paralogs. Two genes encode homologs of the yeast C53 subunit, and either variant can assemble into Pol III. By contrast, only one of two potential C17 variants, and one of two potential C31 variants were detected in Pol III. We introduce a new nomenclature system for plant Pol I and Pol III subunits in which the twelve subunits that are structurally and functionally homologous among Pols I through V are assigned equivalent numbers.

  17. Subunit compositions of Arabidopsis RNA polymerases I and III reveal Pol I- and Pol III-specific forms of the AC40 subunit and alternative forms of the C53 subunit

    DOE PAGES

    Ream, Thomas S.; Haag, Jeremy R.; Pontvianne, Frederic; Nicora, Carrie D.; Norbeck, Angela D.; Pasa-Tolic, Ljiljana; Pikaard, Craig S.

    2015-05-02

    Using affinity purification and mass spectrometry, we identified the subunits of Arabidopsis thaliana multisubunit RNA Polymerases I and III (abbreviated as Pol I and Pol III), providing the first description of their physical compositions in plants. AC40 and AC19 subunits are typically common to Pol I (a.k.a. Pol A) and Pol III (a.k.a. Pol C) and are encoded by single genes whose mutation, in humans, is a cause of the craniofacial disorder, Treacher-Collins Syndrome. Surprisingly, A. thaliana, and related species, express two distinct AC40 paralogs, one of which assembles into Pol I and the other of which assembles into Polmore » III. Changes at eight amino acid positions correlate with this functional divergence of Pol I and Pol III-specific AC40 paralogs. Two genes encode homologs of the yeast C53 subunit, and either variant can assemble into Pol III. By contrast, only one of two potential C17 variants, and one of two potential C31 variants were detected in Pol III. We introduce a new nomenclature system for plant Pol I and Pol III subunits in which the twelve subunits that are structurally and functionally homologous among Pols I through V are assigned equivalent numbers.« less

  18. Photosynthesis of Grass Species Differing in Carbon Dioxide Fixation Pathways : VII. CHROMOSOME NUMBERS, METAPHASE I CHROMOSOME BEHAVIOR, AND MODE OF REPRODUCTION OF PHOTOSYNTHETICALLY DISTINCT PANICUM SPECIES.

    PubMed

    Bouton, J H; Brown, R H; Bolton, J K; Campagnoli, R P

    1981-03-01

    Panicum species of the Laxa group were investigated in a series of published reports and were found to possess C(4), C(3), and intermediate photosynthetic characteristics. Taxonomic and other relationships among these plants, however, are not clear. It was the objective of this investigation to document chromosome number, metaphase I chromosome behavior, and mode of reproduction, including abnormalities in the embryo sac, for these species.Chromosome counts showed a basic number (x) of 10 and ploidy levels of diploid (2n = 2x = 20), tetraploid (2n = 4x = 40), and hexaploid (2n = 6x = 60) in this group of Panicum. One diploid and one tetraploid accession of the C(4) species, Panicum prionitis Griseb., were obtained. Of the intermediate species, Panicum milioides Nees ex Trin. was diploid, Panicum schenckii Hack. was hexaploid, and Panicum decipiens Nees, ex Trin. was found to possess two ploidy levels, one accession being diploid and the other accession being hexaploid. All the C(3) species, which included two accessions of Panicum laxum Sw., three accessions of Panicum hylaeicum Mez., and one accession of Panicum rivulare Trin., were tetraploid.Meiosis was regular with primarily bivalent pairing at metaphase I in all species except the tetraploid accession of P. prionitis which possessed from 4 to 10 tetravalents. Stainable pollen was high in all species, ranging from 70 to 99%. Embryo sac analyses showed a single sac in all plants except the tetraploid accession of P. prionitis, which was found to possess an additional sac at anthesis. An additional sac was also observed in some ovaries of the P. schenckii accession. Self-pollinated seed set was high in all accessions except the diploid accession of P. prionitis and one accession of P. laxum where no seed was set under bagged conditions.This information establishes, within the limits of this collection, a base for future studies on genetic, taxonomic, photosynthetic, and evolutionary relationships among these

  19. Autoimmune epilepsy: distinct subpopulations of epilepsy patients harbor serum autoantibodies to either glutamate/AMPA receptor GluR3, glutamate/NMDA receptor subunit NR2A or double-stranded DNA.

    PubMed

    Ganor, Yonatan; Goldberg-Stern, Hadassa; Lerman-Sagie, Tally; Teichberg, Vivian I; Levite, Mia

    2005-06-01

    We studied 82 patients with different types of epilepsy and 49 neurologically intact non-epileptic controls, and identified three different subpopulations of epilepsy patients bearing significantly elevated levels of autoantibodies to either GluR3B-peptide of glutamate/AMPA receptor subtype 3 (17/82; 21% of patients), or to a peptide of NR2A subunit of glutamate/NMDA receptors (15/82; 18%), or to double-stranded (ds) DNA, the hallmark of systemic lupus erythematosus (13/80; 16%). Most patients had only one antibody type, arguing against cross-reactivity. Nearly all anti-dsDNA Ab-positive patients did not harbor anti-nuclear autoantibodies. Most patients had no history of brain damage, febrile convulsions, early onset epilepsy, acute epilepsy or intractable seizures. We suggest to measure the 'autoimmune-fingerprints' of epilepsy patients for diagnostic and therapeutic purposes. PMID:15978777

  20. The molecular dimension of microbial species: 1. Ecological distinctions among, and homogeneity within, putative ecotypes of Synechococcus inhabiting the cyanobacterial mat of Mushroom Spring, Yellowstone National Park.

    PubMed

    Becraft, Eric D; Wood, Jason M; Rusch, Douglas B; Kühl, Michael; Jensen, Sheila I; Bryant, Donald A; Roberts, David W; Cohan, Frederick M; Ward, David M

    2015-01-01

    Based on the Stable Ecotype Model, evolution leads to the divergence of ecologically distinct populations (e.g., with different niches and/or behaviors) of ecologically interchangeable membership. In this study, pyrosequencing was used to provide deep sequence coverage of Synechococcus psaA genes and transcripts over a large number of habitat types in the Mushroom Spring microbial mat. Putative ecological species [putative ecotypes (PEs)], which were predicted by an evolutionary simulation based on the Stable Ecotype Model (Ecotype Simulation), exhibited distinct distributions relative to temperature-defined positions in the effluent channel and vertical position in the upper 1 mm-thick mat layer. Importantly, in most cases variants predicted to belong to the same PE formed unique clusters relative to temperature and depth in the mat in canonical correspondence analysis, supporting the hypothesis that while the PEs are ecologically distinct, the members of each ecotype are ecologically homogeneous. PEs responded differently to experimental perturbations of temperature and light, but the genetic variation within each PE was maintained as the relative abundances of PEs changed, further indicating that each population responded as a set of ecologically interchangeable individuals. Compared to PEs that predominate deeper within the mat photic zone, the timing of transcript abundances for selected genes differed for PEs that predominate in microenvironments closer to upper surface of the mat with spatiotemporal differences in light and O2 concentration. All of these findings are consistent with the hypotheses that Synechococcus species in hot spring mats are sets of ecologically interchangeable individuals that are differently adapted, that these adaptations control their distributions, and that the resulting distributions constrain the activities of the species in space and time.

  1. The molecular dimension of microbial species: 1. Ecological distinctions among, and homogeneity within, putative ecotypes of Synechococcus inhabiting the cyanobacterial mat of Mushroom Spring, Yellowstone National Park

    PubMed Central

    Becraft, Eric D.; Wood, Jason M.; Rusch, Douglas B.; Kühl, Michael; Jensen, Sheila I.; Bryant, Donald A.; Roberts, David W.; Cohan, Frederick M.; Ward, David M.

    2015-01-01

    Based on the Stable Ecotype Model, evolution leads to the divergence of ecologically distinct populations (e.g., with different niches and/or behaviors) of ecologically interchangeable membership. In this study, pyrosequencing was used to provide deep sequence coverage of Synechococcus psaA genes and transcripts over a large number of habitat types in the Mushroom Spring microbial mat. Putative ecological species [putative ecotypes (PEs)], which were predicted by an evolutionary simulation based on the Stable Ecotype Model (Ecotype Simulation), exhibited distinct distributions relative to temperature-defined positions in the effluent channel and vertical position in the upper 1 mm-thick mat layer. Importantly, in most cases variants predicted to belong to the same PE formed unique clusters relative to temperature and depth in the mat in canonical correspondence analysis, supporting the hypothesis that while the PEs are ecologically distinct, the members of each ecotype are ecologically homogeneous. PEs responded differently to experimental perturbations of temperature and light, but the genetic variation within each PE was maintained as the relative abundances of PEs changed, further indicating that each population responded as a set of ecologically interchangeable individuals. Compared to PEs that predominate deeper within the mat photic zone, the timing of transcript abundances for selected genes differed for PEs that predominate in microenvironments closer to upper surface of the mat with spatiotemporal differences in light and O2 concentration. All of these findings are consistent with the hypotheses that Synechococcus species in hot spring mats are sets of ecologically interchangeable individuals that are differently adapted, that these adaptations control their distributions, and that the resulting distributions constrain the activities of the species in space and time. PMID:26157420

  2. Photolysis of Hi-CO Nitrogenase – Observation of a Plethora of Distinct CO Species using Infrared Spectroscopy

    PubMed Central

    Yan, Lifen; Dapper, Christie H.; George, Simon J.; Wang, Hongxin; Mitra, Devrani; Dong, Weibing; Newton, William E.; Cramer, Stephen P.

    2015-01-01

    Fourier transform infrared spectroscopy (FT-IR) was used to study the photochemistry of CO-inhibited Azotobacter vinelandii nitrogenase using visible light at cryogenic temperatures. The FT-IR difference spectrum of photolyzed hi-CO at 4 K comprises negative bands at 1973 cm−1 and 1679 cm−1 together with positive bands at 1711 cm−1, 2135 and 2123 cm−1. The negative bands are assigned to a hi-CO state that comprises 2 metal-bound CO ligands, one terminally bound, and one bridged and/or protonated species. The positive band at 1711 cm−1 is assigned to a lo-CO product with a single bridged and/or protonated metal-CO group. We term these species ‘Hi-1’ and ‘Lo-1’ respectively. The high-energy bands are assigned to a liberated CO trapped in the protein pocket. Warming results in CO recombination, and the temperature dependence of the recombination rate yields an activation energy of 4 kJ mol−1. Two α-H195 variant enzymes yielded additional signals. Asparagine substitution, α-H195N, gives a spectrum containing 2 negative ‘Hi-2’ bands at 1936 and 1858 cm−1 with a positive ‘Lo-2’ band at 1780 cm−1, while glutamine substitution, α-H195Q, produces a complex spectrum that includes a third CO species, with negative ‘Hi-3’ bands at 1938 and 1911 cm−1 and a positive feature ‘Lo-3’ band at 1921 cm−1. These species can be assigned to a combination of terminal, bridged, and possibly protonated CO groups bound to the FeMo-cofactor active site. The proposed structures are discussed in terms of both CO inhibition and the mechanism nitrogenase catalysis. Given the intractability of observing nitrogenase intermediates by crystallographic methods, IR-monitored photolysis appears to be a promising and information-rich probe of nitrogenase structure and chemistry. PMID:27630531

  3. Adaptation of the Mitochondrial Genome in Cephalopods: Enhancing Proton Translocation Channels and the Subunit Interactions

    PubMed Central

    Almeida, Daniela; Maldonado, Emanuel; Vasconcelos, Vitor; Antunes, Agostinho

    2015-01-01

    Mitochondrial protein-coding genes (mt genes) encode subunits forming complexes of crucial cellular pathways, including those involved in the vital process of oxidative phosphorylation (OXPHOS). Despite the vital role of the mitochondrial genome (mt genome) in the survival of organisms, little is known with respect to its adaptive implications within marine invertebrates. The molluscan Class Cephalopoda is represented by a marine group of species known to occupy contrasting environments ranging from the intertidal to the deep sea, having distinct metabolic requirements, varied body shapes and highly advanced visual and nervous systems that make them highly competitive and successful worldwide predators. Thus, cephalopods are valuable models for testing natural selection acting on their mitochondrial subunits (mt subunits). Here, we used concatenated mt genes from 17 fully sequenced mt genomes of diverse cephalopod species to generate a robust mitochondrial phylogeny for the Class Cephalopoda. We followed an integrative approach considering several branches of interest–covering cephalopods with distinct morphologies, metabolic rates and habitats–to identify sites under positive selection and localize them in the respective protein alignment and/or tridimensional structure of the mt subunits. Our results revealed significant adaptive variation in several mt subunits involved in the energy production pathway of cephalopods: ND5 and ND6 from Complex I, CYTB from Complex III, COX2 and COX3 from Complex IV, and in ATP8 from Complex V. Furthermore, we identified relevant sites involved in protein-interactions, lining proton translocation channels, as well as disease/deficiencies related sites in the aforementioned complexes. A particular case, revealed by this study, is the involvement of some positively selected sites, found in Octopoda lineage in lining proton translocation channels (site 74 from ND5) and in interactions between subunits (site 507 from ND5) of

  4. Adaptation of the Mitochondrial Genome in Cephalopods: Enhancing Proton Translocation Channels and the Subunit Interactions.

    PubMed

    Almeida, Daniela; Maldonado, Emanuel; Vasconcelos, Vitor; Antunes, Agostinho

    2015-01-01

    Mitochondrial protein-coding genes (mt genes) encode subunits forming complexes of crucial cellular pathways, including those involved in the vital process of oxidative phosphorylation (OXPHOS). Despite the vital role of the mitochondrial genome (mt genome) in the survival of organisms, little is known with respect to its adaptive implications within marine invertebrates. The molluscan Class Cephalopoda is represented by a marine group of species known to occupy contrasting environments ranging from the intertidal to the deep sea, having distinct metabolic requirements, varied body shapes and highly advanced visual and nervous systems that make them highly competitive and successful worldwide predators. Thus, cephalopods are valuable models for testing natural selection acting on their mitochondrial subunits (mt subunits). Here, we used concatenated mt genes from 17 fully sequenced mt genomes of diverse cephalopod species to generate a robust mitochondrial phylogeny for the Class Cephalopoda. We followed an integrative approach considering several branches of interest-covering cephalopods with distinct morphologies, metabolic rates and habitats-to identify sites under positive selection and localize them in the respective protein alignment and/or tridimensional structure of the mt subunits. Our results revealed significant adaptive variation in several mt subunits involved in the energy production pathway of cephalopods: ND5 and ND6 from Complex I, CYTB from Complex III, COX2 and COX3 from Complex IV, and in ATP8 from Complex V. Furthermore, we identified relevant sites involved in protein-interactions, lining proton translocation channels, as well as disease/deficiencies related sites in the aforementioned complexes. A particular case, revealed by this study, is the involvement of some positively selected sites, found in Octopoda lineage in lining proton translocation channels (site 74 from ND5) and in interactions between subunits (site 507 from ND5) of Complex I

  5. Adaptation of the Mitochondrial Genome in Cephalopods: Enhancing Proton Translocation Channels and the Subunit Interactions.

    PubMed

    Almeida, Daniela; Maldonado, Emanuel; Vasconcelos, Vitor; Antunes, Agostinho

    2015-01-01

    Mitochondrial protein-coding genes (mt genes) encode subunits forming complexes of crucial cellular pathways, including those involved in the vital process of oxidative phosphorylation (OXPHOS). Despite the vital role of the mitochondrial genome (mt genome) in the survival of organisms, little is known with respect to its adaptive implications within marine invertebrates. The molluscan Class Cephalopoda is represented by a marine group of species known to occupy contrasting environments ranging from the intertidal to the deep sea, having distinct metabolic requirements, varied body shapes and highly advanced visual and nervous systems that make them highly competitive and successful worldwide predators. Thus, cephalopods are valuable models for testing natural selection acting on their mitochondrial subunits (mt subunits). Here, we used concatenated mt genes from 17 fully sequenced mt genomes of diverse cephalopod species to generate a robust mitochondrial phylogeny for the Class Cephalopoda. We followed an integrative approach considering several branches of interest-covering cephalopods with distinct morphologies, metabolic rates and habitats-to identify sites under positive selection and localize them in the respective protein alignment and/or tridimensional structure of the mt subunits. Our results revealed significant adaptive variation in several mt subunits involved in the energy production pathway of cephalopods: ND5 and ND6 from Complex I, CYTB from Complex III, COX2 and COX3 from Complex IV, and in ATP8 from Complex V. Furthermore, we identified relevant sites involved in protein-interactions, lining proton translocation channels, as well as disease/deficiencies related sites in the aforementioned complexes. A particular case, revealed by this study, is the involvement of some positively selected sites, found in Octopoda lineage in lining proton translocation channels (site 74 from ND5) and in interactions between subunits (site 507 from ND5) of Complex I.

  6. Distinct cell-specific expression of homospermidine synthase involved in pyrrolizidine alkaloid biosynthesis in three species of the boraginales.

    PubMed

    Niemüller, Daniel; Reimann, Andreas; Ober, Dietrich

    2012-07-01

    Homospermidine synthase (HSS) is the first specific enzyme in pyrrolizidine alkaloid (PA) biosynthesis, a pathway involved in the plant's chemical defense. HSS has been shown to be recruited repeatedly by duplication of a gene involved in primary metabolism. Within the lineage of the Boraginales, only one gene duplication event gave rise to HSS. Here, we demonstrate that the tissue-specific expression of HSS in three boraginaceous species, Heliotropium indicum, Symphytum officinale, and Cynoglossum officinale, is unique with respect to plant organ, tissue, and cell type. Within H. indicum, HSS is expressed exclusively in nonspecialized cells of the lower epidermis of young leaves and shoots. In S. officinale, HSS expression has been detected in the cells of the root endodermis and in leaves directly underneath developing inflorescences. In young roots of C. officinale, HSS is detected only in cells of the endodermis, but in a later developmental stage, additionally in the pericycle. The individual expression patterns are compared with those within the Senecioneae lineage (Asteraceae), where HSS expression is reproducibly found in specific cells of the endodermis and the adjacent cortex parenchyma of the roots. The individual expression patterns within the Boraginales species are discussed as being a requirement for the successful recruitment of HSS after gene duplication. The diversity of HSS expression within this lineage adds a further facet to the already diverse patterns of expression that have been observed for HSS in other PA-producing plant lineages, making this PA-specific enzyme one of the most diverse expressed proteins described in the literature. PMID:22566491

  7. Molecular identification of sibling species of Sclerodermus (Hymenoptera: Bethylidae) that parasitize buprestid and cerambycid beetles by using partial sequences of mitochondrial DNA cytochrome oxidase subunit 1 and 28S ribosomal RNA gene.

    PubMed

    Jiang, Yuan; Yang, Zhongqi; Wang, Xiaoyi; Hou, Yuxia

    2015-01-01

    The species belonging to Sclerodermus (Hymenoptera: Bethylidae) are currently the most important insect natural enemies of wood borer pests, mainly buprestid and cerambycid beetles, in China. However, some sibling species of this genus are very difficult to distinguish because of their similar morphological features. To address this issue, we conducted phylogenetic and genetic analyses of cytochrome oxidase subunit I (COI) and 28S RNA gene sequences from eight species of Sclerodermus reared from different wood borer pests. The eight sibling species were as follows: S. guani Xiao et Wu, S. sichuanensis Xiao, S. pupariae Yang et Yao, and Sclerodermus spp. (Nos. 1-5). A 594-bp fragment of COI and 750-bp fragment of 28S were subsequently sequenced. For COI, the G-C content was found to be low in all the species, averaging to about 30.0%. Sequence divergences (Kimura-2-parameter distances) between congeneric species averaged to 4.5%, and intraspecific divergences averaged to about 0.09%. Further, the maximum sequence divergences between congeneric species and Sclerodermus sp. (No. 5) averaged to about 16.5%. All 136 samples analyzed were included in six reciprocally monophyletic clades in the COI neighbor-joining (NJ) tree. The NJ tree inferred from the 28S rRNA sequence yielded almost identical results, but the samples from S. guani, S. sichuanensis, S. pupariae, and Sclerodermus spp. (Nos. 1-4) clustered together and only Sclerodermus sp. (No. 5) clustered separately. Our findings indicate that the standard barcode region of COI can be efficiently used to distinguish morphologically similar Sclerodermus species. Further, we speculate that Sclerodermus sp. (No. 5) might be a new species of Sclerodermus.

  8. Molecular Identification of Sibling Species of Sclerodermus (Hymenoptera: Bethylidae) That Parasitize Buprestid and Cerambycid Beetles by Using Partial Sequences of Mitochondrial DNA Cytochrome Oxidase Subunit 1 and 28S Ribosomal RNA Gene

    PubMed Central

    Jiang, Yuan; Yang, Zhongqi; Wang, Xiaoyi; Hou, Yuxia

    2015-01-01

    The species belonging to Sclerodermus (Hymenoptera: Bethylidae) are currently the most important insect natural enemies of wood borer pests, mainly buprestid and cerambycid beetles, in China. However, some sibling species of this genus are very difficult to distinguish because of their similar morphological features. To address this issue, we conducted phylogenetic and genetic analyses of cytochrome oxidase subunit I (COI) and 28S RNA gene sequences from eight species of Sclerodermus reared from different wood borer pests. The eight sibling species were as follows: S. guani Xiao et Wu, S. sichuanensis Xiao, S. pupariae Yang et Yao, and Sclerodermus spp. (Nos. 1–5). A 594-bp fragment of COI and 750-bp fragment of 28S were subsequently sequenced. For COI, the G-C content was found to be low in all the species, averaging to about 30.0%. Sequence divergences (Kimura-2-parameter distances) between congeneric species averaged to 4.5%, and intraspecific divergences averaged to about 0.09%. Further, the maximum sequence divergences between congeneric species and Sclerodermus sp. (No. 5) averaged to about 16.5%. All 136 samples analyzed were included in six reciprocally monophyletic clades in the COI neighbor-joining (NJ) tree. The NJ tree inferred from the 28S rRNA sequence yielded almost identical results, but the samples from S. guani, S. sichuanensis, S. pupariae, and Sclerodermus spp. (Nos. 1–4) clustered together and only Sclerodermus sp. (No. 5) clustered separately. Our findings indicate that the standard barcode region of COI can be efficiently used to distinguish morphologically similar Sclerodermus species. Further, we speculate that Sclerodermus sp. (No. 5) might be a new species of Sclerodermus. PMID:25782000

  9. Genetic variation in walnuts (Juglans regia and J. sigillata; Juglandaceae): Species distinctions, human impacts, and the conservation of agrobiodiversity in Yunnan, China.

    PubMed

    Gunn, Bee F; Aradhya, Mallikarjuna; Salick, Jan M; Miller, Allison J; Yongping, Yang; Lin, Liu; Xian, Hai

    2010-04-01

    Walnuts are a major crop of many countries and mostly cultivated in large-scale plantations with few cultivars. Landraces provide important genetic reservoirs; thus, understanding factors influencing the geographic distribution of genetic variation in crop resources is a fundamental goal of agrobiodiversity conservation. Here, we investigated the role of human settlements and kinship on genetic variation and population structure of two walnut species: Juglans regia, an introduced species widely cultivated for its nuts, and J. sigillata, a native species cultivated locally in Yunnan. The objectives of this study were to characterize sympatric populations of J. regia and J. sigillata using 14 molecular markers and evaluate the role of Tibetan villages and kin groups (related households) on genotypic variation and population structure of J. regia and J. sigillata. Our results based on 220 walnut trees from six Tibetan villages show that although J. regia and J. sigillata are morphologically distinct, the two species are indistinguishable based on microsatellite data. Despite the lack of interspecific differences, AMOVAs partitioned among villages (5.41%, P = 0.0068) and kin groups within villages (3.34%, P = 0.0068) showed significant genetic variation. These findings suggest that village environments and familial relationships are factors contributing to the geographic structure of genetic variation in Tibetan walnuts.

  10. Cu(I)-catalyzed diamination of conjugated dienes. Complementary regioselectivity from two distinct mechanistic pathways involving Cu(II) and Cu(III) species.

    PubMed

    Zhao, Baoguo; Peng, Xingao; Zhu, Yingguang; Ramirez, Thomas A; Cornwall, Richard G; Shi, Yian

    2011-12-28

    Conjugated dienes can be diaminated at the internal and/or terminal double bonds using Cu(I) as catalyst and N,N-di-t-butyldiaziridinone (1) as nitrogen source. The regioselectivity is highly dependent upon the choice of Cu(I) catalyst and the substituents on diene substrates. The diamination likely proceeds via two mechanistically distinct pathways. The N-N bond of N,N-di-t-butyldiaziridinone (1) is first homolytically cleaved by the Cu(I) catalyst to form four-membered Cu(III) species A and Cu(II) radical species B, which are in rapid equilibrium. The internal diamination likely proceeds in a concerted manner via Cu(III) species A, and the terminal diamination likely involves Cu(II) radical species B. Kinetic studies have shown that the diamination is first-order in N,N-di-t-butyldiaziridinone (1), zero-order in olefin, and first-order in total Cu(I) catalyst, and the cleavage of the N-N bond of 1 by the Cu(I) catalyst is the rate-determining step. The internal diamination is favored by use of CuBr without ligand and electron-rich dienes. The terminal diamination is favored when using CuCl-L and dienes with radical-stabilizing groups. PMID:22081888

  11. Cu(I)-Catalyzed Diamination of Conjugated Dienes. Complementary Regioselectivity from Two Distinct Mechanistic Pathways Involving Cu(II) and Cu(III) Species

    PubMed Central

    Zhao, Baoguo; Peng, Xingao; Zhu, Yingguang; Ramirez, Thomas A.; Cornwall, Richard G.

    2011-01-01

    Conjugated dienes can be diaminated at the internal and/or terminal double bonds using Cu(I) as catalyst and N,N-di-t-butyldiaziridinone (1) as nitrogen source. The regioselectivity is highly dependent upon the choice of Cu(I) catalyst and the substituents on diene substrates. The diamination likely proceeds via two mechanistically distinct pathways. The N-N bond of N,N-di-t-butyldiaziridinone (1) is first homolytically cleaved by the Cu(I) catalyst to form four-membered Cu(III) species A and Cu(II) radical species B, which are in rapid equilibrium. The internal diamination likely proceeds in a concerted manner via Cu(III) species A and the terminal diamination likely involves Cu(II) radical species B. Kinetic studies have shown that the diamination is first-order in N,N-di-t-butyldiaziridinone (1), zero-order in olefin, first-order in total Cu(I) catalyst, and the cleavage of the N-N bond of 1 by the Cu(I) catalyst is the rate-determining step. The internal diamination is favored by use of CuBr without ligand and electron-rich dienes. The terminal diamination is favored when using CuCl-L and dienes with radical-stabilizing groups. PMID:22081888

  12. Two distinct abnormalities in patients with C8 alpha-gamma deficiency. Low level of C8 beta chain and presence of dysfunctional C8 alpha-gamma subunit.

    PubMed Central

    Tedesco, F; Roncelli, L; Petersen, B H; Agnello, V; Sodetz, J M

    1990-01-01

    The sera from three C8 alpha-gamma deficient patients previously reported to have a selective C8 alpha-gamma defect were analyzed by SDS-PAGE and Western blot using two polyclonal antisera to C8 alpha-gamma and a monoclonal antibody to C8 alpha. All three sera exhibited C8 alpha-gamma bands that dissociated into alpha and gamma chains under reducing conditions. Quantitation of the alpha-gamma subunit in these sera by a sensitive ELISA revealed an amount approximately 1% of that found in normal human serum. A similar assay performed with a specific antiserum to C8 beta showed unexpectedly low levels of C8 beta in these sera, which were confirmed by hemolytic titration of C8 beta. The remarkable differences between C8 alpha-gamma and C8 beta in the C8 alpha-gamma deficient sera was that in spite of their comparable immunochemical levels, C8 beta still exhibited functional activity whereas C8 alpha-gamma was totally inactive. That the residual C8 alpha-gamma was inactive was also proved by its inability to show lytic bands in an overlay system after SDS-PAGE and subsequent removal of SDS. The implications of these findings for a novel concept of C8 deficiency are discussed. Images PMID:2394837

  13. High-valent nonheme iron. Two distinct iron(IV) species derived from a common iron(II) precursor.

    PubMed

    Jensen, Michael P; Costas, Miquel; Ho, Raymond Y N; Kaizer, József; Mairata i Payeras, Antoni; Münck, Eckard; Que, Lawrence; Rohde, Jan-Uwe; Stubna, Audria

    2005-08-01

    The reaction of [Fe(II)(beta-BPMCN)(OTf)2] (1, BPMCN = N,N'-bis(2-pyridylmethyl)-N,N'-dimethyl-trans-1,2-diaminocyclohexane) with tBuOOH at low-temperature yields alkylperoxoiron(III) intermediates 2 in CH2Cl2 and 2-NCMe in CH3CN. At -45 degrees C and above, 2-NCMe converts to a pale green species 3 (lambda(max) = 753 nm, epsilon = 280 M(-1) cm(-1)) in 90% yield, identified as [Fe(IV)(O)(BPMCN)(NCCH3)]2+ by comparison to other nonheme [Fe(IV)(O)(L)]2+ complexes. Below -55 degrees C in CH2Cl2, 2 decays instead to form deep turquoise 4 (lambda(max) = 656, 845 nm; epsilon = 4000, 3600 M(-1) cm(-1)), formulated to be an unprecedented alkylperoxoiron(IV) complex [Fe(IV)(BPMCN)(OH)(OOtBu)]2+ on the basis of Mössbauer, EXAFS, resonance Raman, NMR, and mass spectral evidence. The reactivity of 1 with tBuOOH in the two solvents reveals an unexpectedly rich iron(IV) chemistry that can be supported by the BPMCN ligand.

  14. Exploring the utility of phylogenetic analysis of cytochrome oxidase gene subunit I as a complementary tool to classical taxonomical identification of phlebotomine sand fly species (Diptera, Psychodidae) from southern Europe.

    PubMed

    Maia, Carla; Parreira, Ricardo; Cristóvão, José Manuel; Afonso, Maria Odete; Campino, Lenea

    2015-04-01

    Phlebotomine sand flies (Diptera, Psychodidae) are known to be vectors of several pathogens such as Leishmania and Phlebovirus genera. The identification of phlebotomine sand fly species is currently based on morphological characters, and requires considerable taxonomic expertise and skilfulness, but may be complemented by DNA-based analyses for (i) accurate species identification and (ii) for estimating sand fly diversity. The aim of this study was to evaluate the utility of mitochondrial cytochrome oxidase gene subunit I (cox1) sequence analysis as a complementary tool to classical taxonomical for the identification of the most prevalent phlebotomine sand fly species from southern Europe (i.e. Phlebotomus ariasi, P. perniciosus, P. sergenti and Sergentomyia minuta). Phylogenetic analyses of cox1 sequences allowed conclusive assignment of most of the sand flies into individual species, and revealed the genetic heterogeneity that characterizes some of the identified genetic clusters. Nevertheless, it showed some limitations, as it failed to (i) allocate correctly all of all species of a given subgenus to a single lineage, or (ii) conclusively identify sequences amplified from individuals classified morphologically as P. ariasi. A more extensive analysis of cox1 sequences together with morphometric characterization of specimens from different geographic areas/regions might be useful for the correct assessment of the phylogenetic relationship within the P. ariasi/P. chadlii cluster and/or help to ascertain the usefulness of cox1 for molecular taxonomy of sand flies.

  15. Local vasotocin modulation of the pacemaker nucleus resembles distinct electric behaviors in two species of weakly electric fish.

    PubMed

    Perrone, Rossana; Migliaro, Adriana; Comas, Virginia; Quintana, Laura; Borde, Michel; Silva, Ana

    2014-01-01

    The neural bases of social behavior diversity in vertebrates have evolved in close association with hypothalamic neuropeptides. In particular, arginine-vasotocin (AVT) is a key integrator underlying differences in behavior across vertebrate taxa. Behavioral displays in weakly electric fish are channeled through specific patterns in their electric organ discharges (EODs), whose rate is ultimately controlled by a medullary pacemaker nucleus (PN). We first explored interspecific differences in the role of AVT as modulator of electric behavior in terms of EOD rate between the solitary Gymnotus omarorum and the gregarious Brachyhypopomus gauderio. In both species, AVT IP injection (10μg/gbw) caused a progressive increase of EOD rate of about 30%, which was persistent in B. gauderio, and attenuated after 30min in G. omarorum. Secondly, we demonstrated by in vitro electrophysiological experiments that these behavioral differences can be accounted by dissimilar effects of AVT upon the PN in itself. AVT administration (1μM) to the perfusion bath of brainstem slices containing the PN produced a small and transient increase of PN activity rate in G. omarorum vs the larger and persistent increase previously reported in B. gauderio. We also identified AVT neurons, for the first time in electric fish, using immunohistochemistry techniques and confirmed the presence of hindbrain AVT projections close to the PN that might constitute the anatomical substrate for AVT influences on PN activity. Taken together, our data reinforce the view of the PN as an extremely plastic medullary central pattern generator that not only responds to higher influences to adapt its function to diverse contexts, but also is able to intrinsically shape its response to neuropeptide actions, thus adding a hindbrain target level to the complexity of the global integration of central neuromodulation of electric behavior. PMID:25125289

  16. Yeast telomerase RNA: a flexible scaffold for protein subunits.

    PubMed

    Zappulla, David C; Cech, Thomas R

    2004-07-01

    In the yeast Saccharomyces cerevisiae, distinct regions of the 1.2-kb telomerase RNA (TLC1) bind to the catalytic subunit Est2p and to accessory proteins. In particular, a bulged stem structure binds the essential regulatory subunit Est1p. We now show that the Est1p-binding domain of the RNA can be moved to three distant locations with retention of telomerase function in vivo. We present the Est1p relocation experiment in the context of a working model for the secondary structure of the entire TLC1 RNA, based on thermodynamic considerations and comparative analysis of sequences from four species. The model for TLC1 has three long quasihelical arms that bind the Ku, Est1p, and Sm proteins. These arms emanate from a central catalytic core that contains the template and Est2p-binding region. Deletion mutagenesis provides evidence that the Sm arm exists in vivo and can be shortened by 42 predicted base pairs with retention of function; therefore, precise positioning of Sm proteins, like Est1p, is not required within telomerase. In the best-studied ribonucleoprotein enzyme, the ribosome, the RNAs have specific three-dimensional structures that orient the functional elements. In the case of yeast telomerase, we propose that the RNA serves a very different function, providing a flexible tether for the protein subunits. PMID:15226497

  17. Characterization of Brassica nigra collections using simple sequence repeat markers reveals distinct groups associated with geographical location, and frequent mislabelling of species identity.

    PubMed

    Pradhan, Aneeta; Nelson, Matthew N; Plummer, Julie A; Cowling, Wallace A; Yan, Guijun

    2011-01-01

    Genetic diversity of 180 Brassica nigra (L.) Kochgenotypes from 60 different accessions was evaluated using 15 simple sequence repeat markers with known locations on the Brassica A, B, and C genomes. Two lines each from Brassica juncea (L.) Czern and Brassica carinata Braunwere also included as comparator species. A total of 218 high quality alleles were used to generate a genetic distance matrix, and clustering and multidimensional scaling analyses were used to investigate genetic relationships among the accessions. Accessions from the same country of origin tended to cluster together. Surprisingly, 13 accessions declared to be B. nigra had A- and B-genome alleles and morphology consistent with them being B. juncea, which was supported by their positioning near B. juncea in the cluster analysis. Two B. nigra accessions possessed alleles associated more closely with the A genome than the B genome, and these may be Brassica rapa L. accessions. One B. nigra accession had B- and C-genome alleles and morphology consistent with it being B. carinata. The remaining 44 accessions (73%) appeared to be truly B. nigra and formed morphologically and genetically distinct groups associated with country or region of origin, notably Ethiopia, Israel, India, and Europe. Most B. nigra accessions were highly heterozygous, consistent with their obligate outcrossing habit. This study demonstrated the value of using molecular markers with known genome locations (in this case, in the Brassica A, B, and C genomes) to confirm species identity in families such as Brassicaceae where species identification based solely on morphological characters is difficult.

  18. IDENTIFICATION OF SPECIES AND SOURCES OF CRYPTOSPORIDIUM OOCYSTS IN STORM WATERS BY A SMALL SUBUNIT RRNA-BASED DIAGNOSTIC AND GENOTYPING TOOL

    EPA Science Inventory

    The identification of Cryptosporidium oocysts in environmental samples is largely made by the use of immunofluorescent assay (IFA). because IFA detects oocysts from all Cryptosporidium parasites, the species distribution and source of Cryptosporidium parasites in environmental sa...

  19. IDENTIFICATION OF CRYPTOSPORIDIUM SPECIES AND THE SOURCES IN RAW WASTEWATER USING A SMALL SUBUNIT RRNA-BASED PCR-RFLP TOOL

    EPA Science Inventory

    The species composition and source of Cryptosporidium oocysts in wastewater have never been determined, even though it is widely assumed that these oocysts are from human sewage. Recent molecular characterizations of Cryptosporidium parasites make it possible to differentiate hum...

  20. Steady state or non-steady state? Identifying driving mechanisms of oxygen isotope signatures of leaf transpiration in functionally distinct plant species

    NASA Astrophysics Data System (ADS)

    Dubbert, Maren; Kübert, Angelika; Cuntz, Matthias; Werner, Christiane

    2015-04-01

    Isotope techniques are widely applied in ecosystem studies. For example, isoflux models are used to separate soil evaporation from transpiration in ecosystems. These models often assume that plant transpiration occurs at isotopic steady state, i.e. that the transpired water shows the same isotopic signature as the source water. Yet, several studies found that transpiration did not occur at isotopic steady state, under both controlled and field conditions. Here we focused on identifying the internal and external factors which drive the isotopic signature of leaf transpiration. Using cavity ring-down spectroscopy (CRDS), the effect of both environmental variables and leaf physiological traits on δ18OT was investigated under controlled conditions. Six plant species with distinct leaf physiological traits were exposed to step changes in relative air humidity (RH), their response in δ18OT and gas exchange parameters and their leaf physiological traits were assessed. Moreover, two functionally distinct plant types (tree, i.e. Quercus suber, and grassland) of a semi-arid Mediterranean oak-woodland where observed under natural conditions throughout an entire growth period in the field. The species differed substantially in their leaf physiological traits and their turn-over times of leaf water. They could be grouped in species with fast (<60 min.), intermediate (ca. 120 min.) and slow (>240 min.) turn-over times, mostly due to differences in stomatal conductance, leaf water content or a combination of both. Changes in RH caused an immediate response in δ18OT, which were similarly strong in all species, while leaf physiological traits affected the subsequent response in δ18OT. The turn-over time of leaf water determined the speed of return to the isotopic steady or a stable δ18OT value (Dubbert & Kübert et al., in prep.). Under natural conditions, changes in environmental conditions over the diurnal cycle had a huge impact on the diurnal development of δ18OT in both

  1. Extremely Low Microsatellite Diversity but Distinct Population Structure in a Long-Lived Threatened Species, the Australian Lungfish Neoceratodus forsteri (Dipnoi)

    PubMed Central

    Hughes, Jane M.; Schmidt, Daniel J.; Huey, Joel A.; Real, Kathryn M.; Espinoza, Thomas; McDougall, Andrew; Kind, Peter K.; Brooks, Steven; Roberts, David T.

    2015-01-01

    The Australian lungfish is a unique living representative of an ancient dipnoan lineage, listed as ‘vulnerable’ to extinction under Australia’s Environment Protection and Biodiversity Conservation Act 1999. Historical accounts indicate this species occurred naturally in two adjacent river systems in Australia, the Burnett and Mary. Current day populations in other rivers are thought to have arisen by translocation from these source populations. Early genetic work detected very little variation and so had limited power to answer questions relevant for management including how genetic variation is partitioned within and among sub-populations. In this study, we use newly developed microsatellite markers to examine samples from the Burnett and Mary Rivers, as well as from two populations thought to be of translocated origin, Brisbane and North Pine. We test whether there is significant genetic structure among and within river drainages; assign putatively translocated populations to potential source populations; and estimate effective population sizes. Eleven polymorphic microsatellite loci genotyped in 218 individuals gave an average within-population heterozygosity of 0.39 which is low relative to other threatened taxa and for freshwater fishes in general. Based on FST values (average over loci = 0.11) and STRUCTURE analyses, we identify three distinct populations in the natural range, one in the Burnett and two distinct populations in the Mary. These analyses also support the hypothesis that the Mary River is the likely source of translocated populations in the Brisbane and North Pine rivers, which agrees with historical published records of a translocation event giving rise to these populations. We were unable to obtain bounded estimates of effective population size, as we have too few genotype combinations, although point estimates were low, ranging from 29 - 129. We recommend that, in order to preserve any local adaptation in the three distinct populations

  2. Extremely low microsatellite diversity but distinct population structure in a long-lived threatened species, the Australian lungfish Neoceratodus forsteri (Dipnoi).

    PubMed

    Hughes, Jane M; Schmidt, Daniel J; Huey, Joel A; Real, Kathryn M; Espinoza, Thomas; McDougall, Andrew; Kind, Peter K; Brooks, Steven; Roberts, David T

    2015-01-01

    The Australian lungfish is a unique living representative of an ancient dipnoan lineage, listed as 'vulnerable' to extinction under Australia's Environment Protection and Biodiversity Conservation Act 1999. Historical accounts indicate this species occurred naturally in two adjacent river systems in Australia, the Burnett and Mary. Current day populations in other rivers are thought to have arisen by translocation from these source populations. Early genetic work detected very little variation and so had limited power to answer questions relevant for management including how genetic variation is partitioned within and among sub-populations. In this study, we use newly developed microsatellite markers to examine samples from the Burnett and Mary Rivers, as well as from two populations thought to be of translocated origin, Brisbane and North Pine. We test whether there is significant genetic structure among and within river drainages; assign putatively translocated populations to potential source populations; and estimate effective population sizes. Eleven polymorphic microsatellite loci genotyped in 218 individuals gave an average within-population heterozygosity of 0.39 which is low relative to other threatened taxa and for freshwater fishes in general. Based on FST values (average over loci = 0.11) and STRUCTURE analyses, we identify three distinct populations in the natural range, one in the Burnett and two distinct populations in the Mary. These analyses also support the hypothesis that the Mary River is the likely source of translocated populations in the Brisbane and North Pine rivers, which agrees with historical published records of a translocation event giving rise to these populations. We were unable to obtain bounded estimates of effective population size, as we have too few genotype combinations, although point estimates were low, ranging from 29 - 129. We recommend that, in order to preserve any local adaptation in the three distinct populations that

  3. A new and distinct species in the genus Caulimovirus exists as an endogenous plant pararetroviral sequence in its host, Dahlia variabilis.

    PubMed

    Pahalawatta, Vihanga; Druffel, Keri; Pappu, Hanu

    2008-07-01

    Viruses in certain genera in family Caulimoviridae were shown to integrate their genomic sequences into their host genomes and exist as endogenous pararetroviral sequences (EPRV). However, members of the genus Caulimovirus remained to be the exception and are known to exist only as episomal elements in the infected cell. We present evidence that the DNA genome of a new and distinct Caulimovirus species, associated with dahlia mosaic, is integrated into its host genome, dahlia (Dahlia variabilis). Using cloned viral genes as probes, Southern blot hybridization of total plant DNA from dahlia seedlings showed the presence of viral DNA in the host DNA. Fluorescent in situ hybridization using labeled DNA probes from the D10 genome localized the viral sequences in dahlia chromosomes. The natural integration of a Caulimovirus genome into its host and its existence as an EPRV suggests the co-evolution of this plant-virus pathosystem. PMID:18462770

  4. RNA sequencing supports distinct reactive oxygen species-mediated pathways of apoptosis by high and low size mass fractions of Bay leaf (Lauris nobilis) in HT-29 cells.

    PubMed

    Rodd, Annabelle L; Ververis, Katherine; Sayakkarage, Dheeshana; Khan, Abdul W; Rafehi, Haloom; Ziemann, Mark; Loveridge, Shanon J; Lazarus, Ross; Kerr, Caroline; Lockett, Trevor; El-Osta, Assam; Karagiannis, Tom C; Bennett, Louise E

    2015-08-01

    Anti-proliferative and pro-apoptotic effects of Bay leaf (Laurus nobilis) in mammalian cancer and HT-29 adenocarcinoma cells have been previously attributed to effects of polyphenolic and essential oil chemical species. Recently, we demonstrated differentiated growth-regulating effects of high (HFBL) versus low molecular mass (LFBL) aqueous fractions of bay leaf and now confirm by comparative effects on gene expression, that HFBL and LFBL suppress HT-29 growth by distinct mechanisms. Induction of intra-cellular lesions including DNA strand breakage by extra-cellular HFBL, invoked the hypothesis that iron-mediated reactive oxygen species with capacity to penetrate cell membrane, were responsible for HFBL-mediated effects, supported by equivalent effects of HFBL in combination with γ radiation. Activities of HFBL and LFBL were interpreted to reflect differentiated responses to iron-mediated reactive oxygen species (ROS), occurring either outside or inside cells. In the presence of LFBL, apoptotic death was relatively delayed compared with HFBL. ROS production by LFBL mediated p53-dependent apoptosis and recovery was suppressed by promoting G1/S phase arrest and failure of cellular tight junctions. In comparison, intra-cellular anti-oxidant protection exerted by LFBL was absent for extra-cellular HFBL (likely polysaccharide-rich), which potentiated more rapid apoptosis by producing DNA double strand breaks. Differentiated effects on expression of genes regulating ROS defense and chromatic condensation by LFBL versus HFBL, were observed. The results support ferrous iron in cell culture systems and potentially in vivo, can invoke different extra-cellular versus intra-cellular ROS-mediated chemistries, that may be regulated by exogenous, including dietary species. PMID:26114728

  5. RNA sequencing supports distinct reactive oxygen species-mediated pathways of apoptosis by high and low size mass fractions of Bay leaf (Lauris nobilis) in HT-29 cells.

    PubMed

    Rodd, Annabelle L; Ververis, Katherine; Sayakkarage, Dheeshana; Khan, Abdul W; Rafehi, Haloom; Ziemann, Mark; Loveridge, Shanon J; Lazarus, Ross; Kerr, Caroline; Lockett, Trevor; El-Osta, Assam; Karagiannis, Tom C; Bennett, Louise E

    2015-08-01

    Anti-proliferative and pro-apoptotic effects of Bay leaf (Laurus nobilis) in mammalian cancer and HT-29 adenocarcinoma cells have been previously attributed to effects of polyphenolic and essential oil chemical species. Recently, we demonstrated differentiated growth-regulating effects of high (HFBL) versus low molecular mass (LFBL) aqueous fractions of bay leaf and now confirm by comparative effects on gene expression, that HFBL and LFBL suppress HT-29 growth by distinct mechanisms. Induction of intra-cellular lesions including DNA strand breakage by extra-cellular HFBL, invoked the hypothesis that iron-mediated reactive oxygen species with capacity to penetrate cell membrane, were responsible for HFBL-mediated effects, supported by equivalent effects of HFBL in combination with γ radiation. Activities of HFBL and LFBL were interpreted to reflect differentiated responses to iron-mediated reactive oxygen species (ROS), occurring either outside or inside cells. In the presence of LFBL, apoptotic death was relatively delayed compared with HFBL. ROS production by LFBL mediated p53-dependent apoptosis and recovery was suppressed by promoting G1/S phase arrest and failure of cellular tight junctions. In comparison, intra-cellular anti-oxidant protection exerted by LFBL was absent for extra-cellular HFBL (likely polysaccharide-rich), which potentiated more rapid apoptosis by producing DNA double strand breaks. Differentiated effects on expression of genes regulating ROS defense and chromatic condensation by LFBL versus HFBL, were observed. The results support ferrous iron in cell culture systems and potentially in vivo, can invoke different extra-cellular versus intra-cellular ROS-mediated chemistries, that may be regulated by exogenous, including dietary species.

  6. A comparative study on Ca content and distribution in two Gesneriaceae species reveals distinctive mechanisms to cope with high rhizospheric soluble calcium.

    PubMed

    Li, Wenlong; Xu, Falun; Chen, Shixuan; Zhang, Zhennan; Zhao, Yan; Jin, Yukuan; Li, Meijing; Zhu, Yan; Liu, Yongxiu; Yang, Yi; Deng, Xin

    2014-01-01

    Excessive Ca is toxic to plants thus significantly affects plant growth and species distribution in Ca-rich karst areas. To understand how plants survive high Ca soil, laboratory experiments were established to compare the physiological responses and internal Ca distribution in organ, tissue, cell, and intracellular levels under different Ca levels for Lysionotus pauciflorus and Boea hygrometrica, two karst habitant Gesneriaceae species in Southwest China. In the controlled condition, L. pauciflorus could survive as high as 200 mM rhizospheric soluble Ca, attributed to a series of physiological responses and preferential storage that limited Ca accumulation in chloroplasts of palisade cells. In contrast, B. hygrometrica could survive only 20 mM rhizospheric soluble Ca, but accumulated a high level of internal Ca in both palisade and spongy cells without disturbance on photosynthetic activity. By phenotype screening of transgenic plants expressing high Ca-inducible genes from B. hygrometrica, the expression of BhDNAJC2 in A. thaliana was found to enhance plant growth and photosynthesis under high soluble Ca stress. BhDNAJC2 encodes a recently reported heat shock protein (HSP) 40 family DnaJ-domain protein. The Ca-resistant phenotype of BhDNAJC2 highlights the important role of chaperone-mediated protein quality control in Ca tolerance in B. hygrometrica. Taken together, our results revealed that distinctive mechanisms were employed in the two Gesneriaceae karst habitants to cope with a high Ca environment.

  7. A comparative study on Ca content and distribution in two Gesneriaceae species reveals distinctive mechanisms to cope with high rhizospheric soluble calcium

    PubMed Central

    Li, Wenlong; Xu, Falun; Chen, Shixuan; Zhang, Zhennan; Zhao, Yan; Jin, Yukuan; Li, Meijing; Zhu, Yan; Liu, Yongxiu; Yang, Yi; Deng, Xin

    2014-01-01

    Excessive Ca is toxic to plants thus significantly affects plant growth and species distribution in Ca-rich karst areas. To understand how plants survive high Ca soil, laboratory experiments were established to compare the physiological responses and internal Ca distribution in organ, tissue, cell, and intracellular levels under different Ca levels for Lysionotus pauciflorus and Boea hygrometrica, two karst habitant Gesneriaceae species in Southwest China. In the controlled condition, L. pauciflorus could survive as high as 200 mM rhizospheric soluble Ca, attributed to a series of physiological responses and preferential storage that limited Ca accumulation in chloroplasts of palisade cells. In contrast, B. hygrometrica could survive only 20 mM rhizospheric soluble Ca, but accumulated a high level of internal Ca in both palisade and spongy cells without disturbance on photosynthetic activity. By phenotype screening of transgenic plants expressing high Ca-inducible genes from B. hygrometrica, the expression of BhDNAJC2 in A. thaliana was found to enhance plant growth and photosynthesis under high soluble Ca stress. BhDNAJC2 encodes a recently reported heat shock protein (HSP) 40 family DnaJ-domain protein. The Ca-resistant phenotype of BhDNAJC2 highlights the important role of chaperone-mediated protein quality control in Ca tolerance in B. hygrometrica. Taken together, our results revealed that distinctive mechanisms were employed in the two Gesneriaceae karst habitants to cope with a high Ca environment. PMID:25477893

  8. Comprehensive genetic analyses reveal evolutionary distinction of a mouse (Zapus hudsonius preblei) proposed for delisting from the US Endangered Species Act

    USGS Publications Warehouse

    King, T.L.; Switzer, J.F.; Morrison, C.L.; Eackles, M.S.; Young, C.C.; Lubinski, B.A.; Cryan, P.

    2006-01-01

    Zapus hudsonius preblei, listed as threatened under the US Endangered Species Act (ESA), is one of 12 recognized subspecies of meadow jumping mice found in North America. Recent morphometric and phylogenetic comparisons among Z. h. preblei and neighbouring conspecifics questioned the taxonomic status of selected subspecies, resulting in a proposal to delist the Z. h. preblei from the ESA. We present additional analyses of the phylogeographic structure within Z. hudsonius that calls into question previously published data (and conclusions) and confirms the original taxonomic designations. A survey of 21 microsatellite DNA loci and 1380 base pairs from two mitochondrial DNA (mtDNA) regions (control region and cytochrome b) revealed that each Z. hudsonius subspecies is genetically distinct. These data do not support the null hypothesis of a homogeneous gene pool among the five subspecies found within the southwestern portion of the species' range. The magnitude of the observed differentiation was considerable and supported by significant findings for nearly every statistical comparison made, regardless of the genome or the taxa under consideration. Structuring of nuclear multilocus genotypes and subspecies-specific mtDNA haplotypes corresponded directly with the disjunct distributions of the subspecies investigated. Given the level of correspondence between the observed genetic population structure and previously proposed taxonomic classification of subspecies (based on the geographic separation and surveys of morphological variation), we conclude that the nominal subspecies surveyed in this study do not warrant synonymy, as has been proposed for Z. h. preblei, Z. h. campestris, and Z. h. intermedius. ?? 2006 The Authors.

  9. IFN-β-inducing, unusual viral RNA species produced by paramyxovirus infection accumulated into distinct cytoplasmic structures in an RNA-type-dependent manner

    PubMed Central

    Yoshida, Asuka; Kawabata, Ryoko; Honda, Tomoyuki; Tomonaga, Keizo; Sakaguchi, Takemasa; Irie, Takashi

    2015-01-01

    The interferon (IFN) system is one of the most important defensive responses of mammals against viruses, and is rapidly evoked when the pathogen-associated molecular patterns (PAMPs) of viruses are sensed. Non-self, virus-derived RNA species have been identified as the PAMPs of RNA viruses. In the present study, we compared different types of IFN-β-inducing and -non-inducing viruses in the context of Sendai virus infection. We found that some types of unusual viral RNA species were produced by infections with IFN-β-inducing viruses and accumulated into distinct cytoplasmic structures in an RNA-type-dependent manner. One of these structures was similar to the so-called antiviral stress granules (avSGs) formed by an infection with IFN-inducing viruses whose C proteins were knocked-out or mutated. Non-encapsidated, unusual viral RNA harboring the 5′-terminal region of the viral genome as well as RIG-I and typical SG markers accumulated in these granules. Another was a non-SG-like inclusion formed by an infection with the Cantell strain; a copyback-type DI genome, but not an authentic viral genome, specifically accumulated in the inclusion, whereas RIG-I and SG markers did not. The induction of IFN-β was closely associated with the production of these unusual RNAs as well as the formation of the cytoplasmic structures. PMID:26300870

  10. Insights into assessing water quality using taxonomic distinctness based on a small species pool of biofilm-dwelling ciliate fauna in coastal waters of the Yellow Sea, northern China.

    PubMed

    Zhang, Wei; Liu, Yuanyuan; Warren, Alan; Xu, Henglong

    2014-12-15

    The aim of this study is to determine the feasibility of using a small species pool from a raw dataset of biofilm-dwelling ciliates for bioassessment based on taxonomic diversity. Samples were collected monthly at four stations within a gradient of environmental stress in coastal waters of the Yellow Sea, northern China from August 2011 to July 2012. A 33-species subset was identified from the raw 137-species dataset using a multivariate method. The spatial patterns of this subset were significantly correlated with the changes in the nutrients and chemical oxygen demand. The taxonomic diversity indices were significantly correlated with nutrients. The pair-wise indices of average taxonomic distinctness (Δ(+)) and the taxonomic distinctness (Λ(+)) showed a clear departure from the expected taxonomic pattern. These findings suggest that this small ciliate assemblage might be used as an adequate species pool for discriminating water quality status based on taxonomic distinctness in marine ecosystems.

  11. Folding, stability, and physical properties of the alpha subunit of bacterial luciferase.

    PubMed

    Noland, B W; Dangott, L J; Baldwin, T O

    1999-12-01

    Bacterial luciferase is a heterodimeric (alphabeta) enzyme composed of homologous subunits. When the Vibrio harveyi luxA gene is expressed in Escherichia coli, the alpha subunit accumulates to high levels. The alpha subunit has a well-defined near-UV circular dichroism spectrum and a higher intrinsic fluorescence than the heterodimer, demonstrating fluorescence quenching in the enzyme which is reduced in the free subunit [Sinclair, J. F., Waddle, J. J., Waddill, W. F., and Baldwin, T. O. (1993) Biochemistry 32, 5036-5044]. Analytical ultracentrifugation of the alpha subunit has revealed a reversible monomer to dimer equilibrium with a dissociation constant of 14.9 +/- 4.0 microM at 18 degrees C in 50 mM phosphate and 100 mM NaCl, pH 7.0. The alpha subunit unfolded and refolded reversibly in urea-containing buffers by a three-state mechanism. The first transition occurred over the range of 0-2 M urea with an associated free-energy change of 2.24 +/- 0.25 kcal/mol at 18 degrees C in 50 mM phosphate buffer, pH 7.0. The second, occurring between 2.5 and 3.5 M urea, comprised a cooperative transition with a free-energy change of 6.50 +/- 0.75 kcal/mol. The intermediate species, populated maximally at ca. 2 M urea, has defined near-UV circular dichroism spectral properties distinct from either the native or the denatured states. The intrinsic fluorescence of the intermediate suggested that, although the quantum yield had decreased, the tryptophanyl residues remained largely buried. The far-UV circular dichroism spectrum of the intermediate indicated that it had lost ca. 40% of its native secondary structure. N-Terminal sequencing of the products of limited proteolysis of the intermediate showed that the C-terminal region of the alpha subunit became protease labile over the urea concentration range at which the intermediate was maximally populated. These observations have led us to propose an unfolding model in which the first transition is the unfolding of a C

  12. Characterization and charge distribution of the asparagine-linked oligosaccharides on secreted mouse thyrotropin and free alpha-subunits

    SciTech Connect

    Gesundheit, N.; Gyves, P.W.; DeCherney, G.S.; Stannard, B.S.; Winston, R.L.; Weintraub, B.D.

    1989-06-01

    Mouse hemipituitaries in vitro secrete TSH, composed of an alpha-beta heterodimer, as well as excess (free) alpha-subunits. By dual metabolic labeling with (35S)sulfate and (3H)mannose, we have characterized oligosaccharides from secreted TSH alpha, TSH beta, and free alpha-subunits released from the apoprotein by enzymatic deglycosylation. Oligosaccharides from each subunit displayed a distinct anion exchange HPLC profile due to a specific pattern of sialylation and sulfation. Six species were obtained from TSH alpha (with two glycosylation sites), including neutral oligosaccharides as well as those with one or two negative charges. For TSH beta (with one glycosylation site) at least eight oligosaccharide species were noted, representing nearly every permutation of sialylation and sulfation; approximately 30% contained three or more negative charges. Analysis of (3H)mannose-labeled oligosaccharides on Concanavalin-A-agarose showed 85% binding for those from TSH alpha, 70% for free alpha, and 50% for those from TSH beta. These data demonstrate that oligosaccharides from secreted TSH beta were more sialylated and sulfated, consistent with a more complex branching pattern, than those from TSH alpha. Oligosaccharides from free alpha-subunit were more sialylated than those from TSH alpha, and the net negative charge was intermediate between those of TSH alpha and TSH beta. Although great microheterogeneity is present even at the single glycosylation site on the beta-subunit of secreted TSH, a pattern of sialylation and sulfation could be discerned.

  13. Distinct Macrophage Fates after in vitro Infection with Different Species of Leishmania: Induction of Apoptosis by Leishmania (Leishmania) amazonensis, but Not by Leishmania (Viannia) guyanensis

    PubMed Central

    DaMata, Jarina Pena; Mendes, Bárbara Pinheiro; Maciel-Lima, Kátia; Menezes, Cristiane Alves Silva; Dutra, Walderez Ornelas; Sousa, Lirlândia Pires; Horta, Maria Fátima

    2015-01-01

    Leishmania is an intracellular parasite in vertebrate hosts, including man. During infection, amastigotes replicate inside macrophages and are transmitted to healthy cells, leading to amplification of the infection. Although transfer of amastigotes from infected to healthy cells is a crucial step that may shape the outcome of the infection, it is not fully understood. Here we compare L. amazonensis and L. guyanensis infection in C57BL/6 and BALB/c mice and investigate the fate of macrophages when infected with these species of Leishmania in vitro. As previously shown, infection of mice results in distinct outcomes: L. amazonensis causes a chronic infection in both strains of mice (although milder in C57BL/6), whereas L. guyanensis does not cause them disease. In vitro, infection is persistent in L. amazonensis-infected macrophages whereas L. guyanensis growth is controlled by host cells from both strains of mice. We demonstrate that, in vitro, L. amazonensis induces apoptosis of both C57BL/6 and BALB/c macrophages, characterized by PS exposure, DNA cleavage into nucleosomal size fragments, and consequent hypodiploidy. None of these signs were seen in macrophages infected with L. guyanensis, which seem to die through necrosis, as indicated by increased PI-, but not Annexin V-, positive cells. L. amazonensis-induced macrophage apoptosis was associated to activation of caspases-3, -8 and -9 in both strains of mice. Considering these two species of Leishmania and strains of mice, macrophage apoptosis, induced at the initial moments of infection, correlates with chronic infection, regardless of its severity. We present evidence suggestive that macrophages phagocytize L. amazonensis-infected cells, which has not been verified so far. The ingestion of apoptotic infected macrophages by healthy macrophages could be a way of amastigote spreading, leading to the establishment of infection. PMID:26513474

  14. Distinct Macrophage Fates after in vitro Infection with Different Species of Leishmania: Induction of Apoptosis by Leishmania (Leishmania) amazonensis, but Not by Leishmania (Viannia) guyanensis.

    PubMed

    DaMata, Jarina Pena; Mendes, Bárbara Pinheiro; Maciel-Lima, Kátia; Menezes, Cristiane Alves Silva; Dutra, Walderez Ornelas; Sousa, Lirlândia Pires; Horta, Maria Fátima

    2015-01-01

    Leishmania is an intracellular parasite in vertebrate hosts, including man. During infection, amastigotes replicate inside macrophages and are transmitted to healthy cells, leading to amplification of the infection. Although transfer of amastigotes from infected to healthy cells is a crucial step that may shape the outcome of the infection, it is not fully understood. Here we compare L. amazonensis and L. guyanensis infection in C57BL/6 and BALB/c mice and investigate the fate of macrophages when infected with these species of Leishmania in vitro. As previously shown, infection of mice results in distinct outcomes: L. amazonensis causes a chronic infection in both strains of mice (although milder in C57BL/6), whereas L. guyanensis does not cause them disease. In vitro, infection is persistent in L. amazonensis-infected macrophages whereas L. guyanensis growth is controlled by host cells from both strains of mice. We demonstrate that, in vitro, L. amazonensis induces apoptosis of both C57BL/6 and BALB/c macrophages, characterized by PS exposure, DNA cleavage into nucleosomal size fragments, and consequent hypodiploidy. None of these signs were seen in macrophages infected with L. guyanensis, which seem to die through necrosis, as indicated by increased PI-, but not Annexin V-, positive cells. L. amazonensis-induced macrophage apoptosis was associated to activation of caspases-3, -8 and -9 in both strains of mice. Considering these two species of Leishmania and strains of mice, macrophage apoptosis, induced at the initial moments of infection, correlates with chronic infection, regardless of its severity. We present evidence suggestive that macrophages phagocytize L. amazonensis-infected cells, which has not been verified so far. The ingestion of apoptotic infected macrophages by healthy macrophages could be a way of amastigote spreading, leading to the establishment of infection. PMID:26513474

  15. Genes encoding the alpha-carboxyltransferase subunit of acetyl-CoA carboxylase from Brassica napus and parental species: cloning, expression patterns, and evolution.

    PubMed

    Li, Zhi-Guo; Yin, Wei-Bo; Guo, Huan; Song, Li-Ying; Chen, Yu-Hong; Guan, Rong-Zhan; Wang, Jing-Qiao; Wang, Richard R-C; Hu, Zan-Min

    2010-05-01

    Heteromeric acetyl coenzyme A carboxylase (ACCase), a rate-limiting enzyme in fatty acid biosynthesis in dicots, is a multi-enzyme complex consisting of biotin carboxylase, biotin carboxyl carrier protein, and carboxyltransferase (alpha-CT and beta-CT). In the present study, four genes encoding alpha-CT were cloned from Brassica napus, and two were cloned from each of the two parental species, B. rapa and B. oleracea. Comparative and cluster analyses indicated that these genes were divided into two major groups. The major divergence between group-1 and group-2 occurred in the second intron. Group-2 alpha-CT genes represented the ancestral form in the genus Brassica. The divergence of group-1 and group-2 genes occurred in their common ancestor 12.96-17.78 million years ago (MYA), soon after the divergence of Arabidopsis thaliana and Brassica (15-20 MYA). This time of divergence is identical to that reported for the paralogous subgenomes of diploid Brassica species (13-17 MYA). Real-time reverse transcription PCR revealed that the expression patterns of the two groups of genes were similar in different organs, except in leaves. To better understand the regulation and evolution of alpha-CT genes, promoter regions from two sets of orthologous gene copies from B. napus, B. rapa, and B. oleracea were cloned and compared. The function of the promoter of gene Bnalpha-CT-1-1 in group-1 and gene Bnalpha-CT-2-1 in group-2 was examined by assaying beta-glucuronidase activity in transgenic A. thaliana. Our results will be helpful in elucidating the evolution and regulation of ACCase in oilseed rape.

  16. Binding interface between the Salmonella σS/RpoS subunit of RNA polymerase and Crl: hints from bacterial species lacking crl

    PubMed Central

    Cavaliere, Paola; Sizun, Christina; Levi-Acobas, Fabienne; Nowakowski, Mireille; Monteil, Véronique; Bontems, François; Bellalou, Jacques; Mayer, Claudine; Norel, Françoise

    2015-01-01

    In many Gram-negative bacteria, including Salmonella enterica serovar Typhimurium (S. Typhimurium), the sigma factor RpoS/σS accumulates during stationary phase of growth, and associates with the core RNA polymerase enzyme (E) to promote transcription initiation of genes involved in general stress resistance and starvation survival. Whereas σ factors are usually inactivated upon interaction with anti-σ proteins, σS binding to the Crl protein increases σS activity by favouring its association to E. Taking advantage of evolution of the σS sequence in bacterial species that do not contain a crl gene, like Pseudomonas aeruginosa, we identified and assigned a critical arginine residue in σS to the S. Typhimurium σS-Crl binding interface. We solved the solution structure of S. Typhimurium Crl by NMR and used it for NMR binding assays with σS and to generate in silico models of the σS-Crl complex constrained by mutational analysis. The σS-Crl models suggest that the identified arginine in σS interacts with an aspartate of Crl that is required for σS binding and is located inside a cavity enclosed by flexible loops, which also contribute to the interface. This study provides the basis for further structural investigation of the σS-Crl complex. PMID:26338235

  17. Polymorphism of genes coding for nuclear 18S rRNA indicates genetic distinctiveness of anastomosis group 10 from other groups in the Rhizoctonia solani species complex.

    PubMed

    Liu, Z L; Domier, L L; Sinclair, J B

    1995-07-01

    DNA polymorphism in the 18S nuclear rRNA gene region was investigated by using 11 restriction endonucleases for 161 isolates of 25 intraspecific groups (ISGs) representing 11 reported anastomosis groups (AGs) of Rhizoctonia solani. A PCR-based restriction mapping method in which enzymatically amplified DNA fragments and subfragments were digested with one or two restriction enzymes was employed. Four types of DNA restriction maps of this region were constructed for these 25 ISGs. Map type I of the 18S rDNA region was represented by isolates of a majority of R. solani ISGs. Map types II and III, represented by ISG 2E and 9 isolates and 5C isolates, respectively, differed from map I by the absence of one (map type II) or two (map type III) restriction sites. Map type IV, represented by ISG 10A and B (or AG 10) isolates, showed significant restriction site variations, with five enzymes in this region compared with those of the remaining ISGs or AGs. Ten of the 25 restriction sites in the 18S rRNA gene region were informative and selected for analysis. Previously reported restriction maps of the 5.8S rRNA gene region, including the internal transcribed spacers, were aligned with each other, and 12 informative restriction sites were identified. These data were used alone and in combination to evaluate group relationships. Analyses derived from these data sets by maximum parsimony and likelihood methods showed that AG 10 isolates were distinct and distantly related to the majority isolates of the other AGs of this species complex.

  18. Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia.

    PubMed

    Yusof, Ruhani; Ahmed, Md Atique; Jelip, Jenarun; Ngian, Hie Ung; Mustakim, Sahlawati; Hussin, Hani Mat; Fong, Mun Yik; Mahmud, Rohela; Sitam, Frankie Anak Thomas; Japning, J Rovie-Ryan; Snounou, Georges; Escalante, Ananias A; Lau, Yee Ling

    2016-08-01

    Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, although most cases have occurred in Malaysia, where P. knowlesi is now the dominant malaria species. This apparently skewed distribution prompted an investigation of the phylogeography of this parasite in 2 geographically separated regions of Malaysia, Peninsular Malaysia and Malaysian Borneo. We investigated samples collected from humans and macaques in these regions. Haplotype network analyses of sequences from 2 P. knowlesi genes, type A small subunit ribosomal 18S RNA and cytochrome c oxidase subunit I, showed 2 genetically distinct divergent clusters, 1 from each of the 2 regions of Malaysia. We propose that these parasites represent 2 distinct P. knowlesi types that independently became zoonotic. These types would have evolved after the sea-level rise at the end of the last ice age, which separated Malaysian Borneo from Peninsular Malaysia.

  19. Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia

    PubMed Central

    Yusof, Ruhani; Ahmed, Md Atique; Jelip, Jenarun; Ngian, Hie Ung; Mustakim, Sahlawati; Hussin, Hani Mat; Fong, Mun Yik; Mahmud, Rohela; Sitam, Frankie Anak Thomas; Japning, J. Rovie-Ryan; Snounou, Georges; Escalante, Ananias A.

    2016-01-01

    Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, although most cases have occurred in Malaysia, where P. knowlesi is now the dominant malaria species. This apparently skewed distribution prompted an investigation of the phylogeography of this parasite in 2 geographically separated regions of Malaysia, Peninsular Malaysia and Malaysian Borneo. We investigated samples collected from humans and macaques in these regions. Haplotype network analyses of sequences from 2 P. knowlesi genes, type A small subunit ribosomal 18S RNA and cytochrome c oxidase subunit I, showed 2 genetically distinct divergent clusters, 1 from each of the 2 regions of Malaysia. We propose that these parasites represent 2 distinct P. knowlesi types that independently became zoonotic. These types would have evolved after the sea-level rise at the end of the last ice age, which separated Malaysian Borneo from Peninsular Malaysia. PMID:27433965

  20. Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia.

    PubMed

    Yusof, Ruhani; Ahmed, Md Atique; Jelip, Jenarun; Ngian, Hie Ung; Mustakim, Sahlawati; Hussin, Hani Mat; Fong, Mun Yik; Mahmud, Rohela; Sitam, Frankie Anak Thomas; Japning, J Rovie-Ryan; Snounou, Georges; Escalante, Ananias A; Lau, Yee Ling

    2016-08-01

    Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, although most cases have occurred in Malaysia, where P. knowlesi is now the dominant malaria species. This apparently skewed distribution prompted an investigation of the phylogeography of this parasite in 2 geographically separated regions of Malaysia, Peninsular Malaysia and Malaysian Borneo. We investigated samples collected from humans and macaques in these regions. Haplotype network analyses of sequences from 2 P. knowlesi genes, type A small subunit ribosomal 18S RNA and cytochrome c oxidase subunit I, showed 2 genetically distinct divergent clusters, 1 from each of the 2 regions of Malaysia. We propose that these parasites represent 2 distinct P. knowlesi types that independently became zoonotic. These types would have evolved after the sea-level rise at the end of the last ice age, which separated Malaysian Borneo from Peninsular Malaysia. PMID:27433965

  1. Subunit organization in cytoplasmic dynein subcomplexes

    PubMed Central

    King, Stephen J.; Bonilla, Myriam; Rodgers, Michael E.; Schroer, Trina A.

    2002-01-01

    Because cytoplasmic dynein plays numerous critical roles in eukaryotic cells, determining the subunit composition and the organization and functions of the subunits within dynein are important goals. This has been difficult partly because of accessory polypeptide heterogeneity of dynein populations. The motor domain containing heavy chains of cytoplasmic dynein are associated with multiple intermediate, light intermediate, and light chain accessory polypeptides. We examined the organization of these subunits within cytoplasmic dynein by separating the molecule into two distinct subcomplexes. These subcomplexes were competent to reassemble into a molecule with dynein-like properties. One subcomplex was composed of the dynein heavy and light intermediate chains whereas the other subcomplex was composed of the intermediate and light chains. The intermediate and light chain subcomplex could be further separated into two pools, only one of which contained dynein light chains. The two pools had distinct intermediate chain compositions, suggesting that intermediate chain isoforms have different light chain–binding properties. When the two intermediate chain pools were characterized by analytical velocity sedimentation, at least four molecular components were seen: intermediate chain monomers, intermediate chain dimers, intermediate chain monomers with bound light chains, and a mixture of intermediate chain dimers with assorted bound light chains. These data provide new insights into the compositional heterogeneity and assembly of the cytoplasmic dynein complex and suggest that individual dynein molecules have distinct molecular compositions in vivo. PMID:11967380

  2. Carbon and Oxygen Isotope Measurements of Ordinary Chondrite (OC) Meteorites from Antarctica Indicate Distinct Terrestrial Carbonate Species using a Stepped Acid Extraction Procedure Impacting Mars Carbonate Research

    NASA Astrophysics Data System (ADS)

    Evans, M. E.; Niles, P. B.; Locke, D.

    2015-12-01

    The purpose of this study is to characterize the stable isotope values of terrestrial, secondary carbonate minerals from five OC meteorites collected in Antarctica. These samples were selected for analysis based upon their size and collection proximity to known Martian meteorites. They were also selected based on petrologic type (3+) such that they were likely to be carbonate-free before falling to Earth. This study has two main tasks: 1) characterize the isotopic composition of terrestrial, secondary carbonate minerals formed on meteorites in Antarctica, and 2) study the mechanisms of carbonate formation in cold and arid environments with Antarctica as an analog for Mars. Two samples from each meteorite, each ~0.5g, was crushed and dissolved in pure phosphoric acid for 3 sequential reactions: a) Rx0 for 1 hour at 30°C, b) Rx1 for 18 hours at 30°C, and c) Rx2 for 3 hours at 150°C. CO2 was distilled by freezing with liquid nitrogen from each sample tube, then separated from organics and sulfides with a TRACE GC using a Restek HayeSep Q 80/100 6' 2mm stainless column, and then analyzed on a Thermo MAT 253 IRMS in Dual Inlet mode. This system was built at NASA/JSC over the past 3 years and proof tested with known carbonate standards to develop procedures, assess yield, and quantify expected uncertainties. Two distinct species of carbonates are found based on the stepped extraction technique: 1) Ca-rich carbonate released at low temperatures, and 2) Mg, or Fe-rich carbonate released at high temperatures. Preliminary results indicate that most of the carbonates present in the ordinary chondrites analyzed have δ13C=+5‰, which is consistent with formation from atmospheric CO2 δ13C=-7‰ at -20°C. The oxygen isotopic compositions of the carbonates vary between +4‰ and +34‰ with the Mg-rich and/or Fe-rich carbonates possessing the lowest δ18O values. This suggests that the carbonates formed under a wide range of temperatures. However, the carbonate oxygen

  3. Carbon and Oxygen Isotope Measurements of Ordinary Chondrite (OC) Meteorites from Antarctica Indicate Distinct Carbonate Species Using a Stepped Acid Extraction Procedure

    NASA Technical Reports Server (NTRS)

    Evans, Michael E.

    2015-01-01

    The purpose of this study is to characterize the stable isotope values of terrestrial, secondary carbonate minerals from five Ordinary Chondrite (OC) meteorites collected in Antarctica. These samples were identified and requested from NASA based upon their size, alteration history, and collection proximity to known Martian meteorites. They are also assumed to be carbonate-free before falling to Earth. This research addresses two questions involving Mars carbonates: 1) characterize terrestrial, secondary carbonate isotope values to apply to Martian meteorites for isolating in-situ carbonates, and 2) increase understanding of carbonates formed in cold and arid environments with Antarctica as an analog for Mars. Two samples from each meteorite, each approximately 0.5 grams, were crushed and dissolved in pure phosphoric acid for 3 sequential reactions: a) R times 0 for 1 hour at 30 degrees Centigrade (fine calcite extraction), b) R times 1 for 18 hours at 30 degrees Centigrade (course calcite extraction), and c) R times 2 for 3 hours at 150 degrees Centigrade (siderite and/or magnesite extraction). CO (sub 2) was distilled by freezing with liquid nitrogen from each sample tube, then separated from organics and sulfides with a TRACE GC using a Restek HayeSep Q 80/100 6 foot 2 millimeter stainless column, and then analyzed on a Thermo MAT 253 Isotope Ratio Mass Spectrometer (IRMS) in Dual Inlet mode. This system was built at NASA/JSC over the past 3 years and proof-tested with known carbonate standards to develop procedures, assess yield, and quantify expected error bands. Two distinct species of carbonates are found: 1) calcite, and 2) non-calcite carbonate (future testing will attempt to differentiate siderite from magnesite). Preliminary results indicate the terrestrial carbonates are formed at approximately sigma (sup 13) C equal to plus 5 per mille, which is consistent with atmospheric CO (sub 2) sigma (sup 13) C equal to minus 7 per mille and fractionation of plus

  4. Normal Hematopoietic Progenitor Subsets Have Distinct Reactive Oxygen Species, BCL2 and Cell-Cycle Profiles That Are Decoupled from Maturation in Acute Myeloid Leukemia

    PubMed Central

    Hills, Robert K.; Knapper, Steve; Steadman, Lora; Qureshi, Ushna; Rector, Jerrald L.; Bradbury, Charlotte; Russell, Nigel H.; Vyas, Paresh; Burnett, Alan K.; Grimwade, David; Hole, Paul S.; Freeman, Sylvie D.

    2016-01-01

    In acute myeloid leukemia (AML) quiescence and low oxidative state, linked to BCL2 mitochondrial regulation, endow leukemic stem cells (LSC) with treatment-resistance. LSC in CD34+ and more mature CD34− AML have heterogeneous immunophenotypes overlapping with normal stem/progenitor cells (SPC) but may be differentiated by functional markers. We therefore investigated the oxidative/reactive oxygen species (ROS) profile, its relationship with cell-cycle/BCL2 for normal SPC, and whether altered in AML and myelodysplasia (MDS). In control BM (n = 24), ROS levels were highest in granulocyte-macrophage progenitors (GMP) and CD34− myeloid precursors but megakaryocyte-erythroid progenitors had equivalent levels to CD34+CD38low immature-SPC although they were ki67high. BCL2 upregulation was specific to GMPs. This profile was also observed for CD34+SPC in MDS-without-excess-blasts (MDS-noEB, n = 12). Erythroid CD34− precursors were, however, abnormally ROS-high in MDS-noEB, potentially linking oxidative stress to cell loss. In pre-treatment AML (n = 93) and MDS-with-excess-blasts (MDS-RAEB) (n = 14), immunophenotypic mature-SPC had similar ROS levels to co-existing immature-SPC. However ROS levels varied between AMLs; Flt3ITD+/NPM1wild-type CD34+SPC had higher ROS than NPM1mutated CD34+ or CD34− SPC. An aberrant ki67lowBCL2high immunophenotype was observed in CD34+AML (most prominent in Flt3ITD AMLs) but also in CD34− AMLs and MDS-RAEB, suggesting a shared redox/pro-survival adaptation. Some patients had BCL2 overexpression in CD34+ ROS-high as well as ROS-low fractions which may be indicative of poor early response to standard chemotherapy. Thus normal SPC subsets have distinct ROS, cell-cycle, BCL2 profiles that in AML /MDS-RAEB are decoupled from maturation. The combined profile of these functional properties in AML subpopulations may be relevant to differential treatment resistance. PMID:27669008

  5. Increased Reactive Oxygen Species Production and Lower Abundance of Complex I Subunits and Carnitine Palmitoyltransferase 1B Protein Despite Normal Mitochondrial Respiration in Insulin-Resistant Human Skeletal Muscle

    PubMed Central

    Lefort, Natalie; Glancy, Brian; Bowen, Benjamin; Willis, Wayne T.; Bailowitz, Zachary; De Filippis, Elena A.; Brophy, Colleen; Meyer, Christian; Højlund, Kurt; Yi, Zhengping; Mandarino, Lawrence J.

    2010-01-01

    OBJECTIVE The contribution of mitochondrial dysfunction to skeletal muscle insulin resistance remains elusive. Comparative proteomics are being applied to generate new hypotheses in human biology and were applied here to isolated mitochondria to identify novel changes in mitochondrial protein abundance present in insulin-resistant muscle. RESEARCH DESIGN AND METHODS Mitochondria were isolated from vastus lateralis muscle from lean and insulin-sensitive individuals and from obese and insulin-resistant individuals who were otherwise healthy. Respiration and reactive oxygen species (ROS) production rates were measured in vitro. Relative abundances of proteins detected by mass spectrometry were determined using a normalized spectral abundance factor method. RESULTS NADH- and FADH2-linked maximal respiration rates were similar between lean and obese individuals. Rates of pyruvate and palmitoyl-dl-carnitine (both including malate) ROS production were significantly higher in obesity. Mitochondria from obese individuals maintained higher (more negative) extramitochondrial ATP free energy at low metabolic flux, suggesting that stronger mitochondrial thermodynamic driving forces may underlie the higher ROS production. Tandem mass spectrometry identified protein abundance differences per mitochondrial mass in insulin resistance, including lower abundance of complex I subunits and enzymes involved in the oxidation of branched-chain amino acids (BCAA) and fatty acids (e.g., carnitine palmitoyltransferase 1B). CONCLUSIONS We provide data suggesting normal oxidative capacity of mitochondria in insulin-resistant skeletal muscle in parallel with high rates of ROS production. Furthermore, we show specific abundance differences in proteins involved in fat and BCAA oxidation that might contribute to the accumulation of lipid and BCAA frequently associated with the pathogenesis of insulin resistance. PMID:20682693

  6. The Arabidopsis KINβγ Subunit of the SnRK1 Complex Regulates Pollen Hydration on the Stigma by Mediating the Level of Reactive Oxygen Species in Pollen

    PubMed Central

    Zhao, Ting Ting; Li, Fei; Jia, Xiao Na; Zhao, Xin-Ying; Zhang, Xian Sheng

    2016-01-01

    Pollen–stigma interactions are essential for pollen germination. The highly regulated process of pollen germination includes pollen adhesion, hydration, and germination on the stigma. However, the internal signaling of pollen that regulates pollen–stigma interactions is poorly understood. KINβγ is a plant-specific subunit of the SNF1-related protein kinase 1 complex which plays important roles in the regulation of plant development. Here, we showed that KINβγ was a cytoplasm- and nucleus-localized protein in the vegetative cells of pollen grains in Arabidopsis. The pollen of the Arabidopsis kinβγ mutant could not germinate on stigma, although it germinated normally in vitro. Further analysis revealed the hydration of kinβγ mutant pollen on the stigma was compromised. However, adding water to the stigma promoted the germination of the mutant pollen in vivo, suggesting that the compromised hydration of the mutant pollen led to its defective germination. In kinβγ mutant pollen, the structure of the mitochondria and peroxisomes was destroyed, and their numbers were significantly reduced compared with those in the wild type. Furthermore, we found that the kinβγ mutant exhibited reduced levels of reactive oxygen species (ROS) in pollen. The addition of H2O2 in vitro partially compensated for the reduced water absorption of the mutant pollen, and reducing ROS levels in pollen by overexpressing Arabidopsis CATALASE 3 resulted in compromised hydration of pollen on the stigma. These results indicate that Arabidopsis KINβγ is critical for the regulation of ROS levels by mediating the biogenesis of mitochondria and peroxisomes in pollen, which is required for pollen–stigma interactions during pollination. PMID:27472382

  7. The Arabidopsis KINβγ Subunit of the SnRK1 Complex Regulates Pollen Hydration on the Stigma by Mediating the Level of Reactive Oxygen Species in Pollen.

    PubMed

    Gao, Xin-Qi; Liu, Chang Zhen; Li, Dan Dan; Zhao, Ting Ting; Li, Fei; Jia, Xiao Na; Zhao, Xin-Ying; Zhang, Xian Sheng

    2016-07-01

    Pollen-stigma interactions are essential for pollen germination. The highly regulated process of pollen germination includes pollen adhesion, hydration, and germination on the stigma. However, the internal signaling of pollen that regulates pollen-stigma interactions is poorly understood. KINβγ is a plant-specific subunit of the SNF1-related protein kinase 1 complex which plays important roles in the regulation of plant development. Here, we showed that KINβγ was a cytoplasm- and nucleus-localized protein in the vegetative cells of pollen grains in Arabidopsis. The pollen of the Arabidopsis kinβγ mutant could not germinate on stigma, although it germinated normally in vitro. Further analysis revealed the hydration of kinβγ mutant pollen on the stigma was compromised. However, adding water to the stigma promoted the germination of the mutant pollen in vivo, suggesting that the compromised hydration of the mutant pollen led to its defective germination. In kinβγ mutant pollen, the structure of the mitochondria and peroxisomes was destroyed, and their numbers were significantly reduced compared with those in the wild type. Furthermore, we found that the kinβγ mutant exhibited reduced levels of reactive oxygen species (ROS) in pollen. The addition of H2O2 in vitro partially compensated for the reduced water absorption of the mutant pollen, and reducing ROS levels in pollen by overexpressing Arabidopsis CATALASE 3 resulted in compromised hydration of pollen on the stigma. These results indicate that Arabidopsis KINβγ is critical for the regulation of ROS levels by mediating the biogenesis of mitochondria and peroxisomes in pollen, which is required for pollen-stigma interactions during pollination. PMID:27472382

  8. GABAA receptor beta subunit heterogeneity: functional expression of cloned cDNAs.

    PubMed Central

    Ymer, S; Schofield, P R; Draguhn, A; Werner, P; Köhler, M; Seeburg, P H

    1989-01-01

    Cloned cDNAs encoding two new beta subunits of the rat and bovine GABAA receptor have been isolated using a degenerate oligonucleotide probe based on a highly conserved peptide sequence in the second transmembrane domain of GABAA receptor subunits. The beta 2 and beta 3 subunits share approximately 72% sequence identity with the previously characterized beta 1 polypeptide. Northern analysis showed that both beta 2 and beta 3 mRNAs are more abundant in the brain than beta 1 mRNA. All three beta subunit encoding cDNAs were also identified in a library constructed from adrenal medulla RNA. Each beta subunit, when co-expressed in Xenopus oocytes with an alpha subunit, forms functional GABAA receptors. These results, together with the known alpha subunit heterogeneity, suggest that a variety of related but functionally distinct GABAA receptor subtypes are generated by different subunit combinations. Images PMID:2548852

  9. Modification of K+ channel–drug interactions by ancillary subunits

    PubMed Central

    Bett, Glenna C L; Rasmusson, Randall L

    2008-01-01

    Reconciling ion channel α-subunit expression with native ionic currents and their pharmacological sensitivity in target organs has proved difficult. In native tissue, many K+ channel α-subunits co-assemble with ancillary subunits, which can profoundly affect physiological parameters including gating kinetics and pharmacological interactions. In this review, we examine the link between voltage-gated potassium ion channel pharmacology and the biophysics of ancillary subunits. We propose that ancillary subunits can modify the interaction between pore blockers and ion channels by three distinct mechanisms: changes in (1) binding site accessibility; (2) orientation of pore-lining residues; (3) the ability of the channel to undergo post-binding conformational changes. Each of these subunit-induced changes has implications for gating, drug affinity and use dependence of their respective channel complexes. A single subunit may modulate its associated α-subunit by more than one of these mechanisms. Voltage-gated potassium channels are the site of action of many therapeutic drugs. In addition, potassium channels interact with drugs whose primary target is another channel, e.g. the calcium channel blocker nifedipine, the sodium channel blocker quinidine, etc. Even when K+ channel block is the intended mode of action, block of related channels in non-target organs, e.g. the heart, can result in major and potentially lethal side-effects. Understanding factors that determine specificity, use dependence and other properties of K+ channel drug binding are therefore of vital clinical importance. Ancillary subunits play a key role in determining these properties in native tissue, and so understanding channel–subunit interactions is vital to understanding clinical pharmacology. PMID:18096604

  10. The Lesser of Two Weevils: Molecular-Genetics of Pest Palm Weevil Populations Confirm Rhynchophorus vulneratus (Panzer 1798) as a Valid Species Distinct from R. ferrugineus (Olivier 1790), and Reveal the Global Extent of Both

    PubMed Central

    Rugman-Jones, Paul F.; Hoddle, Christina D.; Hoddle, Mark S.; Stouthamer, Richard

    2013-01-01

    The red palm weevil (RPW) is a major pest of palms. It is native to southeast Asia and Melanesia, but in recent decades has vastly expanded its range as the result of multiple accidental anthropogenic introductions into the Middle East, Mediterranean Basin, Caribbean, and U.S.A. Currently regarded as a single species, Rhynchophorus ferrugineus (Olivier), RPW displays remarkable color variation across its range, and consequently has a taxonomic history littered with new species descriptions and synonymization. We compared DNA sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene from RPW populations throughout the native and invaded ranges, to investigate the specific status and invasion history of this serious economic pest, and to identify possible common routes of entry. Analyses of COI haplotype data provide conclusive support, corroborated by sequences of additional nuclear gene regions, for the existence of at least two predominantly allopatric species. The true R. ferrugineus is native only to the northern and western parts of continental southeast Asia, Sri Lanka and the Philippines, and is responsible for almost all invasive populations worldwide. In contrast, the second species, which is currently synonymized under R. ferrugineus and should be resurrected under the name R. vulneratus (Panzer), has a more southern distribution across Indonesia, and is responsible for only one invasive population; that in California, U.S.A. The distribution of COI haplotypes is used to discuss the possible existence of further cryptic species, sources and routes of entry of different invasive populations, and the implications of our findings for current control methods. PMID:24143263

  11. Multigene Phylogeography of Bactrocera caudata (Insecta: Tephritidae): Distinct Genetic Lineages in Northern and Southern Hemispheres.

    PubMed

    Yong, Hoi-Sen; Lim, Phaik-Eem; Tan, Ji; Song, Sze-Looi; Suana, I Wayan; Eamsobhana, Praphathip

    2015-01-01

    Bactrocera caudata is a pest of pumpkin flower. Specimens of B. caudata from the northern hemisphere (mainland Asia) and southern hemisphere (Indonesia) were analysed using the partial DNA sequences of the nuclear 28S rRNA and internal transcribed spacer region 2 (ITS-2) genes, and the mitochondrial cytochrome c oxidase subunit I (COI), cytochrome c oxidase subunit II (COII) and 16S rRNA genes. The COI, COII, 16S rDNA and concatenated COI+COII+16S and COI+COII+16S+28S+ITS-2 nucleotide sequences revealed that B. caudata from the northern hemisphere (Peninsular Malaysia, East Malaysia, Thailand) was distinctly different from the southern hemisphere (Indonesia: Java, Bali and Lombok), without common haplotype between them. Phylogenetic analysis revealed two distinct clades (northern and southern hemispheres), indicating distinct genetic lineage. The uncorrected 'p' distance for the concatenated COI+COII+16S nucleotide sequences between the taxa from the northern and southern hemispheres ('p' = 4.46-4.94%) was several folds higher than the 'p' distance for the taxa in the northern hemisphere ('p' = 0.00-0.77%) and the southern hemisphere ('p' = 0.00%). This distinct difference was also reflected by concatenated COI+COII+16S+28S+ITS-2 nucleotide sequences with an uncorrected 'p' distance of 2.34-2.69% between the taxa of northern and southern hemispheres. In accordance with the type locality the Indonesian taxa belong to the nominal species. Thus the taxa from the northern hemisphere, if they were to constitute a cryptic species of the B. caudata species complex based on molecular data, need to be formally described as a new species. The Thailand and Malaysian B. caudata populations in the northern hemisphere showed distinct genetic structure and phylogeographic pattern.

  12. Multigene Phylogeography of Bactrocera caudata (Insecta: Tephritidae): Distinct Genetic Lineages in Northern and Southern Hemispheres

    PubMed Central

    Yong, Hoi-Sen; Lim, Phaik-Eem; Tan, Ji; Song, Sze-Looi; Suana, I Wayan; Eamsobhana, Praphathip

    2015-01-01

    Bactrocera caudata is a pest of pumpkin flower. Specimens of B. caudata from the northern hemisphere (mainland Asia) and southern hemisphere (Indonesia) were analysed using the partial DNA sequences of the nuclear 28S rRNA and internal transcribed spacer region 2 (ITS-2) genes, and the mitochondrial cytochrome c oxidase subunit I (COI), cytochrome c oxidase subunit II (COII) and 16S rRNA genes. The COI, COII, 16S rDNA and concatenated COI+COII+16S and COI+COII+16S+28S+ITS-2 nucleotide sequences revealed that B. caudata from the northern hemisphere (Peninsular Malaysia, East Malaysia, Thailand) was distinctly different from the southern hemisphere (Indonesia: Java, Bali and Lombok), without common haplotype between them. Phylogenetic analysis revealed two distinct clades (northern and southern hemispheres), indicating distinct genetic lineage. The uncorrected ‘p’ distance for the concatenated COI+COII+16S nucleotide sequences between the taxa from the northern and southern hemispheres (‘p’ = 4.46-4.94%) was several folds higher than the ‘p’ distance for the taxa in the northern hemisphere (‘p’ = 0.00-0.77%) and the southern hemisphere (‘p’ = 0.00%). This distinct difference was also reflected by concatenated COI+COII+16S+28S+ITS-2 nucleotide sequences with an uncorrected 'p' distance of 2.34-2.69% between the taxa of northern and southern hemispheres. In accordance with the type locality the Indonesian taxa belong to the nominal species. Thus the taxa from the northern hemisphere, if they were to constitute a cryptic species of the B. caudata species complex based on molecular data, need to be formally described as a new species. The Thailand and Malaysian B. caudata populations in the northern hemisphere showed distinct genetic structure and phylogeographic pattern. PMID:26090853

  13. Multigene Phylogeography of Bactrocera caudata (Insecta: Tephritidae): Distinct Genetic Lineages in Northern and Southern Hemispheres.

    PubMed

    Yong, Hoi-Sen; Lim, Phaik-Eem; Tan, Ji; Song, Sze-Looi; Suana, I Wayan; Eamsobhana, Praphathip

    2015-01-01

    Bactrocera caudata is a pest of pumpkin flower. Specimens of B. caudata from the northern hemisphere (mainland Asia) and southern hemisphere (Indonesia) were analysed using the partial DNA sequences of the nuclear 28S rRNA and internal transcribed spacer region 2 (ITS-2) genes, and the mitochondrial cytochrome c oxidase subunit I (COI), cytochrome c oxidase subunit II (COII) and 16S rRNA genes. The COI, COII, 16S rDNA and concatenated COI+COII+16S and COI+COII+16S+28S+ITS-2 nucleotide sequences revealed that B. caudata from the northern hemisphere (Peninsular Malaysia, East Malaysia, Thailand) was distinctly different from the southern hemisphere (Indonesia: Java, Bali and Lombok), without common haplotype between them. Phylogenetic analysis revealed two distinct clades (northern and southern hemispheres), indicating distinct genetic lineage. The uncorrected 'p' distance for the concatenated COI+COII+16S nucleotide sequences between the taxa from the northern and southern hemispheres ('p' = 4.46-4.94%) was several folds higher than the 'p' distance for the taxa in the northern hemisphere ('p' = 0.00-0.77%) and the southern hemisphere ('p' = 0.00%). This distinct difference was also reflected by concatenated COI+COII+16S+28S+ITS-2 nucleotide sequences with an uncorrected 'p' distance of 2.34-2.69% between the taxa of northern and southern hemispheres. In accordance with the type locality the Indonesian taxa belong to the nominal species. Thus the taxa from the northern hemisphere, if they were to constitute a cryptic species of the B. caudata species complex based on molecular data, need to be formally described as a new species. The Thailand and Malaysian B. caudata populations in the northern hemisphere showed distinct genetic structure and phylogeographic pattern. PMID:26090853

  14. Genetic analysis of the cytoplasmic dynein subunit families.

    PubMed

    Pfister, K Kevin; Shah, Paresh R; Hummerich, Holger; Russ, Andreas; Cotton, James; Annuar, Azlina Ahmad; King, Stephen M; Fisher, Elizabeth M C

    2006-01-01

    Cytoplasmic dyneins, the principal microtubule minus-end-directed motor proteins of the cell, are involved in many essential cellular processes. The major form of this enzyme is a complex of at least six protein subunits, and in mammals all but one of the subunits are encoded by at least two genes. Here we review current knowledge concerning the subunits, their interactions, and their functional roles as derived from biochemical and genetic analyses. We also carried out extensive database searches to look for new genes and to clarify anomalies in the databases. Our analysis documents evolutionary relationships among the dynein subunits of mammals and other model organisms, and sheds new light on the role of this diverse group of proteins, highlighting the existence of two cytoplasmic dynein complexes with distinct cellular roles.

  15. Genetic Analysis of the Cytoplasmic Dynein Subunit Families

    PubMed Central

    2006-01-01

    Cytoplasmic dyneins, the principal microtubule minus-end-directed motor proteins of the cell, are involved in many essential cellular processes. The major form of this enzyme is a complex of at least six protein subunits, and in mammals all but one of the subunits are encoded by at least two genes. Here we review current knowledge concerning the subunits, their interactions, and their functional roles as derived from biochemical and genetic analyses. We also carried out extensive database searches to look for new genes and to clarify anomalies in the databases. Our analysis documents evolutionary relationships among the dynein subunits of mammals and other model organisms, and sheds new light on the role of this diverse group of proteins, highlighting the existence of two cytoplasmic dynein complexes with distinct cellular roles. PMID:16440056

  16. Megraft: A software package to graft ribosomal small subunit (16S/18S) fragments onto full-length sequences for accurate species richness and sequencing depth analysis in pyrosequencing-length metagenomes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Metagenomic libraries represent subsamples of the total DNA found at a study site and offer unprecedented opportunities to study ecological and functional aspects of microbial communities. To examine the depth of the sequencing effort, rarefaction analysis of the ribosomal small sub-unit (SSU/16S/18...

  17. Structural determinants of alpha-bungarotoxin binding to the sequence segment 181-200 of the muscle nicotinic acetylcholine receptor. alpha. subunit: Effects of cysteine/cystine modification and species-specific amino acid substitution

    SciTech Connect

    McLane, K.E.; Wu, Xiadong; Diethelm, B.; Conti-Tronconi, B.M. )

    1991-05-21

    The sequence segment 181-200 of the Torpedo nicotinic acetylcholine receptor (nAChR) {alpha}subunit forms a binding site for {alpha}-bungarotoxin ({alpha}-BTX). Synthetic peptides corresponding to the homologous sequences of human, calf, mouse, chicken, frog, and cobra muscle nAChR {alpha}1 subunits were tested for their ability to bind {sup 125}I-{alpha}-BTX, and differences in {alpha}-BTX affinity were determined by using solution (IC{sub 50}s) and solid-phase (K{sub d}s) assays. Panels of overlapping peptides corresponding to the complete {alpha}1 subunit of mouse and human were also tested for {alpha}-BTX binding, but other sequence segments forming the {alpha}-BTX site were not consistently detectable. The role of a putative vicinal disulfide bound between Cys-192 and -193, relative to the Torpedo sequence, was determined by modifying the peptides with sulfhydryl reagents. Reduction and alkylation of the peptides decreased {alpha}-BTX binding, whereas oxidation of the peptides had little effect. These results indicate that while the adjacent cysteines are likely to be involved in forming the toxin/{alpha}1-subunit interface a vicinal disulfide bound was not required for {alpha}-BTX binding.

  18. Within-island speciation with an exceptional case of distinct separation between two sibling lizard species divided by a narrow stream.

    PubMed

    Tseng, Shu-Ping; Wang, Chao-Jun; Li, Shou-Hsien; Lin, Si-Min

    2015-09-01

    Delimitation of genetic and geographic boundaries between species is a focus of evolutionary biology. In this study, we demonstrated fine-scale differentiation of Takydromus formosanus species complex comprising four insular endemics on Taiwan Island. Phylogeny and ancestral range reconstruction based on mitochondrial DNA sequences of 430 Takydromus lizards (405 lizards of this complex throughout their distribution range, and 25 lizards from 11 other species) indicated that the major branching process occurred within Taiwan, which represented a solid evidence of within-island speciation on this small island. We further demonstrated an exceptional case of a pair of sister species, T. viridipunctatus and T. luyeanus, that were separated by a narrow stream with a width of only 15m. This pattern might be one of the narrowest contact zones ever documented in terrestrial vertebrates. To evaluate the level of genetic introgression between these sister species, a fine-scale collection of another 382 lizards was conducted along a transect line across the stream. A total of 13 microsatellite markers and mtDNA genotyping was used to detect a low proportion of hybrids (5.7-9.9% from STRUCTURE, and 2.3% from DAPC). Our results indicated that the two clades are highly differentiated across this extremely short distance.

  19. Modulation of BK Channel Function by Auxiliary Beta and Gamma Subunits

    PubMed Central

    Li, Q.; Yan, J.

    2016-01-01

    The large-conductance, Ca2+- and voltage-activated K+ (BK) channel is ubiquitously expressed in mammalian tissues and displays diverse biophysical or pharmacological characteristics. This diversity is in part conferred by channel modulation with different regulatory auxiliary subunits. To date, two distinct classes of BK channel auxiliary subunits have been identified: β subunits and γ subunits. Modulation of BK channels by the four auxiliary β (β1–β4) subunits has been well established and intensively investigated over the past two decades. The auxiliary γ subunits, however, were identified only very recently, which adds a new dimension to BK channel regulation and improves our understanding of the physiological functions of BK channels in various tissues and cell types. This chapter will review the current understanding of BK channel modulation by auxiliary β and γ subunits, especially the latest findings. PMID:27238261

  20. Variability of the 5S and 45S rDNA Sites in Passiflora L. Species with Distinct Base Chromosome Numbers

    PubMed Central

    DE MELO, NATONIEL FRANKLIN; GUERRA, MARCELO

    2003-01-01

    Cytologically, the species of Passiflora with known chromosome number can be divided into four groups: (1) 2n = 12, 24, 36; (2) 2n = 24; (3) 2n = 18, 72; and (4) 2n = 20. The base chromosome number proposed for the genus is x = 6, with x = 9, x = 10 and x = 12 being considered secondary base numbers. In the present study, variability of 5S and 45S rDNA sites was investigated in 20 species of these four groups to check the reliability of this hypothesis. In the group with x = 6, five diploid species (2n = 12) exhibit two 5S rDNA sites and two (P. capsularis, P. morifolia and P. rubra) or four (P. misera 2x and P. tricuspis) 45S rDNA sites. The hexaploid cytotype of P. misera had 12 45S rDNA sites and six 5S rDNA. A tetraploid species, P. suberosa, had ten 45S rDNA sites and four 5S rDNA sites, both in the same chromosomes as the 45S rDNA sites. In the group with x = 9, P. actinia, P. amethystina, P. edmundoi, P. elegans, P. galbana, P. glandulosa and P. mucronata displayed six 45S rDNA sites, whereas P. alata, P. cincinnata, P. edulis f. flavicarpa, P. edulis var. roxo and P. laurifolia had four sites. In this group, all species were diploid (2n = 18) and had only two 5S rDNA sites. Passiflora foetida, the only species with 2n = 20, had six 45S rDNA sites and four 5S rDNA sites. The species with x = 12 (2n = 24), P. haematostigma and P. pentagona, showed four 45S rDNA sites and two 5S rDNA. In general, the number and location of 5S and 45S rDNA sites were consistent with the hypothesis of x = 6 as the probable ancestral genome for the genus, while the groups of species with x = 9, x = 10 and x = 12 were considered to be of tetraploid origin with descending dysploidy and gene silencing of some redundant gene sites, mainly those of 5S rDNA. PMID:12876193

  1. Variability of the 5S and 45S rDNA sites in Passiflora L. species with distinct base chromosome numbers.

    PubMed

    de Melo, Natoniel Franklin; Guerra, Marcelo

    2003-08-01

    Cytologically, the species of Passiflora with known chromosome number can be divided into four groups: (1) 2n = 12, 24, 36; (2) 2n = 24; (3) 2n = 18, 72; and (4) 2n = 20. The base chromosome number proposed for the genus is x = 6, with x = 9, x = 10 and x = 12 being considered secondary base numbers. In the present study, variability of 5S and 45S rDNA sites was investigated in 20 species of these four groups to check the reliability of this hypothesis. In the group with x = 6, five diploid species (2n = 12) exhibit two 5S rDNA sites and two (P. capsularis, P. morifolia and P. rubra) or four (P. misera 2x and P. tricuspis) 45S rDNA sites. The hexaploid cytotype of P. misera had 12 45S rDNA sites and six 5S rDNA. A tetraploid species, P. suberosa, had ten 45S rDNA sites and four 5S rDNA sites, both in the same chromosomes as the 45S rDNA sites. In the group with x = 9, P. actinia, P. amethystina, P. edmundoi, P. elegans, P. galbana, P. glandulosa and P. mucronata displayed six 45S rDNA sites, whereas P. alata, P. cincinnata, P. edulis f. flavicarpa, P. edulis var. roxo and P. laurifolia had four sites. In this group, all species were diploid (2n = 18) and had only two 5S rDNA sites. Passiflora foetida, the only species with 2n = 20, had six 45S rDNA sites and four 5S rDNA sites. The species with x = 12 (2n = 24), P. haematostigma and P. pentagona, showed four 45S rDNA sites and two 5S rDNA. In general, the number and location of 5S and 45S rDNA sites were consistent with the hypothesis of x = 6 as the probable ancestral genome for the genus, while the groups of species with x = 9, x = 10 and x = 12 were considered to be of tetraploid origin with descending dysploidy and gene silencing of some redundant gene sites, mainly those of 5S rDNA. PMID:12876193

  2. Complete subunit sequences, structure and evolution of the 6 x 6-mer hemocyanin from the common house centipede, Scutigera coleoptrata.

    PubMed

    Kusche, Kristina; Hembach, Anne; Hagner-Holler, Silke; Gebauer, Wolfgang; Burmester, Thorsten

    2003-07-01

    Hemocyanins are large oligomeric copper-containing proteins that serve for the transport of oxygen in many arthropod species. While studied in detail in the Chelicerata and Crustacea, hemocyanins had long been considered unnecessary in the Myriapoda. Here we report the complete molecular structure of the hemocyanin from the common house centipede Scutigera coleoptrata (Myriapoda: Chilopoda), as deduced from 2D-gel electrophoresis, MALDI-TOF mass spectrometry, protein and cDNA sequencing, and homology modeling. This is the first myriapod hemocyanin to be fully sequenced, and allows the investigation of hemocyanin structure-function relationship and evolution. S. coleoptrata hemocyanin is a 6 x 6-mer composed of four distinct subunit types that occur in an approximate 2 : 2 : 1 : 1 ratio and are 49.5-55.5% identical. The cDNA of a fifth, highly diverged, putative hemocyanin was identified that is not included in the native 6 x 6-mer hemocyanin. Phylogenetic analyses show that myriapod hemocyanins are monophyletic, but at least three distinct subunit types evolved before the separation of the Chilopoda and Diplopoda more than 420 million years ago. In contrast to the situation in the Crustacea and Chelicerata, the substitution rates among the myriapod hemocyanin subunits are highly variable. Phylogenetic analyses do not support a common clade of Myriapoda and Hexapoda, whereas there is evidence in favor of monophyletic Mandibulata.

  3. Transcription, Signaling Receptor Activity, Oxidative Phosphorylation, and Fatty Acid Metabolism Mediate the Presence of Closely Related Species in Distinct Intertidal and Cold-Seep Habitats.

    PubMed

    Van Campenhout, Jelle; Vanreusel, Ann; Van Belleghem, Steven; Derycke, Sofie

    2016-01-01

    Bathyal cold seeps are isolated extreme deep-sea environments characterized by low species diversity while biomass can be high. The Håkon Mosby mud volcano (Barents Sea, 1,280 m) is a rather stable chemosynthetic driven habitat characterized by prominent surface bacterial mats with high sulfide concentrations and low oxygen levels. Here, the nematode Halomonhystera hermesi thrives in high abundances (11,000 individuals 10 cm(-2)). Halomonhystera hermesi is a member of the intertidal Halomonhystera disjuncta species complex that includes five cryptic species (GD1-5). GD1-5's common habitat is characterized by strong environmental fluctuations. Here, we compared the transcriptomes of H. hermesi and GD1, H. hermesi's closest relative. Genes encoding proteins involved in oxidative phosphorylation are more strongly expressed in H. hermesi than in GD1, and many genes were only observed in H. hermesi while being completely absent in GD1. Both observations could in part be attributed to high sulfide concentrations and low oxygen levels. Additionally, fatty acid elongation was also prominent in H. hermesi confirming the importance of highly unsaturated fatty acids in this species. Significant higher amounts of transcription factors and genes involved in signaling receptor activity were observed in GD1 (many of which were completely absent in H. hermesi), allowing fast signaling and transcriptional reprogramming which can mediate survival in dynamic intertidal environments. GC content was approximately 8% higher in H. hermesi coding unigenes resulting in differential codon usage between both species and a higher proportion of amino acids with GC-rich codons in H. hermesi. In general our results showed that most pathways were active in both environments and that only three genes are under natural selection. This indicates that also plasticity should be taken in consideration in the evolutionary history of Halomonhystera species. Such plasticity, as well as possible

  4. Transcription, Signaling Receptor Activity, Oxidative Phosphorylation, and Fatty Acid Metabolism Mediate the Presence of Closely Related Species in Distinct Intertidal and Cold-Seep Habitats

    PubMed Central

    Van Campenhout, Jelle; Vanreusel, Ann; Van Belleghem, Steven; Derycke, Sofie

    2016-01-01

    Bathyal cold seeps are isolated extreme deep-sea environments characterized by low species diversity while biomass can be high. The Håkon Mosby mud volcano (Barents Sea, 1,280 m) is a rather stable chemosynthetic driven habitat characterized by prominent surface bacterial mats with high sulfide concentrations and low oxygen levels. Here, the nematode Halomonhystera hermesi thrives in high abundances (11,000 individuals 10 cm−2). Halomonhystera hermesi is a member of the intertidal Halomonhystera disjuncta species complex that includes five cryptic species (GD1-5). GD1-5’s common habitat is characterized by strong environmental fluctuations. Here, we compared the transcriptomes of H. hermesi and GD1, H. hermesi’s closest relative. Genes encoding proteins involved in oxidative phosphorylation are more strongly expressed in H. hermesi than in GD1, and many genes were only observed in H. hermesi while being completely absent in GD1. Both observations could in part be attributed to high sulfide concentrations and low oxygen levels. Additionally, fatty acid elongation was also prominent in H. hermesi confirming the importance of highly unsaturated fatty acids in this species. Significant higher amounts of transcription factors and genes involved in signaling receptor activity were observed in GD1 (many of which were completely absent in H. hermesi), allowing fast signaling and transcriptional reprogramming which can mediate survival in dynamic intertidal environments. GC content was approximately 8% higher in H. hermesi coding unigenes resulting in differential codon usage between both species and a higher proportion of amino acids with GC-rich codons in H. hermesi. In general our results showed that most pathways were active in both environments and that only three genes are under natural selection. This indicates that also plasticity should be taken in consideration in the evolutionary history of Halomonhystera species. Such plasticity, as well as possible

  5. Genetic variation in the mitochondrial cytochrome c oxidase subunit 1 within Progamotaenia festiva (Cestoda: Anoplocephalidae) from macropodid marsupials.

    PubMed

    Beveridge, I; Shamsi, S; Hu, M; Chilton, N B; Gasser, R B

    2007-09-01

    Genetic variation was examined in the anoplocephalid cestode Progamotaenia festiva, from Australian marsupials, in order to test the hypothesis that P. festiva, is a complex of sibling species and to assess the extent of host switching reported previously based on multilocus enzyme electrophoresis (MEE). Polymerase chain reaction (PCR)-based single-strand conformational polymorphism (SSCP) was used for the analysis of sequence variation in the cytochrome c oxidase subunit 1 (cox1) gene among 179 specimens of P. festiva (identified based on morphology and predilection site in the host) from 13 different host species, followed by selective DNA sequencing. Fifty-three distinct sequence types (haplotypes) representing all specimens were defined. Phylogenetic analyses of these sequence data (utilizing maximum parsimony and neighbour-joining methods) revealed 12 distinct clades. Other heterologous species, P. ewersi and P. macropodis, were used as outgroups and the remaining bile-duct inhabiting species, P. diaphana and P. effigia, were included in the analysis for comparative purposes. The latter 2 species were nested within the clades representing P. festiva. Most clades of P. festiva identified were restricted to a single host species; one clade primarily in Macropus robustus was also found in the related host species M. antilopinus in an area of host sympatry; another clade occurring primarily in M. robustus occurred also in additional kangaroo species, M. rufus and M. dorsalis. High levels of genetic divergence, the existence of distinct clades and their occurrence in sympatry provide support for the hypothesis that P. festiva represents a complex of numerous species, most of which, but not all, are host specific. Three distinct clades of cestodes were found within a single host, M. robustus, but there was no evidence of within-host speciation.

  6. flaB Gene as a Molecular Marker for Distinct Identification of Borrelia Species in Environmental Samples by the PCR-Restriction Fragment Length Polymorphism Method ▿

    PubMed Central

    Wodecka, Beata

    2011-01-01

    A new protocol employing nested PCR-restriction fragment length polymorphism (RFLP) based on the flaB gene and two restriction enzymes was worked out. This protocol allows the identification of all Borrelia species transmitted by Ixodes ricinus in Europe, including Borrelia miyamotoi and 3 genetic variants of B. garinii. A dendrogram of flaB sequence similarity was in accordance with RFLP variants. PMID:21841027

  7. Plasmodium forresteri n. sp., from raptors in Florida and southern Georgia: its distinction from Plasmodium elongatum morphologically within and among host species and by vector susceptibility.

    PubMed

    Telford, S R; Nayar, J K; Foster, G W; Knight, J W

    1997-10-01

    Plasmodium forresteri n. sp. naturally infects eastern screech-owls (Otus asio), great horned owls (Bubo virginianus), barred owls (Strix varia), bald eagles (Haliaeetus leucocephalus), red-shouldered hawks (Buteo lineatus), broad-winged hawks (Buteo platypterus), and red-tailed hawks (Buteo jamaicensis) in Florida and southern Georgia. Schizonts occur in mature or nearly mature erythrocytes, produce 2-6 merozoites arranged most commonly in fan or cruciform configuration, with mean dimensions among host species varying from 3.7 to 4.8 x 2.5 to 3.4 microns. Gametocytes are elongate, with mean dimensions among host species varying from 11.5 to 13.1 x 2.0 to 2.4 microns. One or both gametocyte margins are irregular and often crenulate. Gametocytes seldom fill the space between the erythrocyte nucleus and margin. Species characteristics were maintained in isodiagnostic Japanese quail (Coturnix japonica) and Pekin ducks (Anas platyrhynchos). In mosquito infection studies, only Culex restuans could support sporogony of P. forresteri, in contrast to Plasmodium elongatum of raptor origin that completed sporogony in both Cx. restuans and Culex nigripalpus. PMID:9379302

  8. Plasmodium forresteri n. sp., from raptors in Florida and southern Georgia: its distinction from Plasmodium elongatum morphologically within and among host species and by vector susceptibility.

    PubMed

    Telford, S R; Nayar, J K; Foster, G W; Knight, J W

    1997-10-01

    Plasmodium forresteri n. sp. naturally infects eastern screech-owls (Otus asio), great horned owls (Bubo virginianus), barred owls (Strix varia), bald eagles (Haliaeetus leucocephalus), red-shouldered hawks (Buteo lineatus), broad-winged hawks (Buteo platypterus), and red-tailed hawks (Buteo jamaicensis) in Florida and southern Georgia. Schizonts occur in mature or nearly mature erythrocytes, produce 2-6 merozoites arranged most commonly in fan or cruciform configuration, with mean dimensions among host species varying from 3.7 to 4.8 x 2.5 to 3.4 microns. Gametocytes are elongate, with mean dimensions among host species varying from 11.5 to 13.1 x 2.0 to 2.4 microns. One or both gametocyte margins are irregular and often crenulate. Gametocytes seldom fill the space between the erythrocyte nucleus and margin. Species characteristics were maintained in isodiagnostic Japanese quail (Coturnix japonica) and Pekin ducks (Anas platyrhynchos). In mosquito infection studies, only Culex restuans could support sporogony of P. forresteri, in contrast to Plasmodium elongatum of raptor origin that completed sporogony in both Cx. restuans and Culex nigripalpus.

  9. Developmental and Regulatory Functions of Na(+) Channel Non-pore-forming β Subunits.

    PubMed

    Winters, J J; Isom, L L

    2016-01-01

    Voltage-gated Na(+) channels (VGSCs) isolated from mammalian neurons are heterotrimeric complexes containing one pore-forming α subunit and two non-pore-forming β subunits. In excitable cells, VGSCs are responsible for the initiation of action potentials. VGSC β subunits are type I topology glycoproteins, containing an extracellular amino-terminal immunoglobulin (Ig) domain with homology to many neural cell adhesion molecules (CAMs), a single transmembrane segment, and an intracellular carboxyl-terminal domain. VGSC β subunits are encoded by a gene family that is distinct from the α subunits. While α subunits are expressed in prokaryotes, β subunit orthologs did not arise until after the emergence of vertebrates. β subunits regulate the cell surface expression, subcellular localization, and gating properties of their associated α subunits. In addition, like many other Ig-CAMs, β subunits are involved in cell migration, neurite outgrowth, and axon pathfinding and may function in these roles in the absence of associated α subunits. In sum, these multifunctional proteins are critical for both channel regulation and central nervous system development. PMID:27586289

  10. Microheliella maris (Microhelida ord. n.), an ultrastructurally highly distinctive new axopodial protist species and genus, and the unity of phylum Heliozoa.

    PubMed

    Yabuki, Akinori; Chao, Ema E; Ishida, Ken-Ichiro; Cavalier-Smith, Thomas

    2012-05-01

    A new heliozoan, Microheliella maris, has sufficiently distinctive ultrastructure to merit a new order, Microhelida. Its 18S and 28S rRNA genes were sequenced earlier under the informal name 'marine microheliozoan'; we here sequenced its Hsp90 gene. A three-gene tree suggests that it is distantly related to centrohelids and others in chromist subkingdom Hacrobia; but it is too divergent to be placed accurately by few genes. Unlike centrohelids, its central spherical centrosome has two concentric granular shells and a dense core devoid of a trilaminar central disc. Microtubules radiate from the centrosomal shells. Unlike centrohelids, axopodia have only three microtubules, fixed basally by dense plasma membrane anchors, and bear terminal and lateral haptosome-like extrusomes. As in the heliomonad Heliomorpha, the centrosome is embedded in a nuclear cavity, and centrosomal microtubules traverse the nucleus inside cytoplasmic channels. A novel filogranular network interconnects mitochondria, ER, and plasma membrane. The microbody is attached to the nucleus and mitochondrion, which has vermicular tubular cristae. We group Microhelida and Heliomonadida, purged of dissimilar flagellates, as a new tubulicristate class Endohelea within phylum Heliozoa. Previously misassigned GenBank 18S rDNA sequences reveal Microhelida as diverse and ancient. We discuss principles underlying the biogenesis and diversity of axopodial patterns. PMID:22153838

  11. Genome structure and organization of a member of a novel and distinct species of the genus Caulimovirus associated with dahlia mosaic.

    PubMed

    Pahalawatta, V; Druffel, K L; Wyatt, S D; Eastwell, K C; Pappu, H R

    2008-01-01

    The genome structure and organization of a new and distinct caulimovirus that is widespread in dahlia (Dahlia variabilis) was determined. The double-stranded DNA genome was ca. 7.0 kb in size and shared many of the features of the members of the genus Caulimovirus, such as the presence of genes potentially coding for the movement protein, the inclusion body protein, and the reverse transcriptase (RT), and an intergenic region consisting of a potential 35S promoter. However, the virus differed from the previously described dahlia mosaic caulimovirus and other known caulimoviruses in that the aphid transmission factor (ATF) was absent and the putative coat protein contained a C-terminal deletion and was fused in-frame with the RT. Sequence identity at the amino acid level with known caulimoviruses including a previously reported caulimovirus from dahlia was low and ranged from 32 to 72%. The absence of an ATF and the highly divergent nature of the genomic sequence are characteristics of this new caulimovirus that is widely associated with dahlia. PMID:18253696

  12. The monospecific genus Meredithia (Kallymeniaceae, Gigartinales) is species rich and geographically widespread with species from temperate Atlantic, Pacific, and Indian Oceans.

    PubMed

    Schneider, Craig W; Saunders, Gary W; Lane, Christopher E

    2014-02-01

    Using sequences of 5' region of the cytochrome oxidase subunit 1 gene, large subunit rDNA, and ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit gene as genetic markers to elucidate their phylogenetic positions, six unknown species from Western Australia, Tasmania, Lord Howe Is., and Norfolk Is. cluster with Meredithia in the Kallymeniaceae (Gigartinales), and are described as new members of this previously monospecific genus. Specimens from Bermuda referable to Kallymenia limminghei Mont. in the 20th century also clustered with this genetic grouping, not with the generitype of Kallymenia. The Bermudian specimens are further shown to be morphologically distinct from the type of K. limminghei (Guadeloupe, Caribbean Sea) and are described as a new species, Meredithia crenata. Using these Indo-Pacific and Bermudian collections, our analyses further show that Psaromenia is closely related to Meredithia, and that Cirrulicarpus nanus sensu stricto should be returned to Meredithia.

  13. Distinct Cell-Specific Expression of Homospermidine Synthase Involved in Pyrrolizidine Alkaloid Biosynthesis in Three Species of the Boraginales1[C][W][OA

    PubMed Central

    Niemüller, Daniel; Reimann, Andreas; Ober, Dietrich

    2012-01-01

    Homospermidine synthase (HSS) is the first specific enzyme in pyrrolizidine alkaloid (PA) biosynthesis, a pathway involved in the plant’s chemical defense. HSS has been shown to be recruited repeatedly by duplication of a gene involved in primary metabolism. Within the lineage of the Boraginales, only one gene duplication event gave rise to HSS. Here, we demonstrate that the tissue-specific expression of HSS in three boraginaceous species, Heliotropium indicum, Symphytum officinale, and Cynoglossum officinale, is unique with respect to plant organ, tissue, and cell type. Within H. indicum, HSS is expressed exclusively in nonspecialized cells of the lower epidermis of young leaves and shoots. In S. officinale, HSS expression has been detected in the cells of the root endodermis and in leaves directly underneath developing inflorescences. In young roots of C. officinale, HSS is detected only in cells of the endodermis, but in a later developmental stage, additionally in the pericycle. The individual expression patterns are compared with those within the Senecioneae lineage (Asteraceae), where HSS expression is reproducibly found in specific cells of the endodermis and the adjacent cortex parenchyma of the roots. The individual expression patterns within the Boraginales species are discussed as being a requirement for the successful recruitment of HSS after gene duplication. The diversity of HSS expression within this lineage adds a further facet to the already diverse patterns of expression that have been observed for HSS in other PA-producing plant lineages, making this PA-specific enzyme one of the most diverse expressed proteins described in the literature. PMID:22566491

  14. Comparative Analyses of Three Chlorella Species in Response to Light and Sugar Reveal Distinctive Lipid Accumulation Patterns in the Microalga C. sorokiniana

    PubMed Central

    Barnes, Austin; Noel, Eric A.; Betenbaugh, Michael J.; Oyler, George A.

    2014-01-01

    While photosynthetic microalgae, such as Chlorella, serve as feedstocks for nutritional oils and biofuels, heterotrophic cultivation can augment growth rates, support high cell densities, and increase triacylglycerol (TAG) lipid content. However, these species differ significantly in their photoautotrophic and heterotrophic characteristics. In this study, the phylogeny of thirty Chlorella strains was determined in order to inform bioprospecting efforts and detailed physiological assessment of three species. The growth kinetics and lipid biochemistry of C. protothecoides UTEX 411, C. vulgaris UTEX 265, and C. sorokiniana UTEX 1230 were quantified during photoautotrophy in Bold's basal medium (BBM) and heterotrophy in BBM supplemented with glucose (10 g L−1). Heterotrophic growth rates of UTEX 411, 265, and 1230 were found to be 1.5-, 3.7-, and 5-fold higher than their respective autotrophic rates. With a rapid nine-hour heterotrophic doubling time, Chlorella sorokiniana UTEX 1230 maximally accumulated 39% total lipids by dry weight during heterotrophy compared to 18% autotrophically. Furthermore, the discrete fatty acid composition of each strain was examined in order to elucidate lipid accumulation patterns under the two trophic conditions. In both modes of growth, UTEX 411 and 265 produced 18∶1 as the principal fatty acid while UTEX 1230 exhibited a 2.5-fold enrichment in 18∶2 relative to 18∶1. Although the total lipid content was highest in UTEX 411 during heterotrophy, UTEX 1230 demonstrated a two-fold increase in its heterotrophic TAG fraction at a rate of 28.9 mg L−1 d−1 to reach 22% of the biomass, corresponding to as much as 90% of its total lipids. Interestingly, UTEX 1230 growth was restricted during mixotrophy and its TAG production rate was suppressed to 18.2 mg L−1 d−1. This constraint on carbon flow raises intriguing questions about the impact of sugar and light on the metabolic regulation of microalgal lipid biosynthesis. PMID:24699196

  15. Similar GABAA receptor subunit composition in somatic and axon initial segment synapses of hippocampal pyramidal cells

    PubMed Central

    Kerti-Szigeti, Katalin; Nusser, Zoltan

    2016-01-01

    Hippocampal pyramidal cells (PCs) express many GABAAR subunit types and receive GABAergic inputs from distinct interneurons. Previous experiments revealed input-specific differences in α1 and α2 subunit densities in perisomatic synapses, suggesting distinct IPSC decay kinetics. However, IPSC decays evoked by axo-axonic, parvalbumin- or cholecystokinin-expressing basket cells were found to be similar. Using replica immunogold labeling, here we show that all CA1 PC somatic and AIS synapses contain the α1, α2, β1, β2, β3 and γ2 subunits. In CA3 PCs, 90% of the perisomatic synapses are immunopositive for the α1 subunit and all synapses are positive for the remaining five subunits. Somatic synapses form unimodal distributions based on their immunoreactivity for these subunits. The α2 subunit densities in somatic synapses facing Cav2.1 (i.e. parvalbumin) or Cav2.2 (cholecystokinin) positive presynaptic active zones are comparable. We conclude that perisomatic synapses made by three distinct interneuron types have similar GABAA receptor subunit content. DOI: http://dx.doi.org/10.7554/eLife.18426.001 PMID:27537197

  16. Distinct transcriptome profiles reveal gene expression patterns during fruit development and maturation in five main cultivated species of pear (Pyrus L.).

    PubMed

    Zhang, Ming-Yue; Xue, Cheng; Xu, Linlin; Sun, Honghe; Qin, Meng-Fan; Zhang, Shaoling; Wu, Jun

    2016-01-01

    The transcriptomes of five pear cultivars, 'Hosui' (P. pyrifolia), 'Yali' (P. bretschneideri), 'Kuerlexiangli' (P. sinkiangensis), 'Nanguoli' (P. ussuriensis), and 'Starkrimson' (P. communis) were sequenced at seven key fruit developmental stages, from fruit setting to maturation and fruit senescence after harvesting. In total, 33,136 genes that could be mapped by reads, were analyzed. Most gene expression cluster models showed a steadily decreasing trend. Gene expression patterns had obvious differences according to maturity type, that is, post-ripening cultivars were still vigorous at maturity, and showed a higher proportion of up-regulated genes; non post-ripening cultivars had a gradually decreasing tendency during fruit maturation. Meanwhile, differentially expressed genes related to fruit quality and development, such as stone cells, sugar, acid and hormones, were identified. Co-expression analysis revealed that several ethylene synthesis genes and polyphenoloxidase-related genes interacted with each other directly, and an indirect relationship was reflected between ethylene synthesis genes and ethylene response genes. In addition, the highly diverse SNPs represented the great differences between oriental and occidental pears. Understanding how RNA-seq based gene-expression patterns and differential gene expression contribute to fruit quality allows us to build models for gene-expression for fruit development of Pyrus species. PMID:27305967

  17. Distinct transcriptome profiles reveal gene expression patterns during fruit development and maturation in five main cultivated species of pear (Pyrus L.)

    PubMed Central

    Zhang, Ming-Yue; Xue, Cheng; Xu, Linlin; Sun, Honghe; Qin, Meng-Fan; Zhang, Shaoling; Wu, Jun

    2016-01-01

    The transcriptomes of five pear cultivars, ‘Hosui’ (P. pyrifolia), ‘Yali’ (P. bretschneideri), ‘Kuerlexiangli’ (P. sinkiangensis), ‘Nanguoli’ (P. ussuriensis), and ‘Starkrimson’ (P. communis) were sequenced at seven key fruit developmental stages, from fruit setting to maturation and fruit senescence after harvesting. In total, 33,136 genes that could be mapped by reads, were analyzed. Most gene expression cluster models showed a steadily decreasing trend. Gene expression patterns had obvious differences according to maturity type, that is, post-ripening cultivars were still vigorous at maturity, and showed a higher proportion of up-regulated genes; non post-ripening cultivars had a gradually decreasing tendency during fruit maturation. Meanwhile, differentially expressed genes related to fruit quality and development, such as stone cells, sugar, acid and hormones, were identified. Co-expression analysis revealed that several ethylene synthesis genes and polyphenoloxidase-related genes interacted with each other directly, and an indirect relationship was reflected between ethylene synthesis genes and ethylene response genes. In addition, the highly diverse SNPs represented the great differences between oriental and occidental pears. Understanding how RNA-seq based gene-expression patterns and differential gene expression contribute to fruit quality allows us to build models for gene-expression for fruit development of Pyrus species. PMID:27305967

  18. A study of protein-carotenoid interactions in the astaxanthin-protein crustacyanin by absorption and Stark spectroscopy; evidence for the presence of three spectrally distinct species.

    PubMed

    Krawczyk, S; Britton, G

    2001-01-12

    Molecular mechanisms underlying the peculiar spectral properties of the carotenoid astaxanthin in alpha-crustacyanin, the blue carotenoprotein isolated from the exoskeleton of the lobster Homarus gammarus, were investigated by comparing the basic electrooptical parameters of astaxanthin free in vitro with those of astaxanthin in the complex. Absorption and electroabsorption (Stark effect) spectra were obtained for alpha-crustacyanin in low-temperature glasses to provide information about the molecular interactions that lead to the large bathochromic shift of the spectra resulting from this complexation. The low-temperature spectra reveal the presence of at least three spectral forms of alpha-crustacyanin, with vibronic (0-0) transitions at 14000 cm(-1), 13500 cm(-1) and 11600 cm(-1) (corresponding to approximately 630, 660 and 780 nm, respectively, at room temperature) and with relative aboundance 85%, 10% and 5%. The longer wavelength absorbing species have not previously been detected. The changes in polarizability and in permanent dipole moments associated with the S0-->S2 electronic transition for all these forms are about 1.5 times larger than for isolated astaxanthin. The results are discussed with reference to the symmetric polarization model for astaxanthin in alpha-crustacyanin. PMID:11341939

  19. Distinct transcriptome profiles reveal gene expression patterns during fruit development and maturation in five main cultivated species of pear (Pyrus L.).

    PubMed

    Zhang, Ming-Yue; Xue, Cheng; Xu, Linlin; Sun, Honghe; Qin, Meng-Fan; Zhang, Shaoling; Wu, Jun

    2016-01-01

    The transcriptomes of five pear cultivars, 'Hosui' (P. pyrifolia), 'Yali' (P. bretschneideri), 'Kuerlexiangli' (P. sinkiangensis), 'Nanguoli' (P. ussuriensis), and 'Starkrimson' (P. communis) were sequenced at seven key fruit developmental stages, from fruit setting to maturation and fruit senescence after harvesting. In total, 33,136 genes that could be mapped by reads, were analyzed. Most gene expression cluster models showed a steadily decreasing trend. Gene expression patterns had obvious differences according to maturity type, that is, post-ripening cultivars were still vigorous at maturity, and showed a higher proportion of up-regulated genes; non post-ripening cultivars had a gradually decreasing tendency during fruit maturation. Meanwhile, differentially expressed genes related to fruit quality and development, such as stone cells, sugar, acid and hormones, were identified. Co-expression analysis revealed that several ethylene synthesis genes and polyphenoloxidase-related genes interacted with each other directly, and an indirect relationship was reflected between ethylene synthesis genes and ethylene response genes. In addition, the highly diverse SNPs represented the great differences between oriental and occidental pears. Understanding how RNA-seq based gene-expression patterns and differential gene expression contribute to fruit quality allows us to build models for gene-expression for fruit development of Pyrus species.

  20. Intracellular dissociation and reassembly of prolyl 4-hydroxylase:the alpha-subunits associated with the immunoglobulin-heavy-chain binding protein (BiP) allowing reassembly with the beta-subunit.

    PubMed Central

    John, D C; Bulleid, N J

    1996-01-01

    Prolyl 4-hydroxylase (P4-H) consists of two distinct polypeptides; the catalytically more important alpha-subunit and the beta-subunit, which is identical to the multifunctional enzyme protein disulphide isomerase. The enzyme appears to be assembled in vivo into an alpha 2 beta 2 tetramer from newly synthesized alpha-subunits associating with an endogenous pool of beta-subunits. Using a cell-free system, we have shown previously that enzyme assembly is redox-dependent and that assembled alpha-subunits are intramolecularly disulphide-bonded [John and Bulleid (1994) Biochemistry 33, 14018-14025]. Here we have studied this assembly process within intact cells by expressing both subunits in COS-1 cells. Newly synthesized alpha-subunits were shown to assemble with the beta-subunit, to form insoluble aggregates, or to remain soluble but not associate with the beta-subunit. Treatment of cells with dithiothreitol (DTT) led to dissociation of P4-H into subunits and on removal of DTT the enzyme reassembled. This reassembly was ATP-dependent, suggesting an interaction with an ATP-dependent chaperone. This was confirmed when immunoglobulin-heavy-chain binding protein (BiP) and alpha-subunits were co-immunoprecipitated with antibodies against the alpha-subunit and BiP, respectively. These results indicate that unassembled alpha-subunits are maintained in an assembly-competent form by interacting with the molecular chaperone BiP. PMID:8760347

  1. High ploidy diversity and distinct patterns of cytotype distribution in a widespread species of Oxalis in the Greater Cape Floristic Region

    PubMed Central

    Krejčíková, Jana; Sudová, Radka; Lučanová, Magdalena; Trávníček, Pavel; Urfus, Tomáš; Vít, Petr; Weiss-Schneeweiss, Hanna; Kolano, Bożena; Oberlander, Kenneth; Dreyer, Leanne L.; Suda, Jan

    2013-01-01

    Background and Aims Genome duplication is widely acknowledged as a major force in the evolution of angiosperms, although the incidence of polyploidy in different floras may differ dramatically. The Greater Cape Floristic Region of southern Africa is one of the world's biodiversity hotspots and is considered depauperate in polyploids. To test this assumption, ploidy variation was assessed in a widespread member of the largest geophytic genus in the Cape flora: Oxalis obtusa. Methods DNA flow cytometry complemented by confirmatory chromosome counts was used to determine ploidy levels in 355 populations of O. obtusa (1014 individuals) across its entire distribution range. Ecological differentiation among cytotypes was tested by comparing sets of vegetation and climatic variables extracted for each locality. Key Results Three majority (2x, 4x, 6x) and three minority (3x, 5x, 8x) cytotypes were detected in situ, in addition to a heptaploid individual originating from a botanical garden. While single-cytotype populations predominate, 12 mixed-ploidy populations were also found. The overall pattern of ploidy level distribution is quite complex, but some ecological segregation was observed. Hexaploids are the most common cytotype and prevail in the Fynbos biome. In contrast, tetraploids dominate in the Succulent Karoo biome. Precipitation parameters were identified as the most important climatic variables associated with cytotype distribution. Conclusions Although it would be premature to make generalizations regarding the role of genome duplication in the genesis of hyperdiversity of the Cape flora, the substantial and unexpected ploidy diversity in Oxalis obtusa is unparalleled in comparison with any other cytologically known native Cape plant species. The results suggest that ploidy variation in the Greater Cape Floristic Region may be much greater than currently assumed, which, given the documented role of polyploidy in speciation, has direct implications for radiation

  2. Interaction of factor XIII subunits.

    PubMed

    Katona, Eva; Pénzes, Krisztina; Csapó, Andrea; Fazakas, Ferenc; Udvardy, Miklós L; Bagoly, Zsuzsa; Orosz, Zsuzsanna Z; Muszbek, László

    2014-03-13

    Coagulation factor XIII (FXIII) is a heterotetramer consisting of 2 catalytic A subunits (FXIII-A2) and 2 protective/inhibitory B subunits (FXIII-B2). FXIII-B, a mosaic protein consisting of 10 sushi domains, significantly prolongs the lifespan of catalytic subunits in the circulation and prevents their slow progressive activation in plasmatic conditions. In this study, the biochemistry of the interaction between the 2 FXIII subunits was investigated. Using a surface plasmon resonance technique and an enzyme-linked immunosorbent assay-type binding assay, the equilibrium dissociation constant (Kd) for the interaction was established in the range of 10(-10) M. Based on the measured Kd, it was calculated that in plasma approximately 1% of FXIII-A2 should be in free form. This value was confirmed experimentally by measuring FXIII-A2 in plasma samples immunodepleted of FXIII-A2B2. Free plasma FXIII-A2 is functionally active, and when activated by thrombin and Ca(2+), it can cross-link fibrin. In cerebrospinal fluid and tears with much lower FXIII subunit concentrations, >80% of FXIII-A2 existed in free form. A monoclonal anti-FXIII-B antibody that prevented the interaction between the 2 subunits reacted with the recombinant combined first and second sushi domains of FXIII-B, and its epitope was localized to the peptide spanning positions 96 to 103 in the second sushi domain. PMID:24408323

  3. Fluorescence measurements reveal stoichiometry of K+ channels formed by modulatory and delayed rectifier alpha-subunits.

    PubMed

    Kerschensteiner, Daniel; Soto, Florentina; Stocker, Martin

    2005-04-26

    Modulatory alpha-subunits, which comprise one-fourth of all voltagegated K(+) channel (Kv) alpha-subunits, do not assemble into homomeric channels, but selectively associate with delayed rectifier Kv2 subunits to form heteromeric channels of unknown stoichiometry. Their distinct expression patterns and unique functional properties have made these channels candidate molecular correlates for a broad set of native K(+) currents. Here, we combine FRET and electrophysiological measurements to determine the stoichiometry and geometry of heteromeric channels composed of the delayed rectifier Kv2.1 subunit and the modulatory Kv9.3 alpha-subunit. Kv channel alpha-subunits were fused with GFP variants, and heteromerization of different combinations of tagged and untagged alpha-subunits was studied. FRET, evaluated by acceptor photobleaching, was only observed upon formation of functional channels. Our results, obtained from two independent experimental paradigms, suggest the formation of heteromeric Kv2.1/Kv9.3 channels of fixed stoichiometry consisting of three Kv2.1 subunits and one Kv9.3 subunit. Strikingly, despite this uneven stoichiometry, we find that heteromeric Kv2.1/Kv9.3 channels maintain a pseudosymmetric arrangement of subunits around the central pore. PMID:15827117

  4. Fluorescence measurements reveal stoichiometry of K+ channels formed by modulatory and delayed rectifier α-subunits

    PubMed Central

    Kerschensteiner, Daniel; Soto, Florentina; Stocker, Martin

    2005-01-01

    Modulatory α-subunits, which comprise one-fourth of all voltagegated K+ channel (Kv) α-subunits, do not assemble into homomeric channels, but selectively associate with delayed rectifier Kv2 subunits to form heteromeric channels of unknown stoichiometry. Their distinct expression patterns and unique functional properties have made these channels candidate molecular correlates for a broad set of native K+ currents. Here, we combine FRET and electrophysiological measurements to determine the stoichiometry and geometry of heteromeric channels composed of the delayed rectifier Kv2.1 subunit and the modulatory Kv9.3 α-subunit. Kv channel α-subunits were fused with GFP variants, and heteromerization of different combinations of tagged and untagged α-subunits was studied. FRET, evaluated by acceptor photobleaching, was only observed upon formation of functional channels. Our results, obtained from two independent experimental paradigms, suggest the formation of heteromeric Kv2.1/Kv9.3 channels of fixed stoichiometry consisting of three Kv2.1 subunits and one Kv9.3 subunit. Strikingly, despite this uneven stoichiometry, we find that heteromeric Kv2.1/Kv9.3 channels maintain a pseudosymmetric arrangement of subunits around the central pore. PMID:15827117

  5. Chromosomal localization of human RNA polymerase II subunit genes

    SciTech Connect

    Acker, J.; Wintzerith, M.; Vigneron, M.; Kedinger, C. ); Mattei, M.G.; Roeckel, N.; Depetris, D. )

    1994-04-01

    The eukaryotic DNA-dependent RNA polymerase II (or B) is composed of 10 to 14 polypeptides ranging from 220 to 10 kDa. To gain further insight into the molecular structure and function of these subunits, the authors have undertaken the molecular cloning of nucleotide sequences corresponding to the human enzyme. The cDNAs of five subunits (hRPB220, hRPB140, hRPB33, hRPB25, and hRPB14.5) have been isolated. Using in situ hybridization, they show that the genes of these subunits have distinct chromosomal locations (17p13, 4q12, 16q13-q21, 19p13.3, and 19q12, respectively). Thus, if assembly of active polymerase molecules requires coordinated expression from these independent genes, mechanisms that ensure tight coregulation of the corresponding promoters must exist. 20 refs., 2 figs., 1 tab.

  6. Zinc induces distinct changes in the metabolism of reactive oxygen and nitrogen species (ROS and RNS) in the roots of two Brassica species with different sensitivity to zinc stress

    PubMed Central

    Feigl, Gábor; Lehotai, Nóra; Molnár, Árpád; Ördög, Attila; Rodríguez-Ruiz, Marta; Palma, José M.; Corpas, Francisco J.; Erdei, László; Kolbert, Zsuzsanna

    2015-01-01

    Background and Aims Zinc (Zn) is an essential micronutrient naturally present in soils, but anthropogenic activities can lead to accumulation in the environment and resulting damage to plants. Heavy metals such as Zn can induce oxidative stress and the generation of reactive oxygen and nitrogen species (ROS and RNS), which can reduce growth and yield in crop plants. This study assesses the interplay of these two families of molecules in order to evaluate the responses in roots of two Brassica species under high concentrations of Zn. Methods Nine-day-old hydroponically grown Brassica juncea (Indian mustard) and B. napus (oilseed rape) seedlings were treated with ZnSO4 (0, 50, 150 and 300 µm) for 7 d. Stress intensity was assessed through analyses of cell wall damage and cell viability. Biochemical and cellular techniques were used to measure key components of the metabolism of ROS and RNS including lipid peroxidation, enzymatic antioxidants, protein nitration and content of superoxide radical (O2·−), nitric oxide (NO) and peroxynitrite (ONOO−). Key Results Analysis of morphological root damage and alterations of microelement homeostasis indicate that B. juncea is more tolerant to Zn stress than B. napus. ROS and RNS parameters suggest that the oxidative components are predominant compared with the nitrosative components in the root system of both species. Conclusions The results indicate a clear relationship between ROS and RNS metabolism as a mechanism of response against stress caused by an excess of Zn. The oxidative stress components seem to be more dominant than the elements of the nitrosative stress in the root system of these two Brassica species. PMID:25538112

  7. Molecular and morphological evidence supports the species status of the Mahachai fighter Betta sp. Mahachai and reveals new species of Betta from Thailand.

    PubMed

    Sriwattanarothai, N; Steinke, D; Ruenwongsa, P; Hanner, R; Panijpan, B

    2010-08-01

    Two regions of mitochondrial (mt) DNA, cytochrome c oxidase subunit 1 (COI) and 16S rRNA, were sequenced in nine species of Betta from Thailand and Indonesia. Most species showed little intraspecific COI variation (adjusted mean = 0.48%) including the putative species Betta sp. Mahachai, but one species (Betta smaragdina) included three lineages showing much greater divergence (7.03-13.48%) that probably represent overlooked species. These findings were confirmed by maximum likelihood analysis and Bayesian inference, which revealed well-supported corresponding monophyletic clades. Based on these results and morphological differences, the putative species Betta sp. Mahachai from central Thailand is a species distinct from other members of the B. splendens group and represents a new and hitherto undescribed species. Furthermore, this study also demonstrated the probable existence of two overlooked Betta species found in the Khorat plateau basin, illustrating the utility of mitochondrial genetic markers in the revelation of overlooked diversity.

  8. Molecular and morphological evidence supports the species status of the Mahachai fighter Betta sp. Mahachai and reveals new species of Betta from Thailand.

    PubMed

    Sriwattanarothai, N; Steinke, D; Ruenwongsa, P; Hanner, R; Panijpan, B

    2010-08-01

    Two regions of mitochondrial (mt) DNA, cytochrome c oxidase subunit 1 (COI) and 16S rRNA, were sequenced in nine species of Betta from Thailand and Indonesia. Most species showed little intraspecific COI variation (adjusted mean = 0.48%) including the putative species Betta sp. Mahachai, but one species (Betta smaragdina) included three lineages showing much greater divergence (7.03-13.48%) that probably represent overlooked species. These findings were confirmed by maximum likelihood analysis and Bayesian inference, which revealed well-supported corresponding monophyletic clades. Based on these results and morphological differences, the putative species Betta sp. Mahachai from central Thailand is a species distinct from other members of the B. splendens group and represents a new and hitherto undescribed species. Furthermore, this study also demonstrated the probable existence of two overlooked Betta species found in the Khorat plateau basin, illustrating the utility of mitochondrial genetic markers in the revelation of overlooked diversity. PMID:20646165

  9. Sequence analysis of four acidic beta-crystallin subunits of amphibian lenses: phylogenetic comparison between beta- and gamma-crystallins.

    PubMed

    Lu, S F; Pan, F M; Chiou, S H

    1996-04-16

    beta-Crystallins composed of the most heterogeneous group of subunit chains among the three major crystallin families of vertebrates, i.e. alpha-, beta- and gamma-crystallins, are less well understood at the structural and functional levels than the other two. They comprise a multigene family with at least three basic (betaB1-3) and four acidic (betaA1-4) subunit polypeptides. In order to facilitate the determination of the primary sequences of all these ubiquitous crystallin subunits present in all vertebrate species, cDNA mixture was synthesized from the poly(A)+ mRNA isolated from bullfrog eye lenses. We report here a protocol of Rapid Amplification of cDNA Ends (RACE) was used to amplify cDNAs encoding beta-crystallin acidic subunit polypeptides by polymerase chain reaction (PCR). Four complete full-length reading frames with two each of 597 and 648 base pairs, which cover four deduced protein sequences of 198 (betaA1-1 and betaA1-2) and 215 (betaA3-1 and betaA3-2) amino acids including the universal initiating methionine, were revealed by nucleotide sequencing. They show about 96-98% sequence similarity among themselves and 76-80%, 80-83% to the homologous betaA1/A3 crystallins of bovine and human species respectively, revealing the close structural relationship among acidic subunits of all beta-crystallins even from remotely related species. In this study a phylogenetic comparison based on amino-acid sequences of various betaA1/A3 crystallins plus the major basic beta-crystallin (betaBp) and gamma-crystallin from different vertebrate species is made using a combination of distance matrix and approximate parsimony methods, which correctly groups these betaA crystallin chains together as one family distinct from basic beta-crystallins and gamma-crystallin and further corroborates the supposition that beta- and gamma-crystallins form a superfamily with a common ancestry.

  10. Is Kir6.1 a subunit of mitoK{sub ATP}?

    SciTech Connect

    Brian Foster, D.; Rucker, Jasma J.; Marban, Eduardo

    2008-02-15

    The subunit composition of the mitochondrial ATP-sensitive K{sup +}-channel (mitoK{sub ATP}) is unknown, though some suspect a role for the inward rectifier, Kir6.1, based largely on antibody studies of heart mitochondria. To ascertain the molecular identity of mitoK{sub ATP} we therefore sought to purify this putative mitochondrial Kir6.1, and conclusively identify the subunits by mass spectrometry. Immunoblots, conducted with two commercially available antibodies, revealed two distinct signals in isolated heart mitochondria, of 51 and 48 kDa, respectively. Localization was confirmed by either immuno-gold electron microscopy or by immunofluorescence. Each putative Kir6.1 species was extracted, purified, and identified by LC-MS/MS. The 51 kDa band was identified as NADH-dehydrogenase flavoprotein 1, while the preponderant protein in the 48-kDa band was mitochondrial isocitrate dehydrogenase (NADP form). 1D-, 2D-, and native gel analyses were consistent with these assignments. The data suggest it is premature to assign Kir6.1 a role in mitoK{sub ATP} on the basis of immunoreactivity alone.

  11. The land crabs of the Discoplax longipes A. Milne-Edwards, 1867 species group, with description of a new species from Guam (Crustacea: Decapoda: Brachyura: Gecarcinidae).

    PubMed

    Ng, Peter K L; Shih, Hsi-Te

    2015-06-30

    Specimens of the gecarcinid land crab Discoplax longipes A. Milne-Edwards, 1867, from the western Pacific, can be separated into two distinct groups on the basis of DNA (mitochondrial 16S rDNA and cytochrome oxidase subunit I) and structure of the male first gonopod. On the basis of this data, the material that occurs from the Loyalty Islands to French Polynesia is shown to be D. longipes s. str., whereas specimens from Guam are here referred to a new pseudocryptic species, D. michalis n. sp. The two species are described and figured; and a revised key to the long-legged Discoplax species is provided.

  12. The ribosomal subunit assembly line

    PubMed Central

    Dlakić, Mensur

    2005-01-01

    Recent proteomic studies in Saccharomyces cerevisiae have identified nearly 200 proteins, other than the structural ribosomal proteins, that participate in the assembly of ribosomal subunits and their transport from the nucleus. In a separate line of research, proteomic studies of mature plant ribosomes have revealed considerable variability in the protein composition of individual ribosomes. PMID:16207363

  13. Chemical crosslinking of the subunits of HIV-1 reverse transcriptase.

    PubMed Central

    Debyser, Z.; De Clercq, E.

    1996-01-01

    The reverse transcriptase (RT) of the human immunodeficiency virus type 1 (HIV-1) is composed of two subunits of 66 and 51 kDa in a 1 to 1 ratio. Because dimerization is a prerequisite for enzymatic activity, interference with the dimerization process could constitute an alternative antiviral strategy for RT inhibition. Here we describe an in vitro assay for the study of the dimerization state of HIV-1 reverse transcriptase based on chemical crosslinking of the subunits with dimethylsuberimidate. Crosslinking results in the formation of covalent bonds between the subunits, so that the crosslinked species can be resolved by denaturing gel electrophoresis. Crosslinked RT species with molecular weight greater than that of the dimeric form accumulate during a 1-15-min time course. Initial evidence suggests that those high molecular weight species represent trimers and tetramers and may be the result of intramolecular crosslinking of the subunits of a higher-order RT oligomer. A peptide that corresponds to part of the tryptophan repeat motif in the connection domain of HIV-1 RT inhibits crosslink formation as well as enzymatic activity. The crosslinking assay thus allows the investigation of the effect of inhibitors on the dimerization of HIV-1 RT. PMID:8745406

  14. Balancing acts of two HEAT subunits of condensin I support dynamic assembly of chromosome axes.

    PubMed

    Kinoshita, Kazuhisa; Kobayashi, Tetsuya J; Hirano, Tatsuya

    2015-04-01

    Condensin I is a five-subunit protein complex that plays a central role in mitotic chromosome assembly and segregation in eukaryotes. To dissect its mechanism of action, we reconstituted wild-type and mutant complexes from recombinant subunits and tested their abilities to assemble chromosomes in Xenopus egg cell-free extracts depleted of endogenous condensins. We find that ATP binding and hydrolysis by SMC subunits have distinct contributions to the action of condensin I and that continuous ATP hydrolysis is required for structural maintenance of chromosomes. Mutant complexes lacking either one of two HEAT subunits produce abnormal chromosomes with highly characteristic defects and have contrasting structural effects on chromosome axes preassembled with the wild-type complex. We propose that balancing acts of the two HEAT subunits support dynamic assembly of chromosome axes under the control of the SMC ATPase cycle, thereby governing construction of rod-shaped chromosomes in eukaryotic cells.

  15. The Evolution of the Four Subunits of Voltage-Gated Calcium Channels: Ancient Roots, Increasing Complexity, and Multiple Losses

    PubMed Central

    Moran, Yehu; Zakon, Harold H.

    2014-01-01

    The alpha subunits of voltage-gated calcium channels (Cavs) are large transmembrane proteins responsible for crucial physiological processes in excitable cells. They are assisted by three auxiliary subunits that can modulate their electrical behavior. Little is known about the evolution and roles of the various subunits of Cavs in nonbilaterian animals and in nonanimal lineages. For this reason, we mapped the phyletic distribution of the four channel subunits and reconstructed their phylogeny. Although alpha subunits have deep evolutionary roots as ancient as the split between plants and opistokonths, beta subunits appeared in the last common ancestor of animals and their close-relatives choanoflagellates, gamma subunits are a bilaterian novelty and alpha2/delta subunits appeared in the lineage of Placozoa, Cnidaria, and Bilateria. We note that gene losses were extremely common in the evolution of Cavs, with noticeable losses in multiple clades of subfamilies and also of whole Cav families. As in vertebrates, but not protostomes, Cav channel genes duplicated in Cnidaria. We characterized by in situ hybridization the tissue distribution of alpha subunits in the sea anemone Nematostella vectensis, a nonbilaterian animal possessing all three Cav subfamilies common to Bilateria. We find that some of the alpha subunit subtypes exhibit distinct spatiotemporal expression patterns. Further, all six sea anemone alpha subunit subtypes are conserved in stony corals, which separated from anemones 500 MA. This unexpected conservation together with the expression patterns strongly supports the notion that these subtypes carry unique functional roles. PMID:25146647

  16. Talaromyces rubrifaciens, a new species discovered from heating, ventilation and air conditioning systems in China.

    PubMed

    Luo, Yi; Lu, Xiaohong; Bi, Wu; Liu, Fan; Gao, Weiwei

    2016-01-01

    A new Talaromyces species, T. rubrifaciens, was isolated from supply air outlets of heating, ventilation and air conditioning (HVAC) systems in three kinds of public building in Beijing and Nanjing, China. Morphologically it exhibits many characters of section Trachyspermi but is distinguished from other species of this section by restricted growth and broad and strictly biverticillate conidiophores. Phylogenetic analyses based on the internal transcribed spacer rDNA (ITS), β-tubulin (BenA), calmodulin (CaM) and RNA polymerase second largest subunit (RPB2) genes reveal that T. rubrifaciens is a distinct species in section Trachyspermi. PMID:27055570

  17. Talaromyces rubrifaciens, a new species discovered from heating, ventilation and air conditioning systems in China.

    PubMed

    Luo, Yi; Lu, Xiaohong; Bi, Wu; Liu, Fan; Gao, Weiwei

    2016-01-01

    A new Talaromyces species, T. rubrifaciens, was isolated from supply air outlets of heating, ventilation and air conditioning (HVAC) systems in three kinds of public building in Beijing and Nanjing, China. Morphologically it exhibits many characters of section Trachyspermi but is distinguished from other species of this section by restricted growth and broad and strictly biverticillate conidiophores. Phylogenetic analyses based on the internal transcribed spacer rDNA (ITS), β-tubulin (BenA), calmodulin (CaM) and RNA polymerase second largest subunit (RPB2) genes reveal that T. rubrifaciens is a distinct species in section Trachyspermi.

  18. Emergence of ion channel modal gating from independent subunit kinetics.

    PubMed

    Bicknell, Brendan A; Goodhill, Geoffrey J

    2016-09-01

    Many ion channels exhibit a slow stochastic switching between distinct modes of gating activity. This feature of channel behavior has pronounced implications for the dynamics of ionic currents and the signaling pathways that they regulate. A canonical example is the inositol 1,4,5-trisphosphate receptor (IP3R) channel, whose regulation of intracellular Ca(2+) concentration is essential for numerous cellular processes. However, the underlying biophysical mechanisms that give rise to modal gating in this and most other channels remain unknown. Although ion channels are composed of protein subunits, previous mathematical models of modal gating are coarse grained at the level of whole-channel states, limiting further dialogue between theory and experiment. Here we propose an origin for modal gating, by modeling the kinetics of ligand binding and conformational change in the IP3R at the subunit level. We find good agreement with experimental data over a wide range of ligand concentrations, accounting for equilibrium channel properties, transient responses to changing ligand conditions, and modal gating statistics. We show how this can be understood within a simple analytical framework and confirm our results with stochastic simulations. The model assumes that channel subunits are independent, demonstrating that cooperative binding or concerted conformational changes are not required for modal gating. Moreover, the model embodies a generally applicable principle: If a timescale separation exists in the kinetics of individual subunits, then modal gating can arise as an emergent property of channel behavior. PMID:27551100

  19. Subunit asymmetry in the three-dimensional structure of a human CuZnSOD mutant found in familial amyotrophic lateral sclerosis.

    PubMed Central

    Hart, P. J.; Liu, H.; Pellegrini, M.; Nersissian, A. M.; Gralla, E. B.; Valentine, J. S.; Eisenberg, D.

    1998-01-01

    The X-ray crystal structure of a human copper/zinc superoxide dismutase mutant (G37R CuZnSOD) found in some patients with the inherited form of Lou Gehrig's disease (FALS) has been determined to 1.9 angstroms resolution. The two SOD subunits have distinct environments in the crystal and are different in structure at their copper binding sites. One subunit (subunit[intact]) shows a four-coordinate ligand geometry of the copper ion, whereas the other subunit (subunit[broken]) shows a three-coordinate geometry of the copper ion. Also, subunit(intact) displays higher atomic displacement parameters for backbone atoms ((B) = 30 +/- 10 angstroms2) than subunit(broken) ((B) = 24 +/- 11 angstroms2). This structure is the first CuZnSOD to show large differences between the two subunits. Factors that may contribute to these differences are discussed and a possible link of a looser structure to FALS is suggested. PMID:9541385

  20. Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae) revealed by COI and 16S DNA sequences.

    PubMed

    Lim, Phaik-Eem; Tan, Ji; Suana, I Wayan; Eamsobhana, Praphathip; Yong, Hoi Sen

    2012-01-01

    The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies.

  1. Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae) revealed by COI and 16S DNA sequences.

    PubMed

    Lim, Phaik-Eem; Tan, Ji; Suana, I Wayan; Eamsobhana, Praphathip; Yong, Hoi Sen

    2012-01-01

    The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies. PMID:22615962

  2. Distinct Genetic Lineages of Bactrocera caudata (Insecta: Tephritidae) Revealed by COI and 16S DNA Sequences

    PubMed Central

    Lim, Phaik-Eem; Tan, Ji; Suana, I. Wayan; Eamsobhana, Praphathip; Yong, Hoi Sen

    2012-01-01

    The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected ‘p’ distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The ‘p’ values are distinctly different from intraspecific ‘p’ distance (0–0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus – B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies. PMID:22615962

  3. Production and characterization of monoclonal antibodies to the subunits of human phosphofructokinase: new tools for the immunochemical and genetic analyses of isozymes.

    PubMed

    Vora, S; Wims, L A; Durham, S; Morrison, S L

    1981-10-01

    Recently we have demonstrated that human phosphofructokinase (PFK; ATP: D-fructose-6-P, 1-phosphotransferase; EC.2.7.1.11) is under the control of three structural loci that code for M (muscle-type), L (liver-type), and P (platelet-type) subunits: random tetramerization of these subunits produces various isozymes. In this study, we have produced and characterized BALB/c hybridoma antibodies to the M- and L-type subunits of human PFK. The specific antibodies were detected by an enzyme-immunoprecipitation assay using Staphylococci-bearing protein A as an immunoadsorbent. Of the wells tested using red blood cell (RBC) PFK (M + L), 61% were positive. Only one M-specific hybridoma was identified. The one anti-M and 4 anti-L antibodies were characterized for their biochemical and immunochemical specificities. To define the combining specificities of these antibodies, we compared their reactivity and that of monospecific rabbit anti-M antiserum with muscle and liver PFKs from 15 different vertebrate species. The rabbit anti-M shows strong cross-reactivity with the muscle PFKs from all the species studied. In contrast, the monoclonal anti-M reacts exclusively with muscle PFKs from primates. Two of four anti-L antibodies react only with human L-PFK, whereas the other two react with that from a few other vertebrate species as well. Taken together, these data suggest that primate-specific antibodies recognize evolutionarily, recently acquired antigenic determinants, whereas the antibodies reactive with PFKs from distantly related species recognize conserved determinants. The differential immunoreactivities of muscle and liver PFKs strongly suggest the presence of distinct isozymes in all the vertebrate species studied. These studies demonstrate that it is feasible to produce and characterize monoclonal antibodies that distinguish among isozymes with structural and functional similarities. These antibodies provide sensitive tools in the analyses of isozyme structure, genetics

  4. Subunit gamma of the oxaloacetate decarboxylase Na(+) pump: interaction with other subunits/domains of the complex and binding site for the Zn(2+) metal ion.

    PubMed

    Schmid, Markus; Wild, Markus R; Dahinden, Pius; Dimroth, Peter

    2002-01-29

    The oxaloacetate decarboxylase Na(+) pump of Klebsiella pneumoniae is an enzyme complex composed of the peripheral alpha subunit and the two integral membrane-bound subunits beta and gamma. The alpha subunit consists of the N-terminal carboxyltransferase domain and the C-terminal biotin domain, which are connected by a flexible proline/alanine-rich linker peptide. To probe interactions between the two domains of the alpha subunit and between alpha-subunit domains and the gamma subunit, the relevant polypeptides were synthesized in Escherichia coli and subjected to copurification studies. The two alpha-subunit domains had no distinct affinity toward each other and could, therefore, not be purified as a unit on avidin-sepharose. The two domains reacted together catalytically, however, performing the carboxyl transfer from oxaloacetate to protein-bound biotin. This reaction was enhanced up to 6-fold in the presence of the Zn(2+)-containing gamma subunit. On the basis of copurification with different tagged proteins, the C-terminal biotin domain but not the N-terminal carboxyltransferase domain of the alpha subunit formed a strong complex with the gamma subunit. Upon the mutation of gamma H78 to alanine, the binding affinity to subunit alpha was lost, indicating that this amino acid may be essential for formation of the oxaloacetate decarboxylase enzyme complex. The binding residues for the Zn(2+) metal ion were identified by site-directed and deletion mutagenesis. In the gamma D62A or gamma H77A mutant, the Zn(2+) content of the decarboxylase decreased to 35% or 10% of the wild-type enzyme, respectively. Less than 5% of the Zn(2+) present in the wild-type enzyme was found if the two C-terminal gamma-subunit residues H82 and P83 were deleted. Corresponding with the reduced Zn(2+) contents in these mutants, the oxaloacetate decarboxylase activities were diminished. These results indicate that aspartate 62, histidine 77, and histidine 82 of the gamma subunit are ligands

  5. Specific Subunits of Heterotrimeric G Proteins Play Important Roles during Nodulation in Soybean1[W][OA

    PubMed Central

    Choudhury, Swarup Roy; Pandey, Sona

    2013-01-01

    Heterotrimeric G proteins comprising Gα, Gβ, and Gγ subunits regulate many fundamental growth and development processes in all eukaryotes. Plants possess a relatively limited number of G-protein components compared with mammalian systems, and their detailed functional characterization has been performed mostly in Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa). However, the presence of single Gα and Gβ proteins in both these species has significantly undermined the complexity and specificity of response regulation in plant G-protein signaling. There is ample pharmacological evidence for the role of G proteins in regulation of legume-specific processes such as nodulation, but the lack of genetic data from a leguminous species has restricted its direct assessment. Our recent identification and characterization of an elaborate G-protein family in soybean (Glycine max) and the availability of appropriate molecular-genetic resources have allowed us to directly evaluate the role of G-protein subunits during nodulation. We demonstrate that all G-protein genes are expressed in nodules and exhibit significant changes in their expression in response to Bradyrhizobium japonicum infection and in representative supernodulating and nonnodulating soybean mutants. RNA interference suppression and overexpression of specific G-protein components results in lower and higher nodule numbers, respectively, validating their roles as positive regulators of nodule formation. Our data further show preferential usage of distinct G-protein subunits in the presence of an additional signal during nodulation. Interestingly, the Gα proteins directly interact with the soybean nodulation factor receptors NFR1α and NFR1β, suggesting that the plant G proteins may couple with receptors other than the canonical heptahelical receptors common in metazoans to modulate signaling. PMID:23569109

  6. Structural characterization of recombinant crustacyanin subunits from the lobster Homarus americanus.

    PubMed

    Ferrari, Michele; Folli, Claudia; Pincolini, Elisa; McClintock, Timothy S; Rössle, Manfred; Berni, Rodolfo; Cianci, Michele

    2012-08-01

    Crustacean crustacyanin proteins are linked to the production and modification of carapace colour, with direct implications for fitness and survival. Here, the structural and functional properties of the two recombinant crustacyanin subunits H(1) and H(2) from the American lobster Homarus americanus are reported. The two subunits are structurally highly similar to the corresponding natural apo crustacyanin CRTC and CRTA subunits from the European lobster H. gammarus. Reconstitution studies of the recombinant crustacyanin proteins H(1) and H(2) with astaxanthin reproduced the bathochromic shift of 85-95 nm typical of the natural crustacyanin subunits from H. gammarus in complex with astaxanthin. Moreover, correlations between the presence of crustacyanin genes in crustacean species and the resulting carapace colours with the spectral properties of the subunits in complex with astaxanthin confirmed this genotype-phenotype linkage. PMID:22869108

  7. Structural characterization of recombinant crustacyanin subunits from the lobster Homarus americanus

    PubMed Central

    Ferrari, Michele; Folli, Claudia; Pincolini, Elisa; McClintock, Timothy S.; Rössle, Manfred; Berni, Rodolfo; Cianci, Michele

    2012-01-01

    Crustacean crustacyanin proteins are linked to the production and modification of carapace colour, with direct implications for fitness and survival. Here, the structural and functional properties of the two recombinant crustacyanin subunits H1 and H2 from the American lobster Homarus americanus are reported. The two subunits are structurally highly similar to the corresponding natural apo crustacyanin CRTC and CRTA subunits from the European lobster H. gammarus. Reconstitution studies of the recombinant crustacyanin proteins H1 and H2 with astaxanthin reproduced the bathochromic shift of 85–95 nm typical of the natural crustacyanin subunits from H. gammarus in complex with astaxanthin. Moreover, correlations between the presence of crustacyanin genes in crustacean species and the resulting carapace colours with the spectral properties of the subunits in complex with astaxanthin confirmed this genotype–phenotype linkage. PMID:22869108

  8. Structural characterization of recombinant crustacyanin subunits from the lobster Homarus americanus.

    PubMed

    Ferrari, Michele; Folli, Claudia; Pincolini, Elisa; McClintock, Timothy S; Rössle, Manfred; Berni, Rodolfo; Cianci, Michele

    2012-08-01

    Crustacean crustacyanin proteins are linked to the production and modification of carapace colour, with direct implications for fitness and survival. Here, the structural and functional properties of the two recombinant crustacyanin subunits H(1) and H(2) from the American lobster Homarus americanus are reported. The two subunits are structurally highly similar to the corresponding natural apo crustacyanin CRTC and CRTA subunits from the European lobster H. gammarus. Reconstitution studies of the recombinant crustacyanin proteins H(1) and H(2) with astaxanthin reproduced the bathochromic shift of 85-95 nm typical of the natural crustacyanin subunits from H. gammarus in complex with astaxanthin. Moreover, correlations between the presence of crustacyanin genes in crustacean species and the resulting carapace colours with the spectral properties of the subunits in complex with astaxanthin confirmed this genotype-phenotype linkage.

  9. Activation of pro-(matrix metalloproteinase-2) (pro-MMP-2) by thrombin is membrane-type-MMP-dependent in human umbilical vein endothelial cells and generates a distinct 63 kDa active species.

    PubMed Central

    Lafleur, M A; Hollenberg, M D; Atkinson, S J; Knäuper, V; Murphy, G; Edwards, D R

    2001-01-01

    Thrombin, a critical enzyme in the coagulation cascade, has also been associated with angiogenesis and activation of the zymogen form of matrix metalloproteinase-2 (MMP-2 or gelatinase-A). We show that thrombin activated pro-MMP-2 in a dose- and time-dependent manner in cultured human umbilical-vein endothelial cells (HUVECs) to generate a catalytically active 63 kDa protein that accumulated as the predominant form in the conditioned medium. This 63 kDa thrombin-activated MMP-2 is distinct from the 62 kDa species found following concanavalin A or PMA stimulated pro-MMP-2 activation. Hirudin and leupeptin blocked thrombin-induced pro-MMP-2 activation, demonstrating that the proteolytic activity of thrombin is essential. However, activation was also dependent upon membrane-type-MMP (MT-MMP) action, since it was blocked by EDTA, o-phenanthroline, hydroxamate metalloproteinase inhibitors, tissue inhibitor of metalloproteinase-2 (TIMP-2) and TIMP-4, but not TIMP-1. Thrombin inefficiently cleaved recombinant 72 kDa pro-MMP-2, but efficiently cleaved the 64 kDa MT-MMP-processed intermediate form in the presence of cells. Thrombin also rapidly (within 1 h) increased cellular MT-MMP activity, and at longer time points (>6 h) it increased expression of MT1-MMP mRNA and protein. Thus signalling via proteinase-activated receptors (PARs) may play a role in thrombin-induced MMP-2 activation, though this does not appear to involve PAR1, PAR2, or PAR4 in HUVECs. These results indicate that in HUVECs the activation of pro-MMP-2 by thrombin involves increased MT-MMP activity and preferential cleavage of the MT-MMP-processed 64 kDa MMP-2 form in the presence of cells. The integration of these proteinase systems in the vascular endothelium may be important during thrombogenesis and tissue remodelling associated with neovascularization. PMID:11415441

  10. Subunit Compositions of the RNA-Silencing Enzymes Pol IV and Pol V Reveal Their Origins as Specialized Forms of RNA Polymerase II

    SciTech Connect

    Ream, Thomas S.; Haag, J. R.; Wierzbicki, A. T.; Nicora, Carrie D.; Norbeck, Angela D.; Zhu, Jian-Kang; Hagen, Gretchen; Guilfoyle, Thomas J.; Pasa-Tolic, Ljiljana; Pikaard, Craig S.

    2009-01-30

    In addition to RNA polymerases I, II, and III, the essential RNA polymerases present in all eukaryotes, plants have two additional nuclear RNA polymerases, abbreviated as Pol IV and Pol V, that play nonredundant roles in siRNA-directed DNA methylation and gene silencing. We show that Arabidopsis Pol IV and Pol V are composed of subunits that are paralogous or identical to the 12 subunits of Pol II. Four subunits of Pol IV are distinct from their Pol II paralogs, six subunits of Pol V are distinct from their Pol II paralogs, and four subunits differ between Pol IV and Pol V. Importantly, the subunit differences occur in key positions relative to the template entry and RNA exit paths. Our findings support the hypothesis that Pol IV and Pol V are Pol II-like enzymes that evolved specialized roles in the production of noncoding transcripts for RNA silencing and genome defense.

  11. Small things considered: the small accessory subunits of RNA polymerase in Gram-positive bacteria

    PubMed Central

    Weiss, Andy; Shaw, Lindsey N.

    2015-01-01

    The DNA-dependent RNA polymerase core enzyme in Gram-positive bacteria consists of seven subunits. Whilst four of them (α2ββ′) are essential, three smaller subunits, δ, ε and ω (∼9–21.5 kDa), are considered accessory. Both δ and ω have been viewed as integral components of RNAP for several decades; however, ε has only recently been described. Functionally these three small subunits carry out a variety of tasks, imparting important, supportive effects on the transcriptional process of Gram-positive bacteria. While ω is thought to have a wide range of roles, reaching from maintaining structural integrity of RNAP to σ factor recruitment, the only suggested function for ε thus far is in protecting cells from phage infection. The third subunit, δ, has been shown to have distinct influences in maintaining transcriptional specificity, and thus has a key role in cellular fitness. Collectively, all three accessory subunits, although dispensable under laboratory conditions, are often thought to be crucial for proper RNAP function. Herein we provide an overview of the available literature on each subunit, summarizing landmark findings that have deepened our understanding of these proteins and their function, and outline future challenges in understanding the role of these small subunits in the transcriptional process. PMID:25878038

  12. Molecular Cloning of the Genes Encoding the PR55/Bβ/δ Regulatory Subunits for PP-2A and Analysis of Their Functions in Regulating Development of Goldfish, Carassius auratus.

    PubMed

    Zhao, Jun-Qiong; Xie, Si-Si; Liu, Wen-Bin; Xiao, Ya-Mei; Zeng, Xiao-Ming; Deng, Mi; Gong, Lili; Liu, Jin-Ping; Chen, Pei-Chao; Zhou, Jie; Hu, Xiao-Hui; Lv, Jia-Han; Yu, Xiang-Qian; Wang, Dao; Li, Chi; Peng, Yun-Lei; Liao, Gao-Peng; Liu, Yun; Li, David Wan-Cheng

    2010-01-01

    The protein phosphatase-2A (PP-2A), one of the major phosphatases in eukaryotes, is a heterotrimer, consisting of a scaffold A subunit, a catalytic C subunit and a regulatory B subunit. Previous studies have shown that besides regulating specific PP-2A activity, various B subunits encoded by more than 16 different genes, may have other functions. To explore the possible roles of the regulatory subunits of PP-2A in vertebrate development, we have cloned the PR55/B family regulatory subunits: β and δ, analyzed their tissue specific and developmental expression patterns in Goldfish ( Carassius auratus). Our results revealed that the full-length cDNA for PR55/Bβ consists of 1940 bp with an open reading frame of 1332 nucleotides coding for a deduced protein of 443 amino acids. The full length PR55/Bδ cDNA is 2163 bp containing an open reading frame of 1347 nucleotides encoding a deduced protein of 448 amino acids. The two isoforms of PR55/B display high levels of sequence identity with their counterparts in other species. The PR55/Bβ mRNA and protein are detected in brain and heart. In contrast, the PR55/Bδ is expressed in all 9 tissues examined at both mRNA and protein levels. During development of goldfish, the mRNAs for PR55/Bβ and PR55/Bδ show distinct patterns. At the protein level, PR55/Bδ is expressed at all developmental stages examined, suggesting its important role in regulating goldfish development. Expression of the PR55/Bδ anti-sense RNA leads to significant downregulation of PR55/Bδ proteins and caused severe abnormality in goldfish trunk and eye development. Together, our results suggested that PR55/Bδ plays an important role in governing normal trunk and eye formation during goldfish development.

  13. Developmental expression of cerebellar GABAA-receptor subunit mRNAs. Nature versus nurture.

    PubMed

    Siegel, R E

    1998-01-01

    Recent studies have demonstrated that many of the mRNAs encoding GABAA-receptor subunits in the cerebellum exhibit distinct temporal profiles of expression. The levels of six of these subunit transcripts increase severalfold in the second week of postnatal ontogeny. Findings from a variety of experimental systems suggest that the onset and increases in subunit mRNA expression are mediated by the interaction of genetic and epigenetic programs. The initiation of subunit mRNA expression occurs relatively early in cellular maturation and may be directed by intrinsic mechanisms. However, the levels of expression attained in adult animals may be controlled by extrinsic signals received by neurons during the postnatal maturation process. PMID:9777637

  14. The subunit composition of hinokiresinol synthase controls geometrical selectivity in norlignan formation.

    PubMed

    Suzuki, Shiro; Yamamura, Masaomi; Hattori, Takefumi; Nakatsubo, Tomoyuki; Umezawa, Toshiaki

    2007-12-26

    The selective formation of E- or Z-isomers is an important process in natural product metabolism. We show that the subunit composition of an enzyme can alter the geometrical composition of the enzymatic products. Hinokiresinol synthase, purified from Asparagus officinalis cell cultures, is responsible for the conversion of (7E,7'E)-4-coumaryl 4-coumarate to (Z)-hinokiresinol, the first step in norlignan formation. The protein is most likely a heterodimer composed of two distinct subunits, which share identity with members of the phloem protein 2 gene superfamily. Interestingly, each recombinant subunit of hinokiresinol synthase expressed in Escherichia coli solely converted (7E,7'E)-4-coumaryl 4-coumarate to the unnatural (E)-hinokiresinol, the E-isomer of (Z)-hinokiresinol. By contrast, a mixture of recombinant subunits catalyzed the formation of (Z)-hinokiresinol from the same substrate. PMID:18093914

  15. Glycine activated ion channel subunits encoded by ctenophore glutamate receptor genes.

    PubMed

    Alberstein, Robert; Grey, Richard; Zimmet, Austin; Simmons, David K; Mayer, Mark L

    2015-11-01

    Recent genome projects for ctenophores have revealed the presence of numerous ionotropic glutamate receptors (iGluRs) in Mnemiopsis leidyi and Pleurobrachia bachei, among our earliest metazoan ancestors. Sequence alignments and phylogenetic analysis show that these form a distinct clade from the well-characterized AMPA, kainate, and NMDA iGluR subtypes found in vertebrates. Although annotated as glutamate and kainate receptors, crystal structures of the ML032222a and PbiGluR3 ligand-binding domains (LBDs) reveal endogenous glycine in the binding pocket, whereas ligand-binding assays show that glycine binds with nanomolar affinity; biochemical assays and structural analysis establish that glutamate is occluded from the binding cavity. Further analysis reveals ctenophore-specific features, such as an interdomain Arg-Glu salt bridge, present only in subunits that bind glycine, but also a conserved disulfide in loop 1 of the LBD that is found in all vertebrate NMDA but not AMPA or kainate receptors. We hypothesize that ctenophore iGluRs are related to an early ancestor of NMDA receptors, suggesting a common evolutionary path for ctenophores and bilaterian species, and suggest that future work should consider both glycine and glutamate as candidate neurotransmitters in ctenophore species. PMID:26460032

  16. Glycine activated ion channel subunits encoded by ctenophore glutamate receptor genes

    PubMed Central

    Alberstein, Robert; Grey, Richard; Zimmet, Austin; Simmons, David K.; Mayer, Mark L.

    2015-01-01

    Recent genome projects for ctenophores have revealed the presence of numerous ionotropic glutamate receptors (iGluRs) in Mnemiopsis leidyi and Pleurobrachia bachei, among our earliest metazoan ancestors. Sequence alignments and phylogenetic analysis show that these form a distinct clade from the well-characterized AMPA, kainate, and NMDA iGluR subtypes found in vertebrates. Although annotated as glutamate and kainate receptors, crystal structures of the ML032222a and PbiGluR3 ligand-binding domains (LBDs) reveal endogenous glycine in the binding pocket, whereas ligand-binding assays show that glycine binds with nanomolar affinity; biochemical assays and structural analysis establish that glutamate is occluded from the binding cavity. Further analysis reveals ctenophore-specific features, such as an interdomain Arg-Glu salt bridge, present only in subunits that bind glycine, but also a conserved disulfide in loop 1 of the LBD that is found in all vertebrate NMDA but not AMPA or kainate receptors. We hypothesize that ctenophore iGluRs are related to an early ancestor of NMDA receptors, suggesting a common evolutionary path for ctenophores and bilaterian species, and suggest that future work should consider both glycine and glutamate as candidate neurotransmitters in ctenophore species. PMID:26460032

  17. Phylogenetic position of Gromia oviformis Dujardin inferred from nuclear-encoded small subunit ribosomal DNA.

    PubMed

    Burki, Fabien; Berney, Cédric; Pawlowski, Jan

    2002-09-01

    Gromia oviformis Dujardin is a common marine protist characterised by a large, globular test and filose pseudopodia. First considered a foraminifer, Gromia was later placed within the Filosea and recently included among amoebae of uncertain affinities. In order to clarify the phylogenetic position of this genus, we sequenced the complete small-subunit ribosomal DNA gene of G. oviformis collected at five different geographic localities. The high divergence of obtained sequences suggests that G. oviformis is a species complex composed of several genetically distinct sibling species. Sequence analyses show Gromia to be a member of the Cercozoa, a heterogeneous assemblage which includes filose amoebae, the amoeboflagellate cercomonads, the chlorarachniophytes and the plasmodiophorid plant pathogens. Contrary to traditional classification, Gromia is not closely related to other testate filose amoebae (the Euglyphida), but seems to branch early among the Cercozoa. Our analyses also show a close relationship between the Cercozoa and the Acantharea. Because the Cercozoa are related to the Foraminifera based on other molecular data, we propose that most protists possessing filopodia, reticulopodia and axopodia have a common origin.

  18. Glycine activated ion channel subunits encoded by ctenophore glutamate receptor genes.

    PubMed

    Alberstein, Robert; Grey, Richard; Zimmet, Austin; Simmons, David K; Mayer, Mark L

    2015-11-01

    Recent genome projects for ctenophores have revealed the presence of numerous ionotropic glutamate receptors (iGluRs) in Mnemiopsis leidyi and Pleurobrachia bachei, among our earliest metazoan ancestors. Sequence alignments and phylogenetic analysis show that these form a distinct clade from the well-characterized AMPA, kainate, and NMDA iGluR subtypes found in vertebrates. Although annotated as glutamate and kainate receptors, crystal structures of the ML032222a and PbiGluR3 ligand-binding domains (LBDs) reveal endogenous glycine in the binding pocket, whereas ligand-binding assays show that glycine binds with nanomolar affinity; biochemical assays and structural analysis establish that glutamate is occluded from the binding cavity. Further analysis reveals ctenophore-specific features, such as an interdomain Arg-Glu salt bridge, present only in subunits that bind glycine, but also a conserved disulfide in loop 1 of the LBD that is found in all vertebrate NMDA but not AMPA or kainate receptors. We hypothesize that ctenophore iGluRs are related to an early ancestor of NMDA receptors, suggesting a common evolutionary path for ctenophores and bilaterian species, and suggest that future work should consider both glycine and glutamate as candidate neurotransmitters in ctenophore species.

  19. Evolution, Expression Differentiation and Interaction Specificity of Heterotrimeric G-Protein Subunit Gene Family in the Mesohexaploid Brassica rapa

    PubMed Central

    Arya, Gulab C.; Kumar, Roshan; Bisht, Naveen C.

    2014-01-01

    Heterotrimeric G-proteins, comprising of Gα, Gβ, and Gγ subunits, are important signal transducers which regulate many aspects of fundamental growth and developmental processes in all eukaryotes. Initial studies in model plants Arabidopsis and rice suggest that the repertoire of plant G-protein is much simpler than that observed in metazoans. In order to assess the consequence of whole genome triplication events within Brassicaceae family, we investigated the multiplicity of G-protein subunit genes in mesohexaploid Brassica rapa, a globally important vegetable and oilseed crop. We identified one Gα (BraA.Gα1), three Gβ (BraA.Gβ1, BraA.Gβ2, and BraA.Gβ3), and five Gγ (BraA.Gγ1, BraA.Gγ2, BraA.Gγ3, BraA.Gγ4, and BraA.Gγ5) genes from B. rapa, with a possibility of 15 Gαβγ heterotrimer combinations. Our analysis suggested that the process of genome triplication coupled with gene-loss (gene-fractionation) phenomenon have shaped the quantitative and sequence diversity of G-protein subunit genes in the extant B. rapa genome. Detailed expression analysis using qRT-PCR assays revealed that the G-protein genes have retained ubiquitous but distinct expression profiles across plant development. The expression of multiple G-protein genes was differentially regulated during seed-maturation and germination stages, and in response to various phytohormone treatments and stress conditions. Yeast-based interaction analysis showed that G-protein subunits interacted in most of the possible combinations, with some degree of subunit-specific interaction specificity, to control the functional selectivity of G-protein heterotrimer in different cell and tissue-types or in response to different environmental conditions. Taken together, this research identifies a highly diverse G-protein signaling network known to date from B. rapa, and provides a clue about the possible complexity of G-protein signaling networks present across globally important Brassica species. PMID

  20. Evolution, expression differentiation and interaction specificity of heterotrimeric G-protein subunit gene family in the mesohexaploid Brassica rapa.

    PubMed

    Arya, Gulab C; Kumar, Roshan; Bisht, Naveen C

    2014-01-01

    Heterotrimeric G-proteins, comprising of Gα, Gβ, and Gγ subunits, are important signal transducers which regulate many aspects of fundamental growth and developmental processes in all eukaryotes. Initial studies in model plants Arabidopsis and rice suggest that the repertoire of plant G-protein is much simpler than that observed in metazoans. In order to assess the consequence of whole genome triplication events within Brassicaceae family, we investigated the multiplicity of G-protein subunit genes in mesohexaploid Brassica rapa, a globally important vegetable and oilseed crop. We identified one Gα (BraA.Gα1), three Gβ (BraA.Gβ1, BraA.Gβ2, and BraA.Gβ3), and five Gγ (BraA.Gγ1, BraA.Gγ2, BraA.Gγ3, BraA.Gγ4, and BraA.Gγ5) genes from B. rapa, with a possibility of 15 Gαβγ heterotrimer combinations. Our analysis suggested that the process of genome triplication coupled with gene-loss (gene-fractionation) phenomenon have shaped the quantitative and sequence diversity of G-protein subunit genes in the extant B. rapa genome. Detailed expression analysis using qRT-PCR assays revealed that the G-protein genes have retained ubiquitous but distinct expression profiles across plant development. The expression of multiple G-protein genes was differentially regulated during seed-maturation and germination stages, and in response to various phytohormone treatments and stress conditions. Yeast-based interaction analysis showed that G-protein subunits interacted in most of the possible combinations, with some degree of subunit-specific interaction specificity, to control the functional selectivity of G-protein heterotrimer in different cell and tissue-types or in response to different environmental conditions. Taken together, this research identifies a highly diverse G-protein signaling network known to date from B. rapa, and provides a clue about the possible complexity of G-protein signaling networks present across globally important Brassica species.

  1. Evolution, expression differentiation and interaction specificity of heterotrimeric G-protein subunit gene family in the mesohexaploid Brassica rapa.

    PubMed

    Arya, Gulab C; Kumar, Roshan; Bisht, Naveen C

    2014-01-01

    Heterotrimeric G-proteins, comprising of Gα, Gβ, and Gγ subunits, are important signal transducers which regulate many aspects of fundamental growth and developmental processes in all eukaryotes. Initial studies in model plants Arabidopsis and rice suggest that the repertoire of plant G-protein is much simpler than that observed in metazoans. In order to assess the consequence of whole genome triplication events within Brassicaceae family, we investigated the multiplicity of G-protein subunit genes in mesohexaploid Brassica rapa, a globally important vegetable and oilseed crop. We identified one Gα (BraA.Gα1), three Gβ (BraA.Gβ1, BraA.Gβ2, and BraA.Gβ3), and five Gγ (BraA.Gγ1, BraA.Gγ2, BraA.Gγ3, BraA.Gγ4, and BraA.Gγ5) genes from B. rapa, with a possibility of 15 Gαβγ heterotrimer combinations. Our analysis suggested that the process of genome triplication coupled with gene-loss (gene-fractionation) phenomenon have shaped the quantitative and sequence diversity of G-protein subunit genes in the extant B. rapa genome. Detailed expression analysis using qRT-PCR assays revealed that the G-protein genes have retained ubiquitous but distinct expression profiles across plant development. The expression of multiple G-protein genes was differentially regulated during seed-maturation and germination stages, and in response to various phytohormone treatments and stress conditions. Yeast-based interaction analysis showed that G-protein subunits interacted in most of the possible combinations, with some degree of subunit-specific interaction specificity, to control the functional selectivity of G-protein heterotrimer in different cell and tissue-types or in response to different environmental conditions. Taken together, this research identifies a highly diverse G-protein signaling network known to date from B. rapa, and provides a clue about the possible complexity of G-protein signaling networks present across globally important Brassica species. PMID

  2. Genetic Variation among Human Isolates of Uninucleated Cyst-Producing Entamoeba Species

    PubMed Central

    Verweij, Jaco J.; Polderman, Anton M.; Clark, C. Graham

    2001-01-01

    Twelve human infections with Entamoeba spp. producing uninucleated cysts were studied. DNA was extracted from infected feces and used to amplify part of the ameba small-subunit rRNA gene. Sequence analysis identified four distinct types of Entamoeba, all of which are related to Entamoeba polecki and E. chattoni and two of which have not been reported previously. Whether these genetic types represent different species is unclear. We propose that the agent of all human infections with uninucleated cyst-producing Entamoeba species be reported as “E. polecki-like.” PMID:11283106

  3. The relevance of the storage of subunit c of ATP synthase in different forms and models of Batten disease (NCLs).

    PubMed

    Palmer, David N

    2015-10-01

    The discoveries of specific protein storage in the NCLs, particularly of subunit c of ATP synthase in most, and the sphingolipid activator proteins, SAPs or saposins A and D in CLN1, CLN10 and an unassigned form are reviewed. The subunit c stored in the relevant NCLs is the complete mature molecule including an unusual modification found only in animal species, trimethylation of its lysine-43. Because of its strongly hydrophobic and lipid-like properties subunit c is easily overlooked or incorrectly described. This is becoming more of a problem as subunit c is not detected in standard proteomic investigations. Methods are reviewed that allow its unequivocal characterisation. Subunit c storage and cellular storage body accumulation do not cause the neuropathology characteristic of these diseases. The function of the trimethyl group on lysine-43 of subunit c is considered, along with some indications of where its normal turnover may be disrupted in the NCLs.

  4. Diversity of ribulose-1,5-bisphosphate carboxylase/oxygenase large-subunit genes from groundwater and aquifer microorganisms.

    PubMed

    Alfreider, A; Vogt, C; Hoffmann, D; Babel, W

    2003-05-01

    To test our hypothesis that microbial autotrophic CO2 fixation plays an important role in subsurface systems of two large groundwater remediation projects, several anaerobic/microaerobic aquifer and groundwater samples were taken and used to investigate the distribution and phylogenetic diversity of ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) large-subunit genes. Two primer sets were designed for amplifying partial-subunit genes of RubisCO forms I and II from the DNA, directly extracted from the samples. PCR products were used to construct five clone libraries with putative RubisCO form I sequences, and two libraries of DNA amplified by form II primers. Selected clones were screened for variation by restriction fragment length polymorphism analysis, and a total of 28 clone inserts were sequenced and further analyzed. The phylogenies constructed from amino acid sequences derived from the partial RubisCO large-subunit sequences showed a distinct pattern. Diverse sequences affiliated to the cluster of green-like type IA RubisCO sequences were found, representing various obligate and facultative chemolithoautotrophic Proteobacteria, whereas type II RubisCO sequences detected were most closely related to those of thiobacilli species. An isolate obtained from aquifer enrichment culture, which has been provisionally named Halothiobacillus sp. RA13 on the basis of its 16S rDNA sequence, was found to contain both types of RubisCO genes, i.e., forms I and II. Physiological and ecological considerations are discussed in the context of additional microbial data and physicochemical properties.

  5. Localization of Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Subunits during Intoxication of Live Cells

    PubMed Central

    Damek-Poprawa, Monika; Jang, Jae Yeon; Volgina, Alla; Korostoff, Jonathan

    2012-01-01

    The cytolethal distending toxin (Cdt), produced by some clinically important Gram-negative bacterial species, is related to the family of AB-type toxins. Three heterologous proteins (CdtA, CdtB, and CdtC) and a genotoxin mode of action distinguish the Cdt from others in this toxin class. Crystal structures of several species-specific Cdts have provided a basis for predicting subunit interactions and functions. In addition, empirical studies have yielded significant insights into the in vivo interactions of the Cdt subunits. However, there are still critical gaps in information about the intoxication process. In this study, a novel protein tagging technology was used to localize the subunits in Chinese hamster ovary cells (CHO-K1). A tetracysteine motif was engineered in each subunit, and in subunits with mutations in predicted functional domains, to permit detection with the fluorescein arsenical hairpin binding (FlAsH) dye Lumio green. Live-cell imaging, in conjunction with confocal microscopy, was used to capture the locations of the individual subunits in cells intoxicated, under various conditions, with hybrid heterotrimers. Using this approach, we observed the following. (i) The CdtA subunit remains on the cell surface of CHO cells in association with cholesterol-containing and cholesterol-depleted membrane. (ii) The CdtB subunit is exclusively in the cytosol and, after longer exposure times, localizes to the nucleus. (iii) The CdtC subunit is present on the cell surface and, to a greater extent, in the cytosol. These observations suggest that CdtC, but not CdtA, functions as a chaperone for CdtB entry into cells. PMID:22645284

  6. 28 CFR 51.6 - Political subunits.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 28 Judicial Administration 2 2010-07-01 2010-07-01 false Political subunits. 51.6 Section 51.6 Judicial Administration DEPARTMENT OF JUSTICE (CONTINUED) PROCEDURES FOR THE ADMINISTRATION OF SECTION 5 OF THE VOTING RIGHTS ACT OF 1965, AS AMENDED General Provisions § 51.6 Political subunits. All...

  7. 28 CFR 51.6 - Political subunits.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 28 Judicial Administration 2 2011-07-01 2011-07-01 false Political subunits. 51.6 Section 51.6 Judicial Administration DEPARTMENT OF JUSTICE (CONTINUED) PROCEDURES FOR THE ADMINISTRATION OF SECTION 5 OF THE VOTING RIGHTS ACT OF 1965, AS AMENDED General Provisions § 51.6 Political subunits. All...

  8. 28 CFR 51.6 - Political subunits.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 28 Judicial Administration 2 2012-07-01 2012-07-01 false Political subunits. 51.6 Section 51.6 Judicial Administration DEPARTMENT OF JUSTICE (CONTINUED) PROCEDURES FOR THE ADMINISTRATION OF SECTION 5 OF THE VOTING RIGHTS ACT OF 1965, AS AMENDED General Provisions § 51.6 Political subunits. All...

  9. 28 CFR 51.6 - Political subunits.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 28 Judicial Administration 2 2014-07-01 2014-07-01 false Political subunits. 51.6 Section 51.6 Judicial Administration DEPARTMENT OF JUSTICE (CONTINUED) PROCEDURES FOR THE ADMINISTRATION OF SECTION 5 OF THE VOTING RIGHTS ACT OF 1965, AS AMENDED General Provisions § 51.6 Political subunits. All...

  10. 28 CFR 51.6 - Political subunits.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 28 Judicial Administration 2 2013-07-01 2013-07-01 false Political subunits. 51.6 Section 51.6 Judicial Administration DEPARTMENT OF JUSTICE (CONTINUED) PROCEDURES FOR THE ADMINISTRATION OF SECTION 5 OF THE VOTING RIGHTS ACT OF 1965, AS AMENDED General Provisions § 51.6 Political subunits. All...

  11. Validity of a blue stripe snapper, Lutjanus octolineatus (Cuvier 1828) and a related species, L. bengalensis (Bloch 1790) with a new species (Pisces; Lutjanidae) from the Arabian Sea.

    PubMed

    Iwatsuki, Yukio; Al-Mamry, Juma M; Heemstra, Phillip C

    2016-04-07

    Lutjanus octolineatus (Cuvier 1828), previously considered a junior synonym of Lutjanus bengalensis (Bloch 1790), is shown to be a valid species and lectotypes are designated. Both species are redescribed. The two species have overlapping distributions in the Indian Ocean, but are clearly separable by different dorsal-fin spine counts, blue-striped pattern on the body and the presence or absence of a subocular extension of cheek scales. Lutjanus octovittata (Valenciennes 1830), formerly assigned to synonymy of L. bengalensis, is considered a junior synonym of L. octolineatus based on examination of the holotype. Lutjanus sapphirolineatus n. sp., a species formerly misidentified as L. bengalensis, is described based on 10 specimens from Oman and Somalia. The new species differs from the three species above by a combination of different characters. Analysis of the mitochondrial cytochrome c oxidase subunit 1 (CO1, 603 bp) genetic marker, also strongly supports the validity of each species of the blue-striped snapper complex as distinct.

  12. Recent radiation in a marine and freshwater dinoflagellate species flock.

    PubMed

    Annenkova, Nataliia V; Hansen, Gert; Moestrup, Øjvind; Rengefors, Karin

    2015-08-01

    Processes of rapid radiation among unicellular eukaryotes are much less studied than among multicellular organisms. We have investigated a lineage of cold-water microeukaryotes (protists) that appear to have diverged recently. This lineage stands in stark contrast to known examples of phylogenetically closely related protists, in which genetic difference is typically larger than morphological differences. We found that the group not only consists of the marine-brackish dinoflagellate species Scrippsiella hangoei and the freshwater species Peridinium aciculiferum as discovered previously but also of a whole species flock. The additional species include Peridinium euryceps and Peridinium baicalense, which are restricted to a few lakes, in particular to the ancient Lake Baikal, Russia, and freshwater S. hangoei from Lake Baikal. These species are characterized by relatively large conspicuous morphological differences, which have given rise to the different species descriptions. However, our scanning electron microscopic studies indicate that they belong to a single genus according to traditional morphological characterization of dinoflagellates (thecal plate patterns). Moreover, we found that they have identical SSU (small subunit) rDNA fragments and distinct but very small differences in the DNA markers LSU (large subunit) rDNA, ITS2 (internal transcribed spacer 2) and COB (cytochrome b) gene, which are used to delineate dinoflagellates species. As some of the species co-occur, and all four have small but species-specific sequence differences, we suggest that these taxa are not a case of phenotypic plasticity but originated via recent adaptive radiation. We propose that this is the first clear example among free-living microeukaryotes of recent rapid diversification into several species followed by dispersion to environments with different ecological conditions. PMID:25603395

  13. Recent radiation in a marine and freshwater dinoflagellate species flock.

    PubMed

    Annenkova, Nataliia V; Hansen, Gert; Moestrup, Øjvind; Rengefors, Karin

    2015-08-01

    Processes of rapid radiation among unicellular eukaryotes are much less studied than among multicellular organisms. We have investigated a lineage of cold-water microeukaryotes (protists) that appear to have diverged recently. This lineage stands in stark contrast to known examples of phylogenetically closely related protists, in which genetic difference is typically larger than morphological differences. We found that the group not only consists of the marine-brackish dinoflagellate species Scrippsiella hangoei and the freshwater species Peridinium aciculiferum as discovered previously but also of a whole species flock. The additional species include Peridinium euryceps and Peridinium baicalense, which are restricted to a few lakes, in particular to the ancient Lake Baikal, Russia, and freshwater S. hangoei from Lake Baikal. These species are characterized by relatively large conspicuous morphological differences, which have given rise to the different species descriptions. However, our scanning electron microscopic studies indicate that they belong to a single genus according to traditional morphological characterization of dinoflagellates (thecal plate patterns). Moreover, we found that they have identical SSU (small subunit) rDNA fragments and distinct but very small differences in the DNA markers LSU (large subunit) rDNA, ITS2 (internal transcribed spacer 2) and COB (cytochrome b) gene, which are used to delineate dinoflagellates species. As some of the species co-occur, and all four have small but species-specific sequence differences, we suggest that these taxa are not a case of phenotypic plasticity but originated via recent adaptive radiation. We propose that this is the first clear example among free-living microeukaryotes of recent rapid diversification into several species followed by dispersion to environments with different ecological conditions.

  14. Sarcocysts of an unidentified species of Sarcocystis in the sea otter (Enhydra lutris)

    USGS Publications Warehouse

    Dubey, J.P.; Lindsay, D.S.; Rosenthal, B.M.; Thomas, N.J.

    2003-01-01

    The number of Sarcocystis species that infect sea otters (Enhydra lutris) is unknown. Sea otter tissues were recently shown to harbor sarcocysts of S. neurona and of unidentified species of Sarcocystis. Whereas sarcocysts of S. neurona have walls 1a??3 I?m thick with type 9 villar protrusions, ultrastructure of a distinct thin-walled sarcocyst (0.5a??0.7 I?m thick) lacking villar protrusions, but instead exhibiting minute type 1 undulations on the sarcocyst wall, is described in this report. Parasites characterized from a sea otter infection were inferred to be related to, but distinct from, other species belonging to Sarcocystis, based on sequencing and phylogenetic analysis of a portion of the beta subunit of the plastid-encoded RNA polymerase gene.

  15. Functional Consequences of Subunit Diversity in RNA Polymerases II and V

    SciTech Connect

    Tan, Ek Han; Blevins, Todd; Ream, Thomas S.; Pikaard, Craig S.

    2012-03-01

    Multisubunit RNA polymerases IV and V (Pol IV and Pol V) evolved as specialized forms of Pol II that mediate RNA-directed DNA methylation (RdDM) and transcriptional silencing of transposons, viruses, and endogenous repeats in plants. Among the subunits common to Arabidopsis thaliana Pols II, IV, and V are 93% identical alternative ninth subunits, NRP(B/D/E)9a and NRP(B/D/E)9b. The 9a and 9b subunit variants are incompletely redundant with respect to Pol II; whereas double mutants are embryo lethal, single mutants are viable, yet phenotypically distinct. Likewise, 9a or 9b can associate with Pols IV or V but RNA-directed DNA methylation is impaired only in 9b mutants. Based on genetic and molecular tests, we attribute the defect in RdDM to impaired Pol V function. Collectively, our results reveal a role for the ninth subunit in RNA silencing and demonstrate that subunit diversity generates functionally distinct subtypes of RNA polymerases II and V.

  16. Distinct potassium channels on pain-sensing neurons.

    PubMed

    Rasband, M N; Park, E W; Vanderah, T W; Lai, J; Porreca, F; Trimmer, J S

    2001-11-01

    Differential expression of ion channels contributes functional diversity to sensory neuron signaling. We find nerve injury induced by the Chung model of neuropathic pain leads to striking reductions in voltage-gated K(+) (Kv) channel subunit expression in dorsal root ganglia (DRG) neurons, suggesting a potential molecular mechanism for hyperexcitability of injured nerves. Moreover, specific classes of DRG neurons express distinct Kv channel subunit combinations. Importantly, Kv1.4 is the sole Kv1 alpha subunit expressed in smaller diameter neurons, suggesting that homomeric Kv1.4 channels predominate in A delta and C fibers arising from these cells. These neurons are presumably nociceptors, because they also express the VR-1 capsaicin receptor, calcitonin gene-related peptide, and/or Na(+) channel SNS/PN3/Nav1.8. In contrast, larger diameter neurons associated with mechanoreception and proprioception express high levels of Kv1.1 and Kv1.2 without Kv1.4 or other Kv1 alpha subunits, suggesting that heteromers of these subunits predominate on large, myelinated afferent axons that extend from these cells. PMID:11698689

  17. Archaeal Mo-Containing Glyceraldehyde Oxidoreductase Isozymes Exhibit Diverse Substrate Specificities through Unique Subunit Assemblies

    PubMed Central

    Miyake, Masayuki; Fushinobu, Shinya

    2016-01-01

    Archaea use glycolytic pathways distinct from those found in bacteria and eukaryotes, where unique enzymes catalyze each reaction step. In this study, we isolated three isozymes of glyceraldehyde oxidoreductase (GAOR1, GAOR2 and GAOR3) from the thermoacidophilic archaeon Sulfolobus tokodaii. GAOR1–3 belong to the xanthine oxidoreductase superfamily, and are composed of a molybdo-pyranopterin subunit (L), a flavin subunit (M), and an iron-sulfur subunit (S), forming an LMS hetero-trimer unit. We found that GAOR1 is a tetramer of the STK17810/STK17830/STK17820 hetero-trimer, GAOR2 is a dimer of the STK23390/STK05620/STK05610 hetero-trimer, and GAOR3 is the STK24840/STK05620/STK05610 hetero-trimer. GAOR1–3 exhibited diverse substrate specificities for their electron donors and acceptors, due to their different L-subunits, and probably participate in the non-phosphorylative Entner-Doudoroff glycolytic pathway. We determined the crystal structure of GAOR2, as the first three-dimensional structure of an archaeal molybdenum-containing hydroxylase, to obtain structural insights into their substrate specificities and subunit assemblies. The gene arrangement and the crystal structure suggested that the M/S-complex serves as a structural scaffold for the binding of the L-subunit, to construct the three enzymes with different specificities. Collectively, our findings illustrate a novel principle of a prokaryotic multicomponent isozyme system. PMID:26808202

  18. Role of subunit-9 of mitochondrial ATP synthase in Batten disease

    SciTech Connect

    Johnson, D.W.; Speier, S.; Qian, W.H.

    1995-06-05

    The role of subunit-9 of mitochondrial ATP synthase in Batten disease was defined by characterizing the expression of genes encoding this protein in human tissues. Two genetically distinct neuronal ceroid-lipofuscinoses (NCL) comprise Batten disease: the late-infantile (LINCL) and juvenile (JNCL) types. We tested cell lines and tissues from both types of patients, along with normal controls. Differences in expression between diseased and normal samples were found for both mRNA and protein. Antibody staining of subunit-9 protein was detected in LINCL and JNCL tissues, and in 6 LINCL and 4 of 5 JNCL fibroblast lines. No immunoreactivity was seen in fibroblasts from obligate carriers, normal controls, and 6 other storage disease controls, with the exception of faint staining in Niemann-Pick, type C cells. There was an appreciable difference in staining pattern in both tissue sections and fibroblasts between LINCL and JNCL. Three subunit-9 transcripts (Hum1, Hum2, and Hum3) were specifically detected in NCL and normal human tissue from heart, liver, brain, muscle, and pancreas. Transcriptional regulation of subunit-9 genes was found to be altered in Batten disease. Pseudogenes related to each of the subunit-9 genes were isolated. Sequence analysis of cDNAs spanning the protein-coding regions of the Hum1, Hum2, and Hum3 genes showed conclusively that the primary defect(s) causing NCL are not mutations in the protein-coding regions of the 3 known subunit-9 genes. 29 refs., 6 figs., 5 tabs.

  19. The TCP1γ subunit of Leishmania donovani forms a biologically active homo-oligomeric complex.

    PubMed

    Bhaskar; Mitra, Kalyan; Kuldeep, Jitendra; Siddiqi, Mohammad Imran; Goyal, Neena

    2015-12-01

    Chaperonins are a class of molecular chaperons that encapsulate nascent or stress-denatured proteins and assist their intracellular assembly and folding in an ATP-dependent manner. The ubiquitous eukaryotic chaperonin, TCP1 ring complex is a hetero-oligomeric complex comprising two rings, each formed of eight subunits that may have distinct substrate recognition and ATP hydrolysis properties. In Leishmania, only the TCP1γ subunit has been cloned and characterized. It exhibited differential expression at various growth stages of promastigotes. In the present study, we expressed the TCP1γ subunit in Escherichia coli to investigate whether it forms chaperonin-like complexes and plays a role in protein folding. LdTCP1γ formed high-molecular-weight complexes within E. coli cells as well as in Leishmania cell lysates. The recombinant protein is arranged into two back-to-back rings of seven subunits each, as predicted by homology modelling and observed by negative staining electron microscopy. This morphology is consistent with that of the oligomeric double-ring group I chaperonins found in mitochondria. The LdTCP1γ homo-oligomeric complex hydrolysed ATP, and was active as assayed by luciferase refolding. Thus, the homo-oligomer performs chaperonin reactions without partner subunit(s). Further, co-immunoprecipitation studies revealed that LdTCP1γ interacts with actin and tubulin proteins, suggesting that the complex may have a role in maintaining the structural dynamics of the cytoskeleton of parasites. PMID:26395202

  20. Two classes of regulatory subunits coassemble in the same BK channel and independently regulate gating

    NASA Astrophysics Data System (ADS)

    Gonzalez-Perez, Vivian; Xia, Xiao-Ming; Lingle, Christopher J.

    2015-09-01

    High resolution proteomics increasingly reveals that most native ion channels are assembled in macromolecular complexes. However, whether different partners have additive or cooperative functional effects, or whether some combinations of proteins may preclude assembly of others are largely unexplored topics. The large conductance Ca2+-and-voltage activated potassium channel (BK) is well-suited to discern nuanced differences in regulation arising from combinations of subunits. Here we examine whether assembly of two different classes of regulatory proteins, β and γ, in BK channels is exclusive or independent. Our results show that both γ1 and up to four β2-subunits can coexist in the same functional BK complex, with the gating shift caused by β2-subunits largely additive with that produced by the γ1-subunit(s). The multiplicity of β:γ combinations that can participate in a BK complex therefore allow a range of BK channels with distinct functional properties tuned by the specific stoichiometry of the contributing subunits.

  1. The subunit structure of potato tuber ADPglucose pyrophosphorylase. [Solanum tuberosum L

    SciTech Connect

    Okita, T.W.; Nakata, P.A.; Anderson, J.M. ); Sowokinos, J. ); Morell, M.; Preiss, J. )

    1990-06-01

    ADPglucose pyrophosphorylase has been extensively purified from potato (Solanum tuberosum L.) tuber tissue to study its structure. By employing a modified published procedure together with Mono Q chromatography, a near homogeneous enzyme preparation was obtained with substantial improvement in enzyme yield and specific activity. In single dimensional sodium dodecyl sulfate polyacrylamide gels, the enzyme migrated as a single polypeptide band with a mobility of about 50,000 daltons. Analysis by two-dimensional polyacrylamide gel electrophoresis, however, revealed the presence of two types of subunits which could be distinguished by their slight differences in net charge and molecular weight. The smaller potato tuber subunit was recognized by antiserum prepared against the smaller spinach leaf 51 kilodalton ADPglucose pyrophosphorylase subunit. In contrast, the anti-54 kilodalton raised against the spinach leaf subunit did not significantly react to the tuber enzyme subunits. The results are consistent with the hypothesis that the potato tuber ADPglucose pyrophosphorylase is not composed of a simple homotetramer as previously suggested, but is a product of two separate and distinct subunits as observed for the spinach leaf and maize enzymes.

  2. Cell-free synthesis and assembly of prolyl 4-hydroxylase: the role of the beta-subunit (PDI) in preventing misfolding and aggregation of the alpha-subunit.

    PubMed Central

    John, D C; Grant, M E; Bulleid, N J

    1993-01-01

    Prolyl 4-hydroxylase (P4-H) catalyses a vital post-translational modification in the biosynthesis of collagen. The enzyme consists of two distinct polypeptides forming an alpha 2 beta 2 tetramer (alpha = 64 kDa, beta = 60 kDa), the beta-subunit being identical to the multifunctional enzyme protein disulfide isomerase (PDI). By studying the cell-free synthesis of the rat alpha-subunit of P4-H we have shown that the alpha-subunit can be translocated, glycosylated and the signal peptide cleaved by dog pancreatic microsomal membranes to yield both singly and doubly glycosylated forms. When translations were carried out under conditions which prevent disulfide bond formation, the product synthesized formed aggregates which were associated with the immunoglobulin heavy chain binding protein (BiP). Translations carried out under conditions that promote disulfide bond formation yielded a product that was not associated with BiP but formed a complex with the endogenous beta-subunit (PDI). Complex formation was detected by co-precipitation of the newly synthesized alpha-subunit with antibodies raised against PDI, by sucrose gradient centrifugation and by chemical cross-linking. When microsomal vesicles were depleted of PDI, BiP and other soluble endoplasmic reticulum proteins, no complex formation was observed and the alpha-subunit aggregated even under conditions that promote disulfide bond formation. We have therefore demonstrated that the enzyme P4-H can be assembled at synthesis in a cell-free system and that the solubility of the alpha-subunit is dependent upon its association with PDI. Images PMID:8385607

  3. Subunit arrangement in beef heart complex III

    SciTech Connect

    Gonzalez-Halphen, D.; Lindorfer, M.A.; Capaldi, R.A.

    1988-09-06

    Beef heart mitochondrial complex III was separated into 12 polypeptide bands representing 11 different subunits by using the electrophoresis conditions described previously. Eight of the 12 polypeptide bands were identified from their NH/sub 2/-terminal sequences as obtained by electroblotting directly from the NaDodSO/sub 4/-polyacrylamide gel onto a solid support. The topology of the subunits in complex III was explored by three different approaches. (1) Protease digestion experiments of submitochrondial particles in the presence and absence of detergent showed that subunits II and VI are on the M side of the inner membrane and subunits V and XI on the C side. (2) Labeling experiments with the membrane-intercalated probes (/sup 125/I)TID and arylazidoPE indicated that cytochrome b is the predominant bilayer embedded subunit of complex III, while the non-heme iron protein appears to be peripherally located. (3) Cross-linking studies with carbodiimides and homobifunctional cleavable reagents demonstrated that near-neighbor pairs include subunits I+II, II+VI, III+VI, IV+V, V+X, and V+VII. The cytochrome c binding site was found to include subunits IV, VII, and X. The combined data are used to provide an updated model of the topology of beef heart complex III.

  4. Cleft Lip Repair: The Hybrid Subunit Method.

    PubMed

    Tollefson, Travis T

    2016-04-01

    The unilateral cleft lip repair is one of the most rewarding and challenging of plastic surgery procedures. Surgeons have introduced a variety of straight line, geometric, and rotation-advancement designs, while in practice the majority of North American surgeons have been using hybrids of the rotation-advancement techniques. The anatomic subunit approach was introduced in 2005 by Fisher and has gained popularity, with early adopters of the design touting its simplicity and effectiveness. The objectives of this article are to summarize the basic tenets of respecting the philtral subunit, accurate measurement and planning, and tips for transitioning to this subunit approach.

  5. Cleft Lip Repair: The Hybrid Subunit Method.

    PubMed

    Tollefson, Travis T

    2016-04-01

    The unilateral cleft lip repair is one of the most rewarding and challenging of plastic surgery procedures. Surgeons have introduced a variety of straight line, geometric, and rotation-advancement designs, while in practice the majority of North American surgeons have been using hybrids of the rotation-advancement techniques. The anatomic subunit approach was introduced in 2005 by Fisher and has gained popularity, with early adopters of the design touting its simplicity and effectiveness. The objectives of this article are to summarize the basic tenets of respecting the philtral subunit, accurate measurement and planning, and tips for transitioning to this subunit approach. PMID:27097136

  6. Recent radiation in a marine and freshwater dinoflagellate species flock

    PubMed Central

    Annenkova, Nataliia V; Hansen, Gert; Moestrup, Øjvind; Rengefors, Karin

    2015-01-01

    Processes of rapid radiation among unicellular eukaryotes are much less studied than among multicellular organisms. We have investigated a lineage of cold-water microeukaryotes (protists) that appear to have diverged recently. This lineage stands in stark contrast to known examples of phylogenetically closely related protists, in which genetic difference is typically larger than morphological differences. We found that the group not only consists of the marine-brackish dinoflagellate species Scrippsiella hangoei and the freshwater species Peridinium aciculiferum as discovered previously but also of a whole species flock. The additional species include Peridinium euryceps and Peridinium baicalense, which are restricted to a few lakes, in particular to the ancient Lake Baikal, Russia, and freshwater S. hangoei from Lake Baikal. These species are characterized by relatively large conspicuous morphological differences, which have given rise to the different species descriptions. However, our scanning electron microscopic studies indicate that they belong to a single genus according to traditional morphological characterization of dinoflagellates (thecal plate patterns). Moreover, we found that they have identical SSU (small subunit) rDNA fragments and distinct but very small differences in the DNA markers LSU (large subunit) rDNA, ITS2 (internal transcribed spacer 2) and COB (cytochrome b) gene, which are used to delineate dinoflagellates species. As some of the species co-occur, and all four have small but species–specific sequence differences, we suggest that these taxa are not a case of phenotypic plasticity but originated via recent adaptive radiation. We propose that this is the first clear example among free-living microeukaryotes of recent rapid diversification into several species followed by dispersion to environments with different ecological conditions. PMID:25603395

  7. Purification and subunit heterogeneity of pili of Bordetella bronchiseptica.

    PubMed Central

    Lee, S W; Way, A W; Osen, E G

    1986-01-01

    Pili were isolated and purified from Bordetella bronchiseptica. Electron microscopic observations revealed that pili are ubiquitous in this species. The occurrence of pili and flagella appeared to correlate with growth phase and colonial morphology. Pili were about 3 to 4 nm in diameter and morphologically similar to pili isolated from other gram-negative bacteria. Internal core structure was not evident. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified pili showed that up to three different pilus subunit variants could be observed on a single strain, depending on the colonial phase and culture condition. Enzyme immunoassay and immunoblot, however, showed that these subunit variants are serologically related. Mice vaccinated with purified pili were protected against a virulent intraperitoneal challenge of B. bronchiseptica. B. bronchiseptica pili were also found to be similar to Bordetella pertussis pili in morphology and in the molecular size and antigenic structure of pilus subunits. The intact pili of B. bronchiseptica and B. pertussis, however, appeared to have weak serological cross-reactivity. Images PMID:2867974

  8. Mitochondrial NADH:ubiquinone oxidoreductase (complex I) in eukaryotes: a highly conserved subunit composition highlighted by mining of protein databases.

    PubMed

    Cardol, Pierre

    2011-11-01

    Complex I (NADH:ubiquinone oxidoreductase) is the largest enzyme of the mitochondrial respiratory chain. Compared to its bacterial counterpart which encompasses 14-17 subunits, mitochondrial complex I has almost tripled its subunit composition during evolution of eukaryotes, by recruitment of so-called accessory subunits, part of them being specific to distinct evolutionary lineages. The increasing availability of numerous broadly sampled eukaryotic genomes now enables the reconstruction of the evolutionary history of this large protein complex. Here, a combination of profile-based sequence comparisons and basic structural properties analyses at the protein level enabled to pinpoint homology relationships between complex I subunits from fungi, mammals or green plants, previously identified as "lineage-specific" subunits. In addition, homologs of at least 40 mammalian complex I subunits are present in representatives of all major eukaryote assemblages, half of them having not been investigated so far (Excavates, Chromalveolates, Amoebozoa). This analysis revealed that complex I was subject to a phenomenal increase in size that predated the diversification of extant eukaryotes, followed by very few lineage-specific additions/losses of subunits. The implications of this subunit conservation for studies of complex I are discussed. PMID:21749854

  9. Species from within the Phytophthora cryptogea complex and related species, P. erythroseptica and P. sansomeana, readily hybridize.

    PubMed

    Safaiefarahani, Banafsheh; Mostowfizadeh-Ghalamfarsa, Reza; Hardy, Giles E St J; Burgess, Treena I

    2016-08-01

    During a study on the phylogenetic relationships between species in the Phytophthora cryptogea complex and related species, Phytophthora erythroseptica and Phytophthora sansomeana, 19 hybrid isolates with multiple polymorphisms in the nuclear sequences were observed. Molecular characterization of hybrids was achieved by sequencing three nuclear (internal transcribed spacers, β-tubulin (TUB), heat shock protein 90) and two mitochondrial (cytochrome c oxidase subunit I (coxI), NADH dehydrogenase subunit I (NADH)) gene regions and cloning of the single-copy nuclear gene, TUB. Based on the molecular studies the hybrid isolates belonged to six distinct groups between P. cryptogea, P. erythroseptica, Phytophthora pseudocryptogea, P. sansomeana, and Phytophthora sp. kelmania. In all cases, only a single coxI and NADH allele was detected and nuclear genes were biparentally inherited, suggesting that the hybrids arose from sexual recombination events. Colony morphology, growth rate, cardinal temperatures, breeding system, and morphology of sporangia, oogonia, oospores, and antheridia were also determined. Some morphological differences between the hybrids and the parental species were noted; however, they were not sufficient to reliably distinguish the taxa and DNA markers from nuclear and mitochondrial genes will to be necessary for their identification. The parental species are all important pathogens of agricultural fields that have been transported globally. With the apparent ease of hybridization within this group there is ample opportunity for virulent hybrids to form, perhaps with extended host ranges. PMID:27521629

  10. Scytalidium parasiticum sp. nov., a New Species Parasitizing on Ganoderma boninense Isolated from Oil Palm in Peninsular Malaysia

    PubMed Central

    Goh, Teik Khiang; Marzuki, Nurul Fadhilah; Tung, Hun Jiat; Goh, You Keng; Goh, Kah Joo

    2015-01-01

    A mycoparasite, Scytalidium parasiticum sp. nov., isolated from the basidiomata of Ganoderma boninense causing basal stem rot of oil palm in Johor, Malaysia, is described and illustrated. It is distinct from other Scytalidium species in having smaller asci and ascospores (teleomorphic stage), longer arthroconidia (anamorphic stage), hyaline to yellowish chlamydospores, and producing a fluorescent pigment. The phylogenetic position of S. parasiticum was determined by sequence analyses of the internal transcribed spacers and the small-subunit ribosomal RNA gene regions. A key to identify Scytalidium species with teleomorphic stage is provided. PMID:26190917

  11. Description of Scheffersomyces henanensis sp. nov., a New D-Xylose-Fermenting Yeast Species Isolated from Rotten Wood

    PubMed Central

    Ren, Yongcheng; Chen, Liang; Niu, Qiuhong; Hui, Fengli

    2014-01-01

    Two strains of a D-xylose-fermenting yeast species were isolated from rotten wood samples collected from the Baotianman Nature Reserve in Henan Province, central China. These strains formed hat-shaped ascospores in conjugated and deliquescent asci. Multilocus phylogenetic analysis that included the nearly complete small subunit (SSU), the internal transcribed spacer (ITS) region and the D1/D2 domain of the large subunit (LSU) rRNA genes, as well as RNA polymerase II largest subunit (RPB1) gene demonstrated that the two strains represent a novel yeast species closely related to Scheffersomyces segobiensis. A sequence comparison of xylose reductase (XYL1) gene, which was recently recommended for rapid identification of cryptic species in the Scheffersomyces clade, revealed a significant sequence divergence of 25 nucleotides between the novel strains and their closest relative S. segobiensis, supporting their classification as a distinct species. Furthermore, these new strains can be clearly distinguished from S. segobiensis by a number of morphological and physiological characteristics. Therefore, a novel yeast species, Scheffersomyces henanensis sp. nov., is proposed to accommodate these strains. The type strain is BY-41T ( =  CICC 1974T  =  CBS 12475T). PMID:24647466

  12. Molecular and functional characterization of seven Na+/K+-ATPase β subunit paralogs in Senegalese sole (Solea senegalensis Kaup, 1858).

    PubMed

    Armesto, Paula; Infante, Carlos; Cousin, Xavier; Ponce, Marian; Manchado, Manuel

    2015-04-01

    In the present work, seven genes encoding Na(+),K(+)-ATPase (NKA) β-subunits in the teleost Solea senegalensis are described for the first time. Sequence analysis of the predicted polypeptides revealed a high degree of conservation with those of other vertebrate species and maintenance of important motifs involved in structure and function. Phylogenetic analysis clustered the seven genes into four main clades: β1 (atp1b1a and atp1b1b), β2 (atp1b2a and atp1b2b), β3 (atp1b3a and atp1b3b) and β4 (atp1b4). In juveniles, all paralogous transcripts were detected in the nine tissues examined albeit with different expression patterns. The most ubiquitous expressed gene was atp1b1a whereas atp1b1b was mainly detected in osmoregulatory organs (gill, kidney and intestine), and atp1b2a, atp1b2b, atp1b3a, atp1b3b and atp1b4 in brain. An expression analysis in three brain regions and pituitary revealed that β1-type transcripts were more abundant in pituitary than the other β paralogs with slight differences between brain regions. Quantification of mRNA abundance in gills after a salinity challenge showed an activation of atp1b1a and atp1b1b at high salinity water (60 ppt) and atp1b3a and atp1b3b in response to low salinity (5 ppt). Transcriptional analysis during larval development showed specific expression patterns for each paralog. Moreover, no differences in the expression profiles between larvae cultivated at 10 and 35 ppt were observed except for atp1b4 with higher mRNA levels at 10 than 35 ppt at 18 days post hatch. Whole-mount in situ hybridization analysis revealed that atp1b1b was mainly localized in gut, pronephric tubule, gill, otic vesicle, and chordacentrum of newly hatched larvae. All these data suggest distinct roles of NKA β subunits in tissues, during development and osmoregulation with β1 subunits involved in the adaptation to hyperosmotic conditions and β3 subunits to hypoosmotic environments.

  13. A case of orthologous sequences of hemocyanin subunits for an evolutionary study of horseshoe crabs: amino acid sequence comparison of immunologically identical subunits of Carcinoscorpius rotundicauda and Tachypleus tridentatus.

    PubMed

    Sugita, H; Shishikura, F

    1995-10-01

    About 83% of the amino acid sequence of hemocyanin subunit HR6 from the Southeast Asian horseshoe crab, Carcinoscorpius rotundicauda, has been determined. There is a difference of about 43% between HR6 and complete sequences of chelicerate hemocyanin subunits from the American horseshoe crab, Limulus polyphemus, and a tarantula, Eurypelma californicum. However, the immunologically identical subunits HR6 and HT6 from Tachypleus tridentatus (Japanese horseshoe crab) show 2.7% sequence difference. Based on the amino acid sequences of HR6 and HT6, the divergence between C. rotundicauda and T. tridentatus occurred about 9.6 million years ago. In the case of horseshoe crab hemocyanin subunits, it seems that the orthologous homologues in many homologous subunits between species are immunologically detectable.

  14. Phylogenetic position of Indian termites (Isoptera: Termitidae) with their respective genera inferred from DNA sequence analysis of the mitochondrial cytochrome oxidase gene subunit I compared to subunit II.

    PubMed

    Sharma, Vijay Lakshmi; Singla, Mandakini; Sobti, Ranbir Chander

    2013-08-20

    The present work was aimed to investigate the phylogenetic analysis of different species of Indian termites belonging to the family termitidae based on mitochondrial genes COI and COII. The sequences so obtained from public database revealed grouping of termites according to their ecological distribution. The sequences of the species under investigation were characterized on the basis of frequencies of nucleotide bases and in most of the species, a significantly high percentage of A+T base composition was observed. Phylogenetic tree revealed positioning of species according to the analysis of their cytochrome oxidase subunits.

  15. A consistent terminology for quantifying species diversity? Yes, it does exist.

    PubMed

    Tuomisto, Hanna

    2010-12-01

    The prevailing terminological confusion around the concept 'diversity' has hampered accurate communication and caused diversity issues to appear unnecessarily complicated. In fact, a consistent terminology for phenomena related to (species) diversity is already available. When this terminology is adhered to, diversity emerges as an easily understood concept. It is important to differentiate between diversity itself and a diversity index: an index of something is just a surrogate for the thing itself. The conceptual problem of defining diversity also has to be separated from the practical problem of deciding how to adequately quantify diversity for a community of interest. In practice, diversity can be quantified for any dataset where units of observation (such as individuals) have been classified into types (such as species). All that needs to be known is what proportion of the observed units belong to a type of mean abundance. Diversity equals the inverse of this mean, and it quantifies the effective number of the types of interest. In ecology, interest often (but not always) focuses on species diversity. If the dataset consists of (or gets divided into) subunits, then the total effective number of species (gamma diversity) can be partitioned into the effective number of compositionally distinct subunits (beta diversity) and the mean effective number of species per such subunit (alpha diversity). Species richness is related to species diversity, but they are not the same thing; richness does not take the proportional abundances into account and is therefore the actual-rather than the effective-number of types. Most of the phenomena that have been called 'beta diversity' in the past do not quantify an effective number of types, so they should be referred to by names other than 'diversity' (for example, species turnover or differentiation).

  16. Protein synthesis by ribosomes with tethered subunits.

    PubMed

    Orelle, Cédric; Carlson, Erik D; Szal, Teresa; Florin, Tanja; Jewett, Michael C; Mankin, Alexander S

    2015-08-01

    The ribosome is a ribonucleoprotein machine responsible for protein synthesis. In all kingdoms of life it is composed of two subunits, each built on its own ribosomal RNA (rRNA) scaffold. The independent but coordinated functions of the subunits, including their ability to associate at initiation, rotate during elongation, and dissociate after protein release, are an established model of protein synthesis. Furthermore, the bipartite nature of the ribosome is presumed to be essential for biogenesis, since dedicated assembly factors keep immature ribosomal subunits apart and prevent them from translation initiation. Free exchange of the subunits limits the development of specialized orthogonal genetic systems that could be evolved for novel functions without interfering with native translation. Here we show that ribosomes with tethered and thus inseparable subunits (termed Ribo-T) are capable of successfully carrying out protein synthesis. By engineering a hybrid rRNA composed of both small and large subunit rRNA sequences, we produced a functional ribosome in which the subunits are covalently linked into a single entity by short RNA linkers. Notably, Ribo-T was not only functional in vitro, but was also able to support the growth of Escherichia coli cells even in the absence of wild-type ribosomes. We used Ribo-T to create the first fully orthogonal ribosome-messenger RNA system, and demonstrate its evolvability by selecting otherwise dominantly lethal rRNA mutations in the peptidyl transferase centre that facilitate the translation of a problematic protein sequence. Ribo-T can be used for exploring poorly understood functions of the ribosome, enabling orthogonal genetic systems, and engineering ribosomes with new functions.

  17. Molecular cloning of the. cap alpha. subunit of human and guinea pig leukocyte adhesion glycoprotein Mo1: Chromosomal localization and homology to the. cap alpha. subunits of integrins

    SciTech Connect

    Arnaout, M.A.; Remold-O'Donnell, E.; Pierce, M.W.; Harris, P.; Tenen, D.G.

    1988-04-01

    The cell surface-glycoprotein Mo1 is a member of the family of leukocyte cell adhesion molecules (Leu-CAMs) that includes lymphocyte function-associated antigen 1 (LFA-1) and p150,95. Each Leu-CAM is a heterodimer with a distinct ..cap alpha.. subunit noncovalently associated with a common ..beta.. subunit. The authors describe the isolation and analysis of two partial cDNA clones encoding the ..cap alpha.. subunit of the Leu-CAM Mo1 in humans and guinea pigs. A monoclonal antibody directed against an epitope in the carboxyl-terminal portion of the guinea pig ..cap alpha.. chain was used for immunoscreening a lambdagt11 expression library. The sequence of a 378-base-pair insert from one immunoreactive clone revealed a single continuous open reading frame encoding 126 amino acids including a 26-amino acid tryptic peptide isolated from the purified guinea pig ..cap alpha.. subunit. A cDNA clone of identical size was isolated from a human monocyte/lymphocyte cDNA library by using the guinea pig clone as a probe. The human clone also encoded a 126-amino acid peptide including the sequence of an additional tryptic peptide present in purified human Mo1..cap alpha.. chain. Southern analysis of DNA from hamster-human hybrids localized the human Mo1..cap alpha.. chain to chromosome 16, which has been shown to contain the gene for the ..cap alpha.. chain of lymphocyte function-associated antigen 1. These data suggest that the ..cap alpha.. subunits of Leu-CAMs evolved by gene duplication from a common ancestral gene and strengthen the hypothesis that the ..cap alpha.. subunits of these heterodimeric cell adhesion molecules on myeloid and lymphoid cells, platelets, and fibroblasts are evolutionary related.

  18. The expression profile of acid-sensing ion channel (ASIC) subunits ASIC1a, ASIC1b, ASIC2a, ASIC2b, and ASIC3 in the esophageal vagal afferent nerve subtypes

    PubMed Central

    Dusenkova, Svetlana; Ru, Fei; Surdenikova, Lenka; Nassenstein, Christina; Hatok, Jozef; Dusenka, Robert; Banovcin, Peter; Kliment, Jan; Tatar, Milos

    2014-01-01

    Acid-sensing ion channels (ASICs) have been implicated in esophageal acid sensing and mechanotransduction. However, insufficient knowledge of ASIC subunit expression profile in esophageal afferent nerves hampers the understanding of their role. This knowledge is essential because ASIC subunits form heteromultimeric channels with distinct functional properties. We hypothesized that the esophageal putative nociceptive C-fiber nerves (transient receptor potential vanilloid 1, TRPV1-positive) express multiple ASIC subunits and that the ASIC expression profile differs between the nodose TRPV1-positive subtype developmentally derived from placodes and the jugular TRPV1-positive subtype derived from neural crest. We performed single cell RT-PCR on the vagal afferent neurons retrogradely labeled from the esophagus. In the guinea pig, nearly all (90%–95%) nodose and jugular esophageal TRPV1-positive neurons expressed ASICs, most often in a combination (65–75%). ASIC1, ASIC2, and ASIC3 were expressed in 65–75%, 55–70%, and 70%, respectively, of both nodose and jugular TRPV1-positive neurons. The ASIC1 splice variants ASIC1a and ASIC1b and the ASIC2 splice variant ASIC2b were similarly expressed in both nodose and jugular TRPV1-positive neurons. However, ASIC2a was found exclusively in the nodose neurons. In contrast to guinea pig, ASIC3 was almost absent from the mouse vagal esophageal TRPV1-positive neurons. However, ASIC3 was similarly expressed in the nonnociceptive TRPV1-negative (tension mechanoreceptors) neurons in both species. We conclude that the majority of esophageal vagal nociceptive neurons express multiple ASIC subunits. The placode-derived nodose neurons selectively express ASIC2a, known to substantially reduce acid sensitivity of ASIC heteromultimers. ASIC3 is expressed in the guinea pig but not in the mouse vagal esophageal TRPV1-positive neurons, indicating species differences in ASIC expression. PMID:25190475

  19. Molecular cloning of pituitary glycoprotein alpha-subunit and follicle stimulating hormone and chorionic gonadotropin beta-subunits from New World squirrel monkey and owl monkey.

    PubMed

    Scammell, Jonathan G; Funkhouser, Jane D; Moyer, Felricia S; Gibson, Susan V; Willis, Donna L

    2008-02-01

    The goal of this study was to characterize the gonadotropins expressed in pituitary glands of the New World squirrel monkey (Saimiri sp.) and owl monkey (Aotus sp.). The various subunits were amplified from total RNA from squirrel monkey and owl monkey pituitary glands by reverse transcription-polymerase chain reaction and the deduced amino acid sequences compared to those of other species. Mature squirrel monkey and owl monkey glycoprotein hormone alpha-polypeptides (96 amino acids in length) were determined to be 80% homologous to the human sequence. The sequences of mature beta subunits of follicle stimulating hormone (FSHbeta) from squirrel monkey and owl monkey (111 amino acids in length) are 92% homologous to human FSHbeta. New World primate glycoprotein hormone alpha-polypeptides and FSHbeta subunits showed conservation of all cysteine residues and consensus N-linked glycosylation sites. Attempts to amplify the beta-subunit of luteinizing hormone from squirrel monkey and owl monkey pituitary glands were unsuccessful. Rather, the beta-subunit of chorionic gonadotropin (CG) was amplified from pituitaries of both New World primates. Squirrel monkey and owl monkey CGbeta are 143 and 144 amino acids in length and 77% homologous with human CGbeta. The greatest divergence is in the C terminus, where all four sites for O-linked glycosylation in human CGbeta, responsible for delayed metabolic clearance, are predicted to be absent in New World primate CGbetas. It is likely that CG secreted from pituitary of New World primates exhibits a relatively short half-life compared to human CG.

  20. The resurrection of a species: Sarcocystis bovifelis Heydorn et al., 1975 is distinct from the current Sarcocystis hirsuta in cattle and morphologically indistinguishable from Sarcocystis sinensis in water buffaloes.

    PubMed

    Gjerde, Bjørn

    2016-01-01

    In the mid-1970s, it was established through transmission experiments and ultrastructural studies of sarcocysts by transmission electron microscopy (TEM) that cattle was the intermediate host of three Sarcocystis spp. using dogs, cats and humans, respectively, as definitive hosts. The cat-transmitted species with microscopic sarcocysts was initially named Sarcocystis bovifelis, but it was soon renamed Sarcocystis hirsuta, since it was considered to be identical with a previously named species. In recent years, an apparently new species has been detected in cattle in several countries by molecular methods and TEM and found by both methods to be indistinguishable from Sarcocystis sinensis in water buffaloes. This species was recently named Sarcocystis rommeli. Beginning in August 2014, a thorough review of papers comprising TEM micrographs of thick-walled sarcocysts in cattle was made in order to determine whether S. sinensis-like sarcocysts had been reported previously under other designations. Surprisingly, the review showed that the species S. bovifelis Heydorn et al., 1975 as described from cattle in Germany was S. sinensis-like and that indistinguishable sarcocysts had also been found in cattle in New Zealand and Canada in the 1980s. However, in the New Zealand study, these small sarcocysts were erroneously thought to represent developmental stages of a species with ultrastructurally similar but macroscopic sarcocysts, since the macroscopic cysts were found to be infective for cats. Thus, in the late 1980s, the cat-transmitted S. bovifelis, after having been renamed S. hirsuta, was erroneously synonymised with a second cat-transmitted species in cattle and then slid into obscurity until recently being rediscovered as a S. sinensis-like species in cattle and then named S. rommeli. Following the erroneous synonymisation, the name S. hirsuta has consistently been used for a taxon with macroscopic sarcocysts, and this usage should be continued. The name S. bovifelis

  1. The resurrection of a species: Sarcocystis bovifelis Heydorn et al., 1975 is distinct from the current Sarcocystis hirsuta in cattle and morphologically indistinguishable from Sarcocystis sinensis in water buffaloes.

    PubMed

    Gjerde, Bjørn

    2016-01-01

    In the mid-1970s, it was established through transmission experiments and ultrastructural studies of sarcocysts by transmission electron microscopy (TEM) that cattle was the intermediate host of three Sarcocystis spp. using dogs, cats and humans, respectively, as definitive hosts. The cat-transmitted species with microscopic sarcocysts was initially named Sarcocystis bovifelis, but it was soon renamed Sarcocystis hirsuta, since it was considered to be identical with a previously named species. In recent years, an apparently new species has been detected in cattle in several countries by molecular methods and TEM and found by both methods to be indistinguishable from Sarcocystis sinensis in water buffaloes. This species was recently named Sarcocystis rommeli. Beginning in August 2014, a thorough review of papers comprising TEM micrographs of thick-walled sarcocysts in cattle was made in order to determine whether S. sinensis-like sarcocysts had been reported previously under other designations. Surprisingly, the review showed that the species S. bovifelis Heydorn et al., 1975 as described from cattle in Germany was S. sinensis-like and that indistinguishable sarcocysts had also been found in cattle in New Zealand and Canada in the 1980s. However, in the New Zealand study, these small sarcocysts were erroneously thought to represent developmental stages of a species with ultrastructurally similar but macroscopic sarcocysts, since the macroscopic cysts were found to be infective for cats. Thus, in the late 1980s, the cat-transmitted S. bovifelis, after having been renamed S. hirsuta, was erroneously synonymised with a second cat-transmitted species in cattle and then slid into obscurity until recently being rediscovered as a S. sinensis-like species in cattle and then named S. rommeli. Following the erroneous synonymisation, the name S. hirsuta has consistently been used for a taxon with macroscopic sarcocysts, and this usage should be continued. The name S. bovifelis

  2. Seven new species within western Atlantic Starksia atlantica, S. lepicoelia, and S. sluiteri (Teleostei, Labrisomidae), with comments on congruence of DNA barcodes and species

    PubMed Central

    Baldwin, Carole C.; Castillo, Cristina I.; Weigt, Lee A.; Benjamin C., Victor

    2011-01-01

    Abstract Specimens of Starksia were collected throughout the western Atlantic, and a 650-bp portion of the mitochondrial gene cytochrome oxidase-c subunit I (COl) was sequenced as part of a re-analysis of species diversity of western Central Atlantic shorefishes. A neighbor-joining tree constructed from the sequence data suggests the existence of several cryptic species. Voucher specimens from each genetically distinct lineage and color photographs of vouchers taken prior to dissection and preservation were examined for diagnostic morphological characters. The results suggest that Starksia atlantica, Starksia lepicoelia, and Starksia sluiteri are species complexes, and each comprises three or more species. Seven new species are described. DNA data usually support morphological features, but some incongruence between genetic and morphological data exists. Genetic lineages are only recognized as species if supported by morphology. Genetic lineages within western Atlantic Starksia generally correspond to geography, such that members of each species complex have a very restricted geographical distribution. Increasing geographical coverage of sampling locations will almost certainly increase the number of Starksia species and species complexes recognized in the western Atlantic. Combining molecular and morphological investigations is bringing clarity to the taxonomy of many genera of morphologically similar fishes and increasing the number of currently recognized species. Future phylogenetic studies should help resolve species relationships and shed light on patterns of speciation in western Atlantic Starksia. PMID:21594143

  3. Ecology of cryptic invasions: latitudinal segregation among Watersipora (Bryozoa) species.

    PubMed

    Mackie, Joshua A; Darling, John A; Geller, Jonathan B

    2012-01-01

    Watersipora is an invasive genus of bryozoans, easily dispersed by fouled vessels. We examined Cytochrome c oxidase subunit I haplotypes from introduced populations on the US Pacific coastline to investigate geographic segregation of species and/or haplotypes. In California, the W. subtorquata group fell into three major sub-groups: W. subtorquata clades A and B, and W. "new sp.". W. subtorquata clades A and B were common in southern California south of Point Conception, a recognized biogeographic boundary, whereas further north, W. subtorquata clade A and W. n. sp. were frequent. The southern California region also had colonies of a morphologically distinct species, W. arcuata, also found in southern Australia and Hawaii; COI variation indicates a common ancestral source(s) in these introductions. The distribution of Watersipora-complex lineages on different coastlines is shown to be temperature correlated. Accordingly, pre-exisitng temperature-based adaptations may play a key role in determining invasion patterns.

  4. Glycine Receptors Containing α2 or α3 Subunits Regulate Specific Ethanol-Mediated Behaviors

    PubMed Central

    Blednov, Yuri A.; Benavidez, Jillian M.; Black, Mendy; Leiter, Courtney R.; Osterndorff-Kahanek, Elizabeth

    2015-01-01

    Glycine receptors (GlyRs) are broadly expressed in the central nervous system. Ethanol enhances the function of brain GlyRs, and the GlyRα1 subunit is associated with some of the behavioral actions of ethanol, such as loss of righting reflex. The in vivo role of GlyRα2 and α3 subunits in alcohol responses has not been characterized despite high expression levels in the nucleus accumbens and amygdala, areas that are important for the rewarding properties of drugs of abuse. We used an extensive panel of behavioral tests to examine ethanol actions in mice lacking Glra2 (the gene encoding the glycine receptor alpha 2 subunit) or Glra3 (the gene encoding the glycine receptor alpha 3 subunit). Deletion of Glra2 or Glra3 alters specific ethanol-induced behaviors. Glra2 knockout mice demonstrate reduced ethanol intake and preference in the 24-hour two-bottle choice test and increased initial aversive responses to ethanol and lithium chloride. In contrast, Glra3 knockout mice show increased ethanol intake and preference in the 24-hour intermittent access test and increased development of conditioned taste aversion to ethanol. Mutants and wild-type mice consumed similar amounts of ethanol in the limited access drinking in the dark test. Other ethanol effects, such as anxiolysis, motor incoordination, loss of righting reflex, and acoustic startle response, were not altered in the mutants. The behavioral changes in mice lacking GlyRα2 or α3 subunits were distinct from effects previously observed in mice with knock-in mutations in the α1 subunit. We provide evidence that GlyRα2 and α3 subunits may regulate ethanol consumption and the aversive response to ethanol. PMID:25678534

  5. Chronic Intermittent Ethanol Regulates Hippocampal GABA(A) Receptor Delta Subunit Gene Expression

    PubMed Central

    Follesa, Paolo; Floris, Gabriele; Asuni, Gino P.; Ibba, Antonio; Tocco, Maria G.; Zicca, Luca; Mercante, Beniamina; Deriu, Franca; Gorini, Giorgio

    2015-01-01

    Chronic ethanol consumption causes structural and functional reorganization in the hippocampus and induces alterations in the gene expression of gamma-aminobutyric acid type A receptors (GABAARs). Distinct forced intermittent exposure models have been used previously to investigate changes in GABAAR expression, with contrasting results. Here, we used repeated cycles of a Chronic Intermittent Ethanol paradigm to examine the relationship between voluntary, dependence-associated ethanol consumption, and GABAAR gene expression in mouse hippocampus. Adult male C57BL/6J mice were exposed to four 16-h ethanol vapor (or air) cycles in inhalation chambers alternated with limited-access two-bottle choice between ethanol (15%) and water consumption. The mice exposed to ethanol vapor showed significant increases in ethanol consumption compared to their air-matched controls. GABAAR alpha4 and delta subunit gene expression were measured by qRT-PCR at different stages. There were significant changes in GABAAR delta subunit transcript levels at different time points in ethanol-vapor exposed mice, while the alpha4 subunit levels remained unchanged. Correlated concurrent blood ethanol concentrations suggested that GABAAR delta subunit mRNA levels fluctuate depending on ethanol intoxication, dependence, and withdrawal state. Using a vapor-based Chronic Intermittent Ethanol procedure with combined two-bottle choice consumption, we corroborated previous evidences showing that discontinuous ethanol exposure affects GABAAR delta subunit expression but we did not observe changes in alpha4 subunit. These findings indicate that hippocampal GABAAR delta subunit expression changes transiently over the course of a Chronic Intermittent Ethanol paradigm associated with voluntary intake, in response to ethanol-mediated disturbance of GABAergic neurotransmission. PMID:26617492

  6. Mosquitoes of Anopheles hyrcanus (Diptera, Culicidae) Group: Species Diagnostic and Phylogenetic Relationships

    PubMed Central

    Khrabrova, Natalia V.; Andreeva, Yulia V.; Sibataev, Anuarbek K.; Alekseeva, Svetlana S.; Esenbekova, Perizat A.

    2015-01-01

    Herein, we report the results of study of Anopheles species in Primorsk and Khabarovsk regions of Russia. Three species of the Anopheles hyrcanus group: An. kleini, An. pullus, and An. lesteri were identified by molecular taxonomic diagnostics for the first time in Russia. Surprisingly, An. sinensis, which earlier was considered the only species of Anopheles in Russian Far East, was not observed. We analyzed nucleotide variation in the 610-bp fragment of the 5′ end of the cytochrome c oxidase subunit I (COI) region. All species possessed a distinctive set of COI sequences. A maximum likelihood phylogenetic tree was constructed for members of the hyrcanus group. The examined Anopheles hyrcanus group members could be divided into two major subgroups: subgroup 1 (An. hyrcanus and An. pullus) and subgroup 2 (An. sinensis, An. kleini, and An. lesteri), which were found to be monophyletic. PMID:26149867

  7. Mosquitoes of Anopheles hyrcanus (Diptera, Culicidae) Group: Species Diagnostic and Phylogenetic Relationships.

    PubMed

    Khrabrova, Natalia V; Andreeva, Yulia V; Sibataev, Anuarbek K; Alekseeva, Svetlana S; Esenbekova, Perizat A

    2015-09-01

    Herein, we report the results of study of Anopheles species in Primorsk and Khabarovsk regions of Russia. Three species of the Anopheles hyrcanus group: An. kleini, An. pullus, and An. lesteri were identified by molecular taxonomic diagnostics for the first time in Russia. Surprisingly, An. sinensis, which earlier was considered the only species of Anopheles in Russian Far East, was not observed. We analyzed nucleotide variation in the 610-bp fragment of the 5' end of the cytochrome c oxidase subunit I (COI) region. All species possessed a distinctive set of COI sequences. A maximum likelihood phylogenetic tree was constructed for members of the hyrcanus group. The examined Anopheles hyrcanus group members could be divided into two major subgroups: subgroup 1 (An. hyrcanus and An. pullus) and subgroup 2 (An. sinensis, An. kleini, and An. lesteri), which were found to be monophyletic.

  8. Identification and Characterization of High-Molecular-Weight Glutenin Subunits from Agropyron intermedium

    PubMed Central

    Cao, Shuanghe; Li, Zhixin; Gong, Caiyan; Xu, Hong; Yang, Ran; Hao, Shanting; Wang, Xianping; Wang, Daowen; Zhang, Xiangqi

    2014-01-01

    High-molecular-weight glutenin subunit (HMW-GS) is a primary determinant of processing quality of wheat. Considerable progress has been made in understanding the structure, function and genetic regulation of HMW-GS in wheat and some of its related species, but less is known about their orthologs in Agropyron intermedium, a useful related species for wheat improvement. Here seven HMW-GSs in Ag. intermedium were identified using SDS-PAGE and Western blotting experiments. Subsequently, the seven genes (Glu-1Aix1∼4 and Glu-1Aiy1∼3) encoding the seven HMW-GSs were isolated using PCR technique with degenerate primers, and confirmed by bacterial expression and Western blotting. Sequence analysis indicated that the seven Ag. intermedium HMW-GSs shared high similarity in primary structure to those of wheat, but four of the seven subunits were unusually small compared to the representatives of HMW-GS from wheat and two of them possessed extra cysteine residues. The alignment and clustering analysis of deduced amino acid sequences revealed that 1Aix1 and 1Aiy1 subunits had special molecular structure, belonging to the hybrid type compounding between typical x- and y-type subunit. The xy-type subunit 1Aix1 is composed of the N-terminal of x-type and C-terminal of y-type, whereas yx-type subunit 1Aiy1 comprises the N-terminal of y-type and C-terminal of x-type. This result strongly supported the hypothesis of unequal crossover mechanism that might generate the novel coding sequence for the hybrid type of HMW-GSs. In addition to the aforementioned, the other novel characteristics of the seven subunits were also discussed. Finally, phylogenetic analysis based on HMW-GS genes was carried out and provided new insights into the evolutionary biology of Ag. intermedium. PMID:24503781

  9. Identification and characterization of high-molecular-weight glutenin subunits from Agropyron intermedium.

    PubMed

    Cao, Shuanghe; Li, Zhixin; Gong, Caiyan; Xu, Hong; Yang, Ran; Hao, Shanting; Wang, Xianping; Wang, Daowen; Zhang, Xiangqi

    2014-01-01

    High-molecular-weight glutenin subunit (HMW-GS) is a primary determinant of processing quality of wheat. Considerable progress has been made in understanding the structure, function and genetic regulation of HMW-GS in wheat and some of its related species, but less is known about their orthologs in Agropyron intermedium, a useful related species for wheat improvement. Here seven HMW-GSs in Ag. intermedium were identified using SDS-PAGE and Western blotting experiments. Subsequently, the seven genes (Glu-1Aix1 ∼ 4 and Glu-1Aiy1 ∼ 3) encoding the seven HMW-GSs were isolated using PCR technique with degenerate primers, and confirmed by bacterial expression and Western blotting. Sequence analysis indicated that the seven Ag. intermedium HMW-GSs shared high similarity in primary structure to those of wheat, but four of the seven subunits were unusually small compared to the representatives of HMW-GS from wheat and two of them possessed extra cysteine residues. The alignment and clustering analysis of deduced amino acid sequences revealed that 1Aix1 and 1Aiy1 subunits had special molecular structure, belonging to the hybrid type compounding between typical x- and y-type subunit. The xy-type subunit 1Aix1 is composed of the N-terminal of x-type and C-terminal of y-type, whereas yx-type subunit 1Aiy1 comprises the N-terminal of y-type and C-terminal of x-type. This result strongly supported the hypothesis of unequal crossover mechanism that might generate the novel coding sequence for the hybrid type of HMW-GSs. In addition to the aforementioned, the other novel characteristics of the seven subunits were also discussed. Finally, phylogenetic analysis based on HMW-GS genes was carried out and provided new insights into the evolutionary biology of Ag. intermedium. PMID:24503781

  10. Structural analysis of the α subunit of Na(+)/K(+) ATPase genes in invertebrates.

    PubMed

    Thabet, Rahma; Rouault, J-D; Ayadi, Habib; Leignel, Vincent

    2016-01-01

    The Na(+)/K(+) ATPase is a ubiquitous pump coordinating the transport of Na(+) and K(+) across the membrane of cells and its role is fundamental to cellular functions. It is heteromer in eukaryotes including two or three subunits (α, β and γ which is specific to the vertebrates). The catalytic functions of the enzyme have been attributed to the α subunit. Several complete α protein sequences are available, but only few gene structures were characterized. We identified the genomic sequences coding the α-subunit of the Na(+)/K(+) ATPase, from the whole-genome shotgun contigs (WGS), NCBI Genomes (chromosome), Genomic Survey Sequences (GSS) and High Throughput Genomic Sequences (HTGS) databases across distinct phyla. One copy of the α subunit gene was found in Annelida, Arthropoda, Cnidaria, Echinodermata, Hemichordata, Mollusca, Placozoa, Porifera, Platyhelminthes, Urochordata, but the nematodes seem to possess 2 to 4 copies. The number of introns varied from 0 (Platyhelminthes) to 26 (Porifera); and their localization and length are also highly variable. Molecular phylogenies (Maximum Likelihood and Maximum Parsimony methods) showed some clusters constituted by (Chordata/(Echinodermata/Hemichordata)) or (Plathelminthes/(Annelida/Mollusca)) and a basal position for Porifera. These structural analyses increase our knowledge about the evolutionary events of the α subunit genes in the invertebrates.

  11. Structural analysis of the α subunit of Na(+)/K(+) ATPase genes in invertebrates.

    PubMed

    Thabet, Rahma; Rouault, J-D; Ayadi, Habib; Leignel, Vincent

    2016-01-01

    The Na(+)/K(+) ATPase is a ubiquitous pump coordinating the transport of Na(+) and K(+) across the membrane of cells and its role is fundamental to cellular functions. It is heteromer in eukaryotes including two or three subunits (α, β and γ which is specific to the vertebrates). The catalytic functions of the enzyme have been attributed to the α subunit. Several complete α protein sequences are available, but only few gene structures were characterized. We identified the genomic sequences coding the α-subunit of the Na(+)/K(+) ATPase, from the whole-genome shotgun contigs (WGS), NCBI Genomes (chromosome), Genomic Survey Sequences (GSS) and High Throughput Genomic Sequences (HTGS) databases across distinct phyla. One copy of the α subunit gene was found in Annelida, Arthropoda, Cnidaria, Echinodermata, Hemichordata, Mollusca, Placozoa, Porifera, Platyhelminthes, Urochordata, but the nematodes seem to possess 2 to 4 copies. The number of introns varied from 0 (Platyhelminthes) to 26 (Porifera); and their localization and length are also highly variable. Molecular phylogenies (Maximum Likelihood and Maximum Parsimony methods) showed some clusters constituted by (Chordata/(Echinodermata/Hemichordata)) or (Plathelminthes/(Annelida/Mollusca)) and a basal position for Porifera. These structural analyses increase our knowledge about the evolutionary events of the α subunit genes in the invertebrates. PMID:26812300

  12. Immunochemical characterization of two thyroid-stimulating hormone beta-subunit epitopes.

    PubMed Central

    Fairlie, W D; Stanton, P G; Hearn, M T

    1995-01-01

    The epitopes of human thyroid-stimulating hormone (hTSH) recognized by two murine monoclonal antibodies (MAbs), designated MAb 279 and MAb 299, have been characterized. These MAbs are highly specific for the beta-subunit of TSH. The epitope recognized by MAb 279 appears to be completely conserved between bovine and human TSH and partially conserved in the porcine species. The TSH beta-subunit epitope recognized by MAb 299 is only partially conserved between the human, bovine and porcine species. Both MAbs are capable of inhibiting the binding of TSH to its receptor in a TSH radioreceptor assay, indicating that the epitopes either coincide or are located close to the TSH beta-subunit receptor-binding sites. The carbohydrate moieties of the TSH beta-subunit appear to play little or no role in the epitope recognition by MAb 279 or MAb 299 while the integrity of the disulphide bonds are essential. The epitopic recognition may also involve lysine residues, as determined by the immunoreactivity with both MAbs following citraconylation of TSH. In addition, the amino acid sequence region between residues bTSH beta 34-44 could be excised by trypsin digestion of bovine TSH beta (bTSH beta) without eliminating epitopic recognition by either MAb. These results provide further insight into the relationship between the structure of the TSH beta-subunit epitopes and location of the receptor-binding sites. Images Figure 2 PMID:7538754

  13. Molecular phylogeny of Stentor (Ciliophora: Heterotrichea) based on small subunit ribosomal RNA sequences.

    PubMed

    Gong, Ying-Chun; Yu, Yu-He; Zhu, Fei-Yun; Feng, Wei-Song

    2007-01-01

    To determine the phylogenetic position of Stentor within the Class Heterotrichea, the complete small subunit rRNA genes of three Stentor species, namely Stentor polymorphus, Stentor coeruleus, and Stentor roeseli, were sequenced and used to construct phylogenetic trees using the maximum parsimony, neighbor joining, and Bayesian analysis. With all phylogenetic methods, the genus Stentor was monophyletic, with S. roeseli branching basally. PMID:17300519

  14. Recognition of chimeric small-subunit ribosomal DNAs composed of genes from uncultivated microorganisms

    NASA Technical Reports Server (NTRS)

    Kopczynski, E. D.; Bateson, M. M.; Ward, D. M.

    1994-01-01

    When PCR was used to recover small-subunit (SSU) rRNA genes from a hot spring cyanobacterial mat community, chimeric SSU rRNA sequences which exhibited little or no secondary structural abnormality were recovered. They were revealed as chimeras of SSU rRNA genes of uncultivated species through separate phylogenetic analysis of short sequence domains.

  15. Monoclonal Antibodies to the [alpha]- and [beta]-Subunits of the Plant Mitochondrial F1-ATPase.

    PubMed Central

    Luethy, M. H.; Horak, A.; Elthon, T. E.

    1993-01-01

    We have generated nine monoclonal antibodies against subunits of the maize (Zea mays L.) mitochondrial F1-ATPase. These monoclonal antibodies were generated by immunizing mice against maize mitochondrial fractions and randomly collecting useful hybridomas. To prove that these monoclonal antibodies were directed against ATPase subunits, we tested their cross-reactivity with purified F1-ATPase from pea cotyledon mitochondria. One of the antibodies ([alpha]-ATPaseD) cross-reacted with the pea F1-ATPase [alpha]-subunit and two ([beta]-ATPaseD and [beta]-ATPaseE) cross-reacted with the pea F1-ATPase [beta]-subunit. This established that, of the nine antibodies, four react with the maize [alpha]-ATPase subunit and the other five react with the maize [beta]-ATPase subunit. Most of the monoclonal antibodies cross-react with the F1-ATPase from a wide range of plant species. Each of the four monoclonal antibodies raised against the [alpha]-subunit recognizes a different epitope. Of the five [beta]-subunit antibodies, at least three different epitopes are recognized. Direct incubation of the monoclonal antibodies with the F1-ATPase failed to inhibit the ATPase activity. The monoclonal antibodies [alpha]-ATPaseD and [beta]-ATPaseD were bound to epoxide-glass QuantAffinity beads and incubated with a purified preparation of pea F1-ATPase. The ATPase activity was not inhibited when the antibodies bound the ATPase. The antibodies were used to help map the pea F1-ATPase subunits on a two-dimensional map of whole pea cotyledon mitochondrial protein. In addition, the antibodies have revealed antigenic similarities between various isoforms observed for the [alpha]- and [beta]-subunits of the purified F1-ATPase. The specificity of these monoclonal antibodies, along with their cross-species recognition and their ability to bind the F1-ATPase without inhibiting enzymic function, makes these antibodies useful and invaluable tools for the further purification and characterization of plant

  16. Human CCT4 and CCT5 Chaperonin Subunits Expressed in Escherichia coli Form Biologically Active Homo-oligomers*

    PubMed Central

    Sergeeva, Oksana A.; Chen, Bo; Haase-Pettingell, Cameron; Ludtke, Steven J.; Chiu, Wah; King, Jonathan A.

    2013-01-01

    Chaperonins are a family of chaperones that encapsulate their substrates and assist their folding in an ATP-dependent manner. The ubiquitous eukaryotic chaperonin, TCP-1 ring complex (TRiC), is a hetero-oligomeric complex composed of two rings, each formed from eight different CCT (chaperonin containing TCP-1) subunits. Each CCT subunit may have distinct substrate recognition and ATP hydrolysis properties. We have expressed each human CCT subunit individually in Escherichia coli to investigate whether they form chaperonin-like double ring complexes. CCT4 and CCT5, but not the other six CCT subunits, formed high molecular weight complexes within the E. coli cells that sedimented about 20S in sucrose gradients. When CCT4 and CCT5 were purified, they were both organized as two back-to-back rings of eight subunits each, as seen by negative stain and cryo-electron microscopy. This morphology is consistent with that of the hetero-oligomeric double-ring TRiC purified from bovine testes and HeLa cells. Both CCT4 and CCT5 homo-oligomers hydrolyzed ATP at a rate similar to human TRiC and were active as assayed by luciferase refolding and human γD-crystallin aggregation suppression and refolding. Thus, both CCT4 and CCT5 homo-oligomers have the property of forming 8-fold double rings absent the other subunits, and these complexes carry out chaperonin reactions without other partner subunits. PMID:23612981

  17. Essential role of BRG, the ATPase subunit of BAF chromatin remodeling complexes, in leukemia maintenance

    PubMed Central

    Buscarlet, Manuel; Krasteva, Veneta; Ho, Lena; Simon, Camille; Hébert, Josée; Wilhelm, Brian; Crabtree, Gerald R.; Sauvageau, Guy; Thibault, Pierre

    2014-01-01

    In mammals, combinatorial assembly of alternative families of subunits confers functional specificity to adenosine triphosphate (ATP)-dependent SWI/SNF-like Brg/Brm-associated factor (BAF) chromatin remodeling complexes by creating distinct polymorphic surfaces for interaction with regulatory elements and DNA-binding factors. Although redundant in terms of biochemical activity, the core ATPase subunits, BRG/SMARCA4 and BRM/SMARCA2, are functionally distinct and may contribute to complex specificity. Here we show using quantitative proteomics that BAF complexes expressed in leukemia are specifically assembled around the BRG ATPase. Moreover, using a mouse model of acute myeloid leukemia, we demonstrate that BRG is essential for leukemia maintenance, as leukemic cells lacking BRG rapidly undergo cell-cycle arrest and apoptosis. Most importantly, we show that BRG is dispensable for the maintenance of immunophenotypic long-term repopulating hematopoietic stem cells, suggesting that adroit targeting of BRG in leukemia may have potent and specific therapeutic effects. PMID:24478402

  18. Auxiliary subunits of the CKAMP family differentially modulate AMPA receptor properties

    PubMed Central

    Farrow, Paul; Khodosevich, Konstantin; Sapir, Yechiam; Schulmann, Anton; Aslam, Muhammad

    2015-01-01

    AMPA receptor (AMPAR) function is modulated by auxiliary subunits. Here, we report on three AMPAR interacting proteins—namely CKAMP39, CKAMP52 and CKAMP59—that, together with the previously characterized CKAMP44, constitute a novel family of auxiliary subunits distinct from other families of AMPAR interacting proteins. The new members of the CKAMP family display distinct regional and developmental expression profiles in the mouse brain. Notably, despite their structural similarities they exert diverse modulation on AMPAR gating by influencing deactivation, desensitization and recovery from desensitization, as well as glutamate and cyclothiazide potency to AMPARs. This study indicates that AMPAR function is very precisely controlled by the cell-type specific expression of the CKAMP family members. DOI: http://dx.doi.org/10.7554/eLife.09693.001 PMID:26623514

  19. First record and a new species of Alvinocaris Williams & Chace, 1982 (Crustacea: Decapoda: Caridea: Alvinocarididae) from the Indian Ocean.

    PubMed

    Yahagi, Takuya; Watanabe, Hiromi; Kojima, Shigeaki; Beedessee, Girish; Komai, Tomoyuki

    2014-12-05

    A new species of the alvinocaridid shrimp genus Alvinocaris Williams & Chace, 1982 is described from the Solitaire hydrothermal vent field at 2606 m depth on the Central Indian Ridge. Alvinocaris solitaire sp. nov., the first species of the genus to be recorded from the Indian Ocean, is morphologically most similar to A. lusca Williams & Chace, 1982 from the Galapagos Rift, East Pacific Rise. The new species is distinguished from A. lusca by the less produced pterygostomial angle of the carapace, the presence of small teeth on the posterolateral margin of the third pleuron, and the lack of short plumose setae on the posteromedian margin of the telson. The genetic divergence of the mitochondrial cytochrome c oxidase subunit I (COI) gene (600 bp) among the nine Alvinocaris species analyzed clearly indicates that the new taxon is distinct from the congeneric species for which genetic data are available.

  20. A new species of Macrophthalmus Desmarest, 1823 (Crustacea: Decapoda: Brachyura: Macrophthalmidae) from Taiwan, with notes on four new records.

    PubMed

    Teng, Shao-Jyun; Shih, Hsi-Te

    2015-12-18

    A new species of sentinel crab (family Macrophthalmidae), Macrophthalmus (Mareotis) purpureocheir sp. nov., from coral reefs is herein described from southern and eastern Taiwan. The new species can be distinguished from other species by a suite of characters, including the narrower and more convex carapace, the first anterolateral tooth protruding forward, the short and stout chelipeds, and the male first gonopod with short apical process, as well as the distinctive reddish purple chelae of the male. The identity of this new species is also supported by molecular evidence using the mitochondrial 16S rDNA and cytochrome oxidase subunit I (COI). Three additional species of Macrophthalmus, viz. M. crinitus Rathbun, 1913, M. definitus Adams & White, 1849, and M. milloti Crosnier, 1965, as well as Ilyograpsus paludicola (Rathbun, 1909) of the subfamily Ilyograpsinae Števčić, 2005, are also confirmed from Taiwan.

  1. A new species of Centropages (Copepoda: Calanoida: Centropagidae) from the central Red Sea based on morphological and molecular evidence.

    PubMed

    El-Sherbiny, Mohsen M; Al-Aidaroos, Ali M

    2015-01-01

    Centropages mohamedi sp. nov. (Copepoda: Calanoida) is described from specimens collected in zooplankton samples off Obhur Creek on the Saudi Arabian coast of the central Red Sea. The new species is most closely related to C. orsinii Giesbrecht, 1889, a species described from the Red Sea and widely distributed in the neritic waters of the Indo-West Pacific region. The new species is mainly distinguished by the female genital double somite, male antennules, male leg 4, and leg 5 of both sexes. DNA sequences of the mitochondrial cytochrome C oxidase subunit one (mtCOI) of the two species differ by 20.8%, supporting their morphology-based identification as distinct species.  PMID:25661620

  2. A new species of Centropages (Copepoda: Calanoida: Centropagidae) from the central Red Sea based on morphological and molecular evidence.

    PubMed

    El-Sherbiny, Mohsen M; Al-Aidaroos, Ali M

    2015-01-20

    Centropages mohamedi sp. nov. (Copepoda: Calanoida) is described from specimens collected in zooplankton samples off Obhur Creek on the Saudi Arabian coast of the central Red Sea. The new species is most closely related to C. orsinii Giesbrecht, 1889, a species described from the Red Sea and widely distributed in the neritic waters of the Indo-West Pacific region. The new species is mainly distinguished by the female genital double somite, male antennules, male leg 4, and leg 5 of both sexes. DNA sequences of the mitochondrial cytochrome C oxidase subunit one (mtCOI) of the two species differ by 20.8%, supporting their morphology-based identification as distinct species

  3. Subunit constituent of the porin trimers that form the permeability channels in the outer membrane of Salmonella typhimurium.

    PubMed Central

    Ishii, J; Nakae, T

    1980-01-01

    The polypeptide composition of the functional porin trimers that produced the permeability channels in the outer membrane of Salmonella typhimurium was examined on two-dimensional slab gels. The results suggested that the majority of porin trimers from strains producing mixed species of porin polypeptides consisted of homologous subunit polypeptides. The present results do not exclude the possibility that a small fraction of porin trimer is constructed from heterologous subunit polypeptides. Images PMID:6246065

  4. Genetics of the mammalian phenylalanine hydroxylase system. Studies of human liver phenylalanine hydroxylase subunit structure and of mutations in phenylketonuria.

    PubMed Central

    Choo, K H; Cotton, R G; Danks, D M; Jennings, I G

    1979-01-01

    Phenylalanine hydroxylase was purified from crude extracts of human livers which show enzyme activity by usine two different methods: (a) affinity chromatography and (b) immunoprecipitation with an antiserum against highly purified monkey liver phenylalanine hydroxylase. Purified human liver phenylalanine hydroxylase has an estimated mol. wt. of 275 000, and subunit mol. wts. of approx. 50 000 and 49 000. These two molecular-weight forms are designated H and L subunits. On two-dimensional polyacrylamide gel under dissociating conditions, enzyme purified by the two methods revealed at least six subunit species, which were resolved into two size classes. Two of these species have a molecular weight corresponding to that of the H subunit, whereas the other four have a molecular weight corresponding to that of the L subunit. This evidence indicates that active phenylalanine hydroxylase purified from human liver is composed of a mixture of sununits which are different in charge and size. None of the subunit species could be detected in crude extracts of livers from two patients with classical phenylketonuria by either the affinity or the immunoprecipitation method. However, they were present in liver from a patient with malignant hyperphenylalaninaemia with normal activity of dihydropteridine reductase. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:496890

  5. Site Specific Phosphorylation of Cytochrome c Oxidase Subunits I, IVi1 and Vb in Rabbit Hearts Subjected to Ischemia/Reperfusion

    PubMed Central

    Fang, Ji-Kang; Prabu, Subbuswamy K.; Sepuri, Naresh B.; Raza, Haider; Anandatheerthavarada, Hindupur K.; Galati, Domenico; Spear, Joseph; Avadhani, Narayan G.

    2007-01-01

    We have mapped the sites of ischemia/reperfusion induced phosphorylation of cytochrome c oxidase (CcO) subunits in rabbit hearts by using a combination of Blue Native gel/Tricine gel electrophoresis and nano-LC-MS/MS approaches. We used precursor ion scanning combined with neutral loss scanning and found that mature CcO subunit I was phosphorylated at tandem Ser115/Ser116 positions, subunit IVi1 at Thr52 and subunit Vb at Ser40. These sites are highly conserved in mammalian species. Molecular modeling suggests that phosphorylation sites of subunit I face the inter membrane space while those of subunits IVi1 and Vb face the matrix side. PMID:17349628

  6. Novel subunit-subunit interactions in the structure of glutamine synthetase.

    PubMed

    Almassy, R J; Janson, C A; Hamlin, R; Xuong, N H; Eisenberg, D

    We present an atomic model for glutamine synthetase, an enzyme of central importance in bacterial nitrogen metabolism, from X-ray crystallography. The 12 identical subunits are arranged as the carbon atoms in two face-to-face benzene rings, with unusual subunit contacts. Our model, which places the active sites at the subunit interfaces, suggests a mechanism for the main functional role of glutamine synthetase: how the enzyme regulates the rate of synthesis of glutamine in response to covalent modification and feedback inhibition. PMID:2876389

  7. The investigation of substrate-induced changes in subunit interactions in glyceraldehyde 3-phosphate dehydrogenases by measurement of the kinetics and thermodynamics of subunit exchange.

    PubMed Central

    Osborne, H H; Hollaway, M R

    1975-01-01

    An investigation was made of changes in subunit interactions in glyceraldehyde 3-phosphate dehydrogenase on binding NAD+, NADH and other substrates by using the previously developed method of measurement of rates and extent of subunit exchange between the rabbit enzyme (R4), yeast enzyme (Y4) and rabbit-yeast hybrid (R2Y2) [Osborne & Hollaway (1974) Biochem. J. 143, 651-662]. The free energy of activation for the conversion of tetramer into dimer for the rabbit enzyme (R4 leads to 2R2) is increased by at least 12kJ/mol in the presence of NAD+. This increase is interpreted in terms of an NAD+-induced 'tightening' of the tetrameric structure probably involving increased interaction at the subunit interfaces across the QR plane of the molecule [see Buehner et al. (1974) J. Mol. Biol. 82, 563-585]. This tightening of the structure only occurs on binding the third NAD+ molecule to a given enzyme molecule. Conversely, binding of NADH causes a decrease in the free energy of activation for the R4 leads to 2R2 and Y4 leads to 2Y2 conversions by at least 10kJ/mol. This is interpreted as a NADH-induced 'loosening' of the structures arising from decreased interactions across the subunit interfaces involving the QR dissociation plane. In the presence of NADH the increase in the rate of subunit exchange is such that it is not possible to separate the hybrid from the other species if electrophoresis is carried out with NADH in the separation media. In the presence of a mixture of NADH and NAD+ the effect of NAD+ on subunit exchange is dominant. The results are discussed in terms of the known co-operativty between binding sites in glyceraldehyde 3-phosphate dehydrogenases. Images PLATE 1(a) PLATE 1(b) PLATE 2(a) PLATE 2(b) PLATE 2(c) PMID:174555

  8. Subunit dissociation in fish hemoglobins.

    PubMed

    Edelstein, S J; McEwen, B; Gibson, Q H

    1976-12-10

    The tetramer-dimer dissociation equilibria (K 4,2) of several fish hemoglobins have been examined by sedimentation velocity measurements with a scanner-computer system for the ultracentrifuge and by flash photolysis measurements using rapid kinetic methods. Samples studied in detail included hemoglobins from a marine teleost, Brevoortia tyrannus (common name, menhaden); a fresh water teleost, Cyprinus carpio, (common name, carp); and an elasmobranch Prionace glauca (common name, blue shark). For all three species in the CO form at pH 7, in 0.1 M phosphate buffer, sedimentation coefficients of 4.3 S (typical of tetrameric hemoglobin) are observed in the micromolar concentration range. In contrast, mammalian hemoglobins dissociate appreciably to dimers under these conditions. The inability to detect dissociation in three fish hemoglobins at the lowest concentrations examined indicates that K 4,2 must have a value of 10(-8) M or less. In flash photolysis experiments on very dilute solutions in long path length cells, two kinetic components were detected with their proportions varying as expected for an equilibrium between tetramers (the slower component) and dimers (the faster component); values of K 4,2 for the three fish hemoglobins in the range 10(-9) to 10(-8) M were calculated from these data. Thus, the values of K 4,2 for liganded forms of the fish hemoglobins appear to be midway between the value for liganded human hemoglobin (K 4,2 approximately 10(-6) M) and unliganded human hemoglobin (K 4,2 approximately 10(-12) M). This conclusion is supported by measurements on solutions containing guanidine hydrochloride to enhance the degree of dissociation. All three fish hemoglobins are appreciably dissociated at guanidine concentrations of about 0.8 M, which is roughly midway between the guanidine concentrations needed to cause comparable dissociation of liganded human hemoglobin (about 0.4 M) and unliganded human hemoglobin (about 1.6 M). Kinetic measurements on

  9. Flexible subunit stoichiometry of functional human P2X2/3 heteromeric receptors.

    PubMed

    Kowalski, Maria; Hausmann, Ralf; Schmid, Julia; Dopychai, Anke; Stephan, Gabriele; Tang, Yong; Schmalzing, Günther; Illes, Peter; Rubini, Patrizia

    2015-12-01

    The aim of the present work was to clarify whether heterotrimeric P2X2/3 receptors have a fixed subunit stoichiometry consisting of one P2X2 and two P2X3 subunits as previously suggested, or a flexible stoichiometry containing also the inverse subunit composition. For this purpose we transfected HEK293 cells with P2X2 and P2X3 encoding cDNA at the ratios of 1:2 and 4:1, and analysed the biophysical and pharmacological properties of the generated receptors by means of the whole-cell patch-clamp technique. The concentration-response curves for the selective agonist α,β-meATP did not differ from each other under the two transfection ratios. However, co-expression of an inactive P2X2 mutant and the wild type P2X3 subunit and vice versa resulted in characteristic distortions of the α,β-meATP concentration-response relationships, depending on which subunit was expressed in excess, suggesting that HEK293 cells express mixtures of (P2X2)1/(P2X3)2 and (P2X2)2/(P2X3)1 receptors. Whereas the allosteric modulators H+ and Zn2+ failed to discriminate between the two possible heterotrimeric receptor variants, the α,β-meATP-induced responses were blocked more potently by the competitive antagonist A317491, when the P2X2 subunit was expressed in deficit of the P2X3 subunit. Furthermore, blue-native PAGE analysis of P2X2 and P2X3 subunits co-expressed in Xenopus laevis oocytes and HEK293 cells revealed that plasma membrane-bound P2X2/3 receptors appeared in two clearly distinct heterotrimeric complexes: a (P2X2-GFP)2/(P2X3)1 complex and a (P2X2-GFP)1/(P2X3)2 complex. These data strongly indicate that the stoichiometry of the heteromeric P2X2/3 receptor is not fixed, but determined in a permutational manner by the relative availability of P2X2 and P2X3 subunits. PMID:26184350

  10. Flexible subunit stoichiometry of functional human P2X2/3 heteromeric receptors.

    PubMed

    Kowalski, Maria; Hausmann, Ralf; Schmid, Julia; Dopychai, Anke; Stephan, Gabriele; Tang, Yong; Schmalzing, Günther; Illes, Peter; Rubini, Patrizia

    2015-12-01

    The aim of the present work was to clarify whether heterotrimeric P2X2/3 receptors have a fixed subunit stoichiometry consisting of one P2X2 and two P2X3 subunits as previously suggested, or a flexible stoichiometry containing also the inverse subunit composition. For this purpose we transfected HEK293 cells with P2X2 and P2X3 encoding cDNA at the ratios of 1:2 and 4:1, and analysed the biophysical and pharmacological properties of the generated receptors by means of the whole-cell patch-clamp technique. The concentration-response curves for the selective agonist α,β-meATP did not differ from each other under the two transfection ratios. However, co-expression of an inactive P2X2 mutant and the wild type P2X3 subunit and vice versa resulted in characteristic distortions of the α,β-meATP concentration-response relationships, depending on which subunit was expressed in excess, suggesting that HEK293 cells express mixtures of (P2X2)1/(P2X3)2 and (P2X2)2/(P2X3)1 receptors. Whereas the allosteric modulators H+ and Zn2+ failed to discriminate between the two possible heterotrimeric receptor variants, the α,β-meATP-induced responses were blocked more potently by the competitive antagonist A317491, when the P2X2 subunit was expressed in deficit of the P2X3 subunit. Furthermore, blue-native PAGE analysis of P2X2 and P2X3 subunits co-expressed in Xenopus laevis oocytes and HEK293 cells revealed that plasma membrane-bound P2X2/3 receptors appeared in two clearly distinct heterotrimeric complexes: a (P2X2-GFP)2/(P2X3)1 complex and a (P2X2-GFP)1/(P2X3)2 complex. These data strongly indicate that the stoichiometry of the heteromeric P2X2/3 receptor is not fixed, but determined in a permutational manner by the relative availability of P2X2 and P2X3 subunits.

  11. Betaretroviral Envelope Subunits Are Noncovalently Associated and Restricted to the Mammalian Class

    PubMed Central

    Henzy, Jamie E.

    2013-01-01

    The structure of the transmembrane subunit (TM) of the retroviral envelope glycoprotein (Env) is highly conserved among most retrovirus genera and includes a pair of cysteines that forms an intramolecular disulfide loop within the ectodomain. Alpha-, gamma-, and deltaretroviruses have a third cysteine, adjacent to the loop, which forms a disulfide bond between TM and the surface subunit (SU) of Env, while lentiviruses, which have noncovalently associated subunits, lack this third cysteine. The Betaretrovirus genus includes Jaagsiekte sheep retrovirus (JSRV) and mouse mammary tumor virus (MMTV), as well as many endogenous retroviruses. Envelope subunit association had not been characterized in the betaretroviruses, but lack of a third cysteine in the TM ectodomain suggested noncovalently associated subunits. We tested the Env proteins of JSRV and MMTV, as well as human endogenous retrovirus K (HERV-K)108—a betaretrovirus-like human endogenous retrovirus—for intersubunit bonding and found that, as in the lentiviruses, the Env subunits lack an intersubunit disulfide bond. Since these results suggest that the number of cysteines in the TM loop region readily distinguishes between covalent and noncovalent structure, we surveyed endogenous retroviral TM sequences in the genomes of vertebrates represented in public databases and found that (i) retroviruses with noncovalently associated subunits have been present during all of anthropoid evolution and (ii) the noncovalent env motif is limited to mammals, while the covalent type is found among five vertebrate classes. We discuss implications of these findings for retroviral evolution, cross-species transmissions, and recombination events involving the env gene. PMID:23221553

  12. Foundations of Distinctive Feature Theory.

    ERIC Educational Resources Information Center

    Baltaxe, Christiane A. M.

    This treatise on the theoretical and historical foundations of distinctive feature theory traces the evolution of the distinctive features concept in the context of related notions current in linguistic theory, discusses the evolution of individual distinctive features, and criticizes certain acoustic and perceptual correlates attributed to these…

  13. Striatal α5 Nicotinic Receptor Subunit Regulates Dopamine Transmission in Dorsal Striatum

    PubMed Central

    Exley, Richard; McIntosh, J. Michael; Marks, Michael J.; Maskos, Uwe

    2012-01-01

    Polymorphisms in the gene for the α5 nicotinic acetylcholine receptor (nAChR) subunit are associated with vulnerability to nicotine addiction. However, the underlying normal functions of α5-containing nAChRs in the brain are poorly understood. Striatal dopamine (DA) transmission is critical to the acquisition and maintenance of drug addiction and is modulated strongly by nicotine acting at heteromeric β2-containing (β2*) nAChRs. We explored whether α5 subunits, as well as α4, α6, and β3 subunits, participate in the powerful regulation of DA release probability by β2* nAChRs in nucleus accumbens (NAc) core and in dorsal striatum [caudatoputamen (CPu)]. We detected evoked dopamine release using fast-scan cyclic voltammetry at carbon-fiber microelectrodes in striatal slices from mice with deletions of α4, α5, α6, or β3 subunits. We show that the nAChR subtypes that dominantly regulate dopamine transmission depend critically upon α5 subunits in the dorsal CPu in α4α5(non-α6)β2-nAChRs but not in NAc core, where α4α6β2β3-nAChRs are required. These data reveal the distinct populations of nAChRs that govern DA transmission in NAc core versus dorsal CPu. Furthermore, they indicate that α5 subunits are critical to the regulation of DA transmission by α4β2* nAChRs in regions of striatum associated with habitual and instrumental responses (dorsal CPu) rather than pavlovian associations (NAc). PMID:22396410

  14. Gonadotropin hormone and receptor sequences from model teleost species.

    PubMed

    Wong, Andrew C; Van Eenennaam, Alison L

    2004-01-01

    Fish offer some advantages for the study of vertebrate reproductive physiology. Only a few of the genes encoding the components of the hypothalamic-pituitary-gonadal axis have been identified from model teleosts. This study describes a combination of database searching and molecular approaches to identify the FSH and LH gonadotropin beta-subunits (fshb and lhb, respectively), and the LH receptor (lhr) from two model teleost species: zebrafish (Danio rerio) and Fugu (Takifugu rubripes). Sequence and phylogenetic analyses were used to examine the relationships that exist between gonadotropins and their receptors from species representing several piscine orders. The gonadotropin alpha-subunit (Cga) is highly conserved among teleosts and tetrapods. The presence of a genomic pseudogene (cgap) was also noted in zebrafish. Generally, teleostean FSHbeta protein sequences share less identity with each other than do LHbeta protein sequences, supporting the hypothesis that FSHbeta diverged more rapidly during teleost evolution. Interestingly, and uniquely, zebrafish Fshb lacked two highly conserved cysteine residues in the "determinant loop" which is thought to contribute towards receptor binding and specificity. Teleost gonadotropin receptor sequences clearly diverged into two distinct groups, FSHR and LHR. As has been seen with mammalian gonadotropin receptor transcripts, splice variants of zebrafish lhr were also observed.

  15. Expression of nicotinic acetylcholine receptor subunits from parasitic nematodes in Caenorhabditis elegans.

    PubMed

    Sloan, Megan A; Reaves, Barbara J; Maclean, Mary J; Storey, Bob E; Wolstenholme, Adrian J

    2015-11-01

    The levamisole-sensitive nicotinic acetylcholine receptor present at nematode neuromuscular junctions is composed of multiple different subunits, with the exact composition varying between species. We tested the ability of two well-conserved nicotinic receptor subunits, UNC-38 and UNC-29, from Haemonchus contortus and Ascaris suum to rescue the levamisole-resistance and locomotion defects of Caenorhabditis elegans strains with null deletion mutations in the unc-38 and unc-29 genes. The parasite cDNAs were cloned downstream of the relevant C. elegans promoters and introduced into the mutant strains via biolistic transformation. The UNC-38 subunit of H. contortus was able to completely rescue both the locomotion defects and levamisole resistance of the null deletion mutant VC2937 (ok2896), but no C. elegans expressing the A. suum UNC-38 could be detected. The H. contortus UNC-29.1 subunit partially rescued the levamisole resistance of a C. elegans null mutation in unc-29 VC1944 (ok2450), but did cause increased motility in a thrashing assay. In contrast, only a single line of worms containing the A. suum UNC-29 subunit showed a partial rescue of levamisole resistance, with no effect on thrashing.

  16. Na+/K+-ATPase α1 subunit, a novel therapeutic target for hepatocellular carcinoma

    PubMed Central

    Wang, Peng; Jiang, Yan; Yong, Pan; Zhang, Chenyue; Zhang, Haibin; Meng, Zhiqiang; Yang, Peiying

    2015-01-01

    We aimed to identify the expression patterns of Na+/K+-ATPase (NKA) α subunits in human hepatocellular carcinoma (HCC) samples and evaluate these subunits as potential targets for HCC treatment. The mRNA expression profiles of NKA α subunits in human HCC samples were analyzed. We found that the mRNA expression for NKA α1 subunit (ATP1A1) was higher than that for other NKA α subunits. Also, ATP1A1 gene expression was markedly higher in HCC samples than in adjacent nontumor tissue samples. Western blotting data suggested that 6 of 14 (43%) HCC samples had elevated ATP1A1 protein expression. Furthermore, knockdown of ATP1A1 expression in human HCC HepG2 and MHCC97H cells markedly reduced their proliferation in vitro and suppressed the tumorigenicity of MHCC97H cells in vivo. Downregulation of ATP1A1 expression resulted in cell-cycle arrest at G2/M phase and apoptosis in HepG2 cells as well as decreased migration in Hep3B cells. We further validated that ATP1A1 downregulation caused intracellular accumulation of reactive oxygen species. Pretreatment with N-acetyl cysteine blocked cell-growth inhibition induced by ATP1A1 downregulation. Collectively, these data suggested that targeting ATP1A1 is a novel approach to the treatment of HCC. PMID:26334094

  17. Na+/K+-ATPase α1 subunit, a novel therapeutic target for hepatocellular carcinoma.

    PubMed

    Zhuang, Liping; Xu, Litao; Wang, Peng; Jiang, Yan; Yong, Pan; Zhang, Chenyue; Zhang, Haibin; Meng, Zhiqiang; Yang, Peiying

    2015-09-29

    We aimed to identify the expression patterns of Na+/K+-ATPase (NKA) α subunits in human hepatocellular carcinoma (HCC) samples and evaluate these subunits as potential targets for HCC treatment. The mRNA expression profiles of NKA α subunits in human HCC samples were analyzed. We found that the mRNA expression for NKA α1 subunit (ATP1A1) was higher than that for other NKA α subunits. Also, ATP1A1 gene expression was markedly higher in HCC samples than in adjacent nontumor tissue samples. Western blotting data suggested that 6 of 14 (43%) HCC samples had elevated ATP1A1 protein expression. Furthermore, knockdown of ATP1A1 expression in human HCC HepG2 and MHCC97H cells markedly reduced their proliferation in vitro and suppressed the tumorigenicity of MHCC97H cells in vivo. Downregulation of ATP1A1 expression resulted in cell-cycle arrest at G2/M phase and apoptosis in HepG2 cells as well as decreased migration in Hep3B cells. We further validated that ATP1A1 downregulation caused intracellular accumulation of reactive oxygen species. Pretreatment with N-acetyl cysteine blocked cell-growth inhibition induced by ATP1A1 downregulation. Collectively, these data suggested that targeting ATP1A1 is a novel approach to the treatment of HCC.

  18. Na+/K+-ATPase α1 subunit, a novel therapeutic target for hepatocellular carcinoma.

    PubMed

    Zhuang, Liping; Xu, Litao; Wang, Peng; Jiang, Yan; Yong, Pan; Zhang, Chenyue; Zhang, Haibin; Meng, Zhiqiang; Yang, Peiying

    2015-09-29

    We aimed to identify the expression patterns of Na+/K+-ATPase (NKA) α subunits in human hepatocellular carcinoma (HCC) samples and evaluate these subunits as potential targets for HCC treatment. The mRNA expression profiles of NKA α subunits in human HCC samples were analyzed. We found that the mRNA expression for NKA α1 subunit (ATP1A1) was higher than that for other NKA α subunits. Also, ATP1A1 gene expression was markedly higher in HCC samples than in adjacent nontumor tissue samples. Western blotting data suggested that 6 of 14 (43%) HCC samples had elevated ATP1A1 protein expression. Furthermore, knockdown of ATP1A1 expression in human HCC HepG2 and MHCC97H cells markedly reduced their proliferation in vitro and suppressed the tumorigenicity of MHCC97H cells in vivo. Downregulation of ATP1A1 expression resulted in cell-cycle arrest at G2/M phase and apoptosis in HepG2 cells as well as decreased migration in Hep3B cells. We further validated that ATP1A1 downregulation caused intracellular accumulation of reactive oxygen species. Pretreatment with N-acetyl cysteine blocked cell-growth inhibition induced by ATP1A1 downregulation. Collectively, these data suggested that targeting ATP1A1 is a novel approach to the treatment of HCC. PMID:26334094

  19. Reconfiguration of yeast 40S ribosomal subunit domains by the translation initiation multifactor complex.

    PubMed

    Gilbert, Robert J C; Gordiyenko, Yulya; von der Haar, Tobias; Sonnen, Andreas F-P; Hofmann, Gregor; Nardelli, Maria; Stuart, David I; McCarthy, John E G

    2007-04-01

    In the process of protein synthesis, the small (40S) subunit of the eukaryotic ribosome is recruited to the capped 5' end of the mRNA, from which point it scans along the 5' untranslated region in search of a start codon. However, the 40S subunit alone is not capable of functional association with cellular mRNA species; it has to be prepared for the recruitment and scanning steps by interactions with a group of eukaryotic initiation factors (eIFs). In budding yeast, an important subset of these factors (1, 2, 3, and 5) can form a multifactor complex (MFC). Here, we describe cryo-EM reconstructions of the 40S subunit, of the MFC, and of 40S complexes with MFC factors plus eIF1A. These studies reveal the positioning of the core MFC on the 40S subunit, and show how eIF-binding induces mobility in the head and platform and reconfigures the head-platform-body relationship. This is expected to increase the accessibility of the mRNA channel, thus enabling the 40S subunit to convert to a recruitment-competent state.

  20. Genetic Diversity within Schistosoma haematobium: DNA Barcoding Reveals Two Distinct Groups

    PubMed Central

    Webster, Bonnie L.; Emery, Aiden M.; Webster, Joanne P.; Gouvras, Anouk; Garba, Amadou; Diaw, Oumar; Seye, Mohmoudane M.; Tchuente, Louis Albert Tchuem; Simoonga, Christopher; Mwanga, Joseph; Lange, Charles; Kariuki, Curtis; Mohammed, Khalfan A.; Stothard, J. Russell; Rollinson, David

    2012-01-01

    Background Schistosomiasis in one of the most prevalent parasitic diseases, affecting millions of people and animals in developing countries. Amongst the human-infective species S. haematobium is one of the most widespread causing urogenital schistosomiasis, a major human health problem across Africa, however in terms of research this human pathogen has been severely neglected. Methodology/Principal Findings To elucidate the genetic diversity of Schistosoma haematobium, a DNA ‘barcoding’ study was performed on parasite material collected from 41 localities representing 18 countries across Africa and the Indian Ocean Islands. Surprisingly low sequence variation was found within the mitochondrial cytochrome oxidase subunit I (cox1) and the NADH-dehydrogenase subunit 1 snad1). The 61 haplotypes found within 1978 individual samples split into two distinct groups; one (Group 1) that is predominately made up of parasites from the African mainland and the other (Group 2) that is made up of samples exclusively from the Indian Ocean Islands and the neighbouring African coastal regions. Within Group 1 there was a dominance of one particular haplotype (H1) representing 1574 (80%) of the samples analyzed. Population genetic diversity increased in samples collected from the East African coastal regions and the data suggest that there has been movement of parasites between these areas and the Indian Ocean Islands. Conclusions/Significance The high occurrence of the haplotype (H1) suggests that at some point in the recent evolutionary history of S. haematobium in Africa the population may have passed through a genetic ‘bottleneck’ followed by a population expansion. This study provides novel and extremely interesting insights into the population genetics of S. haematobium on a large geographic scale, which may have consequence for control and monitoring of urogenital schistosomiasis. PMID:23145200

  1. Distinct functions of neuromedin u and neuromedin s in orange-spotted grouper.

    PubMed

    Li, Shuisheng; Xiao, Ling; Liu, Qiongyu; Zheng, Binbin; Chen, Huapu; Liu, Xiaochun; Zhang, Yong; Lin, Haoran

    2015-10-01

    Neuromedin U (NMU) and neuromedin S (NMS) play inhibitory roles in the regulation of food intake and energy homeostasis in mammals. However, their functions are not clearly established in teleost fish. In the present study, nmu and nms homologs were identified in several fish species. Subsequently, their cDNA sequences were cloned from the orange-spotted grouper (Epinephelus coioides). Sequence analysis showed that the orange-spotted grouper Nmu proprotein contains a 21-amino acid mature Nmu peptide (Nmu-21). The Nms proprotein lost the typical mature Nms peptide, but it retains a putative 34-amino acid peptide (Nmsrp). In situ hybridization revealed that nmu- and nms-expressing cells are mainly localized in the hypothalamic regions associated with appetite regulation. Food deprivation decreased the hypothalamic nmu mRNA levels but induced an increase of nms mRNA levels. Periprandial expression analysis showed that hypothalamic expression of nmu increased significantly at 3 h post-feeding, while nms expression was elevated at the normal feeding time. I.p. injection of synthetic Nmu-21 peptide suppressed the hypothalamic neuropeptide y (npy) expression, while Nmsrp administration significantly increased the expression of npy and orexin in orange-spotted grouper. Furthermore, the mRNA levels of LH beta subunit (lhβ) and gh in the pituitary were significantly down-regulated after Nmu-21 peptide administration, while Nmsrp was able to significantly stimulate the expression of FSH beta subunit (fshβ), prolactin (prl), and somatolaction (sl). Our results indicate that nmu and nms possess distinct neuroendocrine functions and pituitary functions in the orange spotted grouper.

  2. Studies of double-labeled mouse thyrotropin and free alpha-subunits to estimate relative fucose content

    SciTech Connect

    Magner, J.; Papagiannes, E.

    1986-11-01

    The composition and structure of the complex oligosaccharides of thyrotropin (TSH) and free alpha-subunits are not well established, but are believed to be important determinants of the biological properties of these glycoproteins. We employed a simple double-label technique to learn the relative fucose content of mouse thyrotropin and free alpha-subunits. Thyrotropic tumor minces were incubated simultaneously with (/sup 35/S)methionine and (/sup 3/H)fucose. Thyrotropin and free alpha-subunits were labeled with both isotopes, and the ratio of /sup 3/H//sup 35/S was higher in free alpha-subunits than in thyrotropin; free alpha-subunits were approximately fivefold richer in fucose than was thyrotropin. The /sup 3/H//sup 35/S ratio was not substantially altered in TSH or free alpha-subunits secreted after a brief incubation with 10(-7) M thyrotropin-releasing hormone. Species which incorporated (/sup 3/H)fucose were resistant to endoglycosidase H. Thus, mouse free alpha-subunits secreted by thyrotropic tumor are relatively rich in fucose. Double-isotope labeling using an amino acid and a sugar appears to be a useful technique for studies of the glycoprotein hormones.

  3. Examination of the Montastraea annularis Species Complex (Cnidaria: Scleractinia) Using ITS and COI Sequences.

    PubMed

    Medina; Weil; Szmant

    1999-01-01

    : The Caribbean coral Montastraea annularis has recently been proposed to be a complex of at least three sibling species. To test the validity of this proposal, we sequenced the ITS region of the nuclear ribosomal RNA gene family (ITS-1, 5.8S, and ITS-2), and a portion of the mitochondrial DNA gene cytochrome c oxidase subunit I (COI) from the three proposed species (M. annularis, M. faveolata, and M. franksi) from Florida reefs. The ITS fragment was 665 nucleotides long and had 19 variable sites, of which 6 were parsimony-informative sites. None of these sites was fixed within the proposed species. The COI fragment was 658 nucleotides long with only two sites variable in one individual. Thus, under both the biological species concept and the phylogenetic species concept, the molecular evidence gathered in this study indicates the Montastraea annularis species complex to be a single evolutionary entity as opposed to three distinct species. The three proposed Montastraea species can interbreed, ruling out prezygotic barriers to gene flow (biological species concept), and the criterion of monophyly is not satisfied if hybridization is occurring among taxa (phylogenetic species concept).

  4. Isolation of the alpha subunits of GTP-binding regulatory proteins by affinity chromatography with immobilized beta gamma subunits.

    PubMed Central

    Pang, I H; Sternweis, P C

    1989-01-01

    Immobilized beta gamma subunits of GTP-binding regulatory proteins (G proteins) were used to isolate alpha subunits from solubilized membranes of bovine tissues and to separate specific alpha subunits based on their differential affinities for beta gamma subunits. The beta gamma subunits were cross-linked to omega-aminobutyl agarose. Up to 7 nmol of alpha subunit could bind to each milliliter of beta gamma-agarose and be recovered by elution with AIF4-. This affinity resin effectively separated the alpha subunits of Gi1 and Gi2 from "contaminating" alpha subunits of Go, the most abundant G protein in bovine brain, by taking advantage of the apparent lower affinity of the alpha subunits of Go for beta gamma subunits. The beta gamma-agarose was also used to isolate mixtures of alpha subunits from cholate extracts of membranes from different bovine tissues. alpha subunits of 39-41 kDa (in various ratios) as well as the alpha subunits of Gs were purified. The yields from extracts exceeded 60% for all alpha subunits examined and apparently represented the relative content of alpha subunits in the tissues. This technique can rapidly isolate and identify, from a small amount of sample, the endogenous G proteins in various tissues and cells. So far, only polypeptides in the range of 39-52 kDa have been detected with this approach. If other GTP-binding proteins interact with these beta gamma subunits, the interaction is either of low affinity or mechanistically unique from the alpha subunits isolated in this study. Images PMID:2510152

  5. Dissociation of ribosomes into large and small subunits.

    PubMed

    Rivera, Maria C; Maguire, Bruce; Lake, James A

    2015-04-01

    Structural and functional studies of ribosomal subunits require the dissociation of intact ribosomes into individual small and large ribosomal subunits. The dissociation of the prokaryotic 70S ribosomes into the 50S and 30S subunits is achieved by dialysis against a buffer containing a lower Mg(2+) concentration. Eukaryotic 80S ribosomes are dissociated into 60S and 40S subunits by incubation in a buffer containing puromycin and higher KCl and Mg(2+) concentrations.

  6. Wickerhamomyces mori sp. nov., an anamorphic yeast species found in the guts of wood-boring insect larvae.

    PubMed

    Hui, Feng-Li; Chen, Liang; Chu, Xue-Ying; Niu, Qiu-Hong; Ke, Tao

    2013-03-01

    A novel anamorphic yeast species is described to accommodate three isolates recovered from the guts of three different wood-boring insect larvae collected in Henan, central China. On the basis of sequence analyses of the D1/D2 domains of the large-subunit rRNA gene and the internal transcribed spacer regions, the three strains are assigned to a novel species of the genus Wickerhamomyces, although the formation of ascospores was not observed. These strains also exhibited a number of distinct morphological and physiological characteristics that clearly differentiated them from Wickerhamomyces mucosus, Candida odintsovae and Wickerhamomyces rabaulensis, the most closely related species. In view of the phenotypic differences and unique rRNA gene sequences, we consider that these three isolates represent a novel species of the genus Wickerhamomyces, Wickerhamomyces mori sp. nov. The type strain is NYNU 1216(T) ( = CICC 1983(T)  = CBS 12678(T)).

  7. Serological reactivity of patients with Echinococcus infections (E. granulosus, E. vogeli, and E. multilocularis) against three antigen B subunits.

    PubMed

    de la Rue, Mário L; Yamano, Kimiaki; Almeida, Cybele E; Iesbich, Margarete P; Fernandes, Cloé D; Goto, Akiko; Kouguchi, Hirokazu; Takahashi, Kenichi

    2010-02-01

    In serodiagnosis of cystic echinococcosis (CE) by Echinococcus granulosus infection, antigen B (AgB) has been utilized worldwide. However, it is known that about 40% of sera with alveolar echinococcosis (AE) by Echinococcus multilocularis infection recognize AgB. Furthermore, cross-reaction against AgB was also reported in sera from polycystic echinococcosis (PE) patients with Echinococcus vogeli infection. These findings indicate that AgB is widely common to the genus Echinococcus. On the other hand, AgB has several subunits, which are composed of the smallest 8-kDa subunit. In this study, reactivities of patient sera with three kinds of Echinococcus infections (CE, PE, and AE) were compared simultaneously under the same condition against three subunits of AgB (8, 16, and 24 kDa). Many articles have referred the fundamental 8- kDa subunit as a diagnostic antigen for CE. However, the reactivity for the 8-kDa subunit of the CE patient was not so high (47.7%) in this study. Furthermore, there are many cases in which serum of patients with PE or AE also recognizes this subunit (66.7% in PE; 45.9% in AE). AgB is effective for the detection of the genus Echinococcus infections, but it does not have high species specificity. Therefore, we need to pay attention to cross-reaction in serodiagnosis of CE in areas where plural species coexist.

  8. Evolutionary analyses of the 12-kDa acidic ribosomal P-proteins reveal a distinct protein of higher plant ribosomes

    PubMed Central

    Szick, Kathleen; Springer, Mark; Bailey-Serres, Julia

    1998-01-01

    The P-protein complex of eukaryotic ribosomes forms a lateral stalk structure in the active site of the large ribosomal subunit and is thought to assist in the elongation phase of translation by stimulating GTPase activity of elongation factor-2 and removal of deacylated tRNA. The complex in animals, fungi, and protozoans is composed of the acidic phosphoproteins P0 (35 kDa), P1 (11–12 kDa), and P2 (11–12 kDa). Previously we demonstrated by protein purification and microsequencing that ribosomes of maize (Zea mays L.) contain P0, one type of P1, two types of P2, and a distinct P1/P2 type protein designated P3. Here we implemented distance matrices, maximum parsimony, and neighbor-joining analyses to assess the evolutionary relationships between the 12 kDa P-proteins of maize and representative eukaryotic species. The analyses identify P3, found to date only in mono- and dicotyledonous plants, as an evolutionarily distinct P-protein. Plants possess three distinct groups of 12 kDa P-proteins (P1, P2, and P3), whereas animals, fungi, and protozoans possess only two distinct groups (P1 and P2). These findings demonstrate that the P-protein complex has evolved into a highly divergent complex with respect to protein composition despite its critical position within the active site of the ribosome. PMID:9482893

  9. Recent Advances in Subunit Vaccine Carriers

    PubMed Central

    Vartak, Abhishek; Sucheck, Steven J.

    2016-01-01

    The lower immunogenicity of synthetic subunit antigens, compared to live attenuated vaccines, is being addressed with improved vaccine carriers. Recent reports indicate that the physio-chemical properties of these carriers can be altered to achieve optimal antigen presentation, endosomal escape, particle bio-distribution, and cellular trafficking. The carriers can be modified with various antigens and ligands for dendritic cells targeting. They can also be modified with adjuvants, either covalently or entrapped in the matrix, to improve cellular and humoral immune responses against the antigen. As a result, these multi-functional carrier systems are being explored for use in active immunotherapy against cancer and infectious diseases. Advancing technology, improved analytical methods, and use of computational methodology have also contributed to the development of subunit vaccine carriers. This review details recent breakthroughs in the design of nano-particulate vaccine carriers, including liposomes, polymeric nanoparticles, and inorganic nanoparticles. PMID:27104575

  10. PKA regulatory subunit expression in tooth development.

    PubMed

    de Sousa, Sílvia Ferreira; Kawasaki, Katsushige; Kawasaki, Maiko; Volponi, Ana Angelova; Gomez, Ricardo Santiago; Gomes, Carolina Cavaliéri; Sharpe, Paul T; Ohazama, Atsushi

    2014-05-01

    Protein kinase A (PKA) plays critical roles in many biological processes including cell proliferation, cell differentiation, cellular metabolism and gene regulation. Mutation in PKA regulatory subunit, PRKAR1A has previously been identified in odontogenic myxomas, but it is unclear whether PKA is involved in tooth development. The aim of the present study was to assess the expression of alpha isoforms of PKA regulatory subunit (Prkar1a and Prkar2a) in mouse and human odontogenesis by in situ hybridization. PRKAR1A and PRKAR2A mRNA transcription was further confirmed in a human deciduous germ by qRT-PCR. Mouse Prkar1a and human PRKAR2A exhibited a dynamic spatio-temporal expression in tooth development, whereas neither human PRKAR1A nor mouse Prkar2a showed their expression in odontogenesis. These isoforms thus showed different expression pattern between human and mouse tooth germs. PMID:24755349

  11. Recent Advances in Subunit Vaccine Carriers.

    PubMed

    Vartak, Abhishek; Sucheck, Steven J

    2016-01-01

    The lower immunogenicity of synthetic subunit antigens, compared to live attenuated vaccines, is being addressed with improved vaccine carriers. Recent reports indicate that the physio-chemical properties of these carriers can be altered to achieve optimal antigen presentation, endosomal escape, particle bio-distribution, and cellular trafficking. The carriers can be modified with various antigens and ligands for dendritic cells targeting. They can also be modified with adjuvants, either covalently or entrapped in the matrix, to improve cellular and humoral immune responses against the antigen. As a result, these multi-functional carrier systems are being explored for use in active immunotherapy against cancer and infectious diseases. Advancing technology, improved analytical methods, and use of computational methodology have also contributed to the development of subunit vaccine carriers. This review details recent breakthroughs in the design of nano-particulate vaccine carriers, including liposomes, polymeric nanoparticles, and inorganic nanoparticles. PMID:27104575

  12. PKA regulatory subunit expression in tooth development.

    PubMed

    de Sousa, Sílvia Ferreira; Kawasaki, Katsushige; Kawasaki, Maiko; Volponi, Ana Angelova; Gomez, Ricardo Santiago; Gomes, Carolina Cavaliéri; Sharpe, Paul T; Ohazama, Atsushi

    2014-05-01

    Protein kinase A (PKA) plays critical roles in many biological processes including cell proliferation, cell differentiation, cellular metabolism and gene regulation. Mutation in PKA regulatory subunit, PRKAR1A has previously been identified in odontogenic myxomas, but it is unclear whether PKA is involved in tooth development. The aim of the present study was to assess the expression of alpha isoforms of PKA regulatory subunit (Prkar1a and Prkar2a) in mouse and human odontogenesis by in situ hybridization. PRKAR1A and PRKAR2A mRNA transcription was further confirmed in a human deciduous germ by qRT-PCR. Mouse Prkar1a and human PRKAR2A exhibited a dynamic spatio-temporal expression in tooth development, whereas neither human PRKAR1A nor mouse Prkar2a showed their expression in odontogenesis. These isoforms thus showed different expression pattern between human and mouse tooth germs.

  13. Structure–Function Relationships in Fungal Large-Subunit Catalases

    SciTech Connect

    Diaz, A.; Valdez, V; Rudino-Pinera, E; Horjales, E; Hansberg, W

    2009-01-01

    Neurospora crassa has two large-subunit catalases, CAT-1 and CAT-3. CAT-1 is associated with non-growing cells and accumulates particularly in asexual spores; CAT-3 is associated with growing cells and is induced under different stress conditions. It is our interest to elucidate the structure-function relationships in large-subunit catalases. Here we have determined the CAT-3 crystal structure and compared it with the previously determined CAT-1 structure. Similar to CAT-1, CAT-3 hydrogen peroxide (H{sub 2}O{sub 2}) saturation kinetics exhibited two components, consistent with the existence of two active sites: one saturated in the millimolar range and the other in the molar range. In the CAT-1 structure, we found three interesting features related to its unusual kinetics: (a) a constriction in the channel that conveys H{sub 2}O{sub 2} to the active site; (b) a covalent bond between the tyrosine, which forms the fifth coordination bound to the iron of the heme, and a vicinal cysteine; (c) oxidation of the pyrrole ring III to form a cis-hydroxyl group in C5 and a cis-{gamma}-spirolactone in C6. The site of heme oxidation marks the starts of the central channel that communicates to the central cavity and the shortest way products can exit the active site. CAT-3 has a similar constriction in its major channel, which could function as a gating system regulated by the H{sub 2}O{sub 2} concentration before the gate. CAT-3 functional tyrosine is not covalently bonded, but has instead the electron relay mechanism described for the human catalase to divert electrons from it. Pyrrole ring III in CAT-3 is not oxidized as it is in other large-subunit catalases whose structure has been determined. Different in CAT-3 from these enzymes is an occupied central cavity. Results presented here indicate that CAT-3 and CAT-1 enzymes represent a functional group of catalases with distinctive structural characteristics that determine similar kinetics.

  14. Fusarium foetens, a new species pathogenic to begonia elatior hybrids (Begonia x hiemalis) and the sister taxon of the Fusarium oxysporum species complex.

    PubMed

    Schroers, H-J; Baayen, R P; Meffert, J P; de Gruyter, J; Hooftman, M; O'Donnell, K

    2004-01-01

    A new disease recently was discovered in begonia elatior hybrid (Begonia × hiemalis) nurseries in The Netherlands. Diseased plants showed a combination of basal rot, vein yellowing and wilting and the base of collapsing plants was covered by unusually large masses of Fusarium macroconidia. A species of Fusarium was isolated consistently from the discolored veins of leaves and stems. It differed morphologically from F. begoniae, a known agent of begonia flower, leaf and stem blight. The Fusarium species resembled members of the F. oxysporum species complex in producing short monophialides on the aerial mycelium and abundant chlamydospores. Other phenotypic characters such as polyphialides formed occasionally in at least some strains, relatively long monophialides intermingled with the short monophialides formed on the aerial mycelium, distinct sporodochial conidiomata, and distinct pungent colony odor distinguished it from the F. oxysporum species complex. Phylogenetic analyses of partial sequences of the mitochondrial small subunit of the ribosomal DNA (mtSSU rDNA), nuclear translation elongation factor 1α (EF-1α) and β-tubulin gene exons and introns indicate that the Fusarium species represents a sister group of the F. oxysporum species complex. Begonia × hiemalis cultivars Bazan, Bellona and Netja Dark proved to be highly susceptible to the new species. Inoculated plants developed tracheomycosis within 4 wk, and most died within 8 wk. The new taxon was not pathogenic to Euphorbia pulcherrima, Impatiens walleriana and Saintpaulia ionantha that commonly are grown in nurseries along with B. × hiemalis. Inoculated plants of Cyclamen persicum did not develop the disease but had discolored vessels from which the inoculated fungus was isolated. Given that the newly discovered begonia pathogen is distinct in pathogenicity, morphology and phylogeny from other fusaria, it is described here as a new species, Fusarium foetens. PMID:21148861

  15. CCTα and CCTδ Chaperonin Subunits Are Essential and Required for Cilia Assembly and Maintenance in Tetrahymena

    PubMed Central

    Seixas, Cecilia; Cruto, Teresa; Tavares, Alexandra; Gaertig, Jacek; Soares, Helena

    2010-01-01

    Background The eukaryotic cytosolic chaperonin CCT is a hetero-oligomeric complex formed by two rings connected back-to-back, each composed of eight distinct subunits (CCTα to CCTζ). CCT complex mediates the folding, of a wide range of newly synthesised proteins including tubulin (α, β and γ) and actin, as quantitatively major substrates. Methodology/Principal Findings We disrupted the genes encoding CCTα and CCTδ subunits in the ciliate Tetrahymena. Cells lacking the zygotic expression of either CCTα or CCTδ showed a loss of cell body microtubules, failed to assemble new cilia and died within 2 cell cycles. We also show that loss of CCT subunit activity leads to axoneme shortening and splaying of tips of axonemal microtubules. An epitope-tagged CCTα rescued the gene knockout phenotype and localized primarily to the tips of cilia. A mutation in CCTα, G346E, at a residue also present in the related protein implicated in the Bardet Biedel Syndrome, BBS6, also caused defects in cilia and impaired CCTα localization in cilia. Conclusions/Significance Our results demonstrate that the CCT subunits are essential and required for ciliary assembly and maintenance of axoneme structure, especially at the tips of cilia. PMID:20502701

  16. Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions

    PubMed Central

    Wasserman, Michael R.; Pulk, Arto; Zhou, Zhou; Altman, Roger B.; Zinder, John C.; Green, Keith D.; Garneau-Tsodikova, Sylvie; Doudna Cate, Jamie H.; Blanchard, Scott C.

    2015-01-01

    Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin—paromomycin, ribostamycin and neamine—each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6′-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6′-substituent and the drug's net positive charge. By solving the crystal structure of the paromomycin–ribosome complex, we observe specific contacts between the apical tip of H69 and the 6′-hydroxyl on paromomycin from within the drug's canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation. PMID:26224058

  17. Solution structure of the NDH-1 complex subunit CupS from Thermosynechococcus elongatus.

    PubMed

    Korste, Annika; Wulfhorst, Hannes; Ikegami, Takahisa; Nowaczyk, Marc M; Stoll, Raphael

    2015-10-01

    The cyanobacterial multi-subunit membrane protein complex NDH-1 is both structurally and functionally related to Complex I of eubacteria and mitochondria. In addition to functions in respiration and cyclic electron transfer around photosystem I (PSI), the cyanobacterial NDH-1 complex is involved in a unique mechanism for inorganic carbon concentration. Although the crystal structures of the similar respiratory Complex I from Thermus thermophilus and Escherichia coli are known, atomic structural information is not available for the cyanobacterial NDH-1 complex yet. In particular, the structures of those subunits that are not homologous to Complex I will help to understand their distinct functions. The 15.7kDa protein CupS is a small soluble subunit of the complex variant NDH-1MS, which is thought to play a role in CO2 conversion. Here, we present the NMR structure of CupS from Thermosynechococcus elongatus, which is the very first structure of a specific cyanobacterial NDH-1 complex subunit. CupS shares a structural similarity with members of the Fasciclin protein superfamily. The structural comparison to Fasciclin type proteins based on known NMR structures and protein sequences of human TGFBIp, MPB70 from Mycobacterium bovis, and Fdp from Rhodobacter sphaeroides, together with a virtual docking model of CupS and NdhF3, provide first insight into the specific binding of CupS to the NDH-1MS complex at atomic resolution.

  18. Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions.

    PubMed

    Wasserman, Michael R; Pulk, Arto; Zhou, Zhou; Altman, Roger B; Zinder, John C; Green, Keith D; Garneau-Tsodikova, Sylvie; Cate, Jamie H Doudna; Blanchard, Scott C

    2015-01-01

    Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin-paromomycin, ribostamycin and neamine-each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6'-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6'-substituent and the drug's net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6'-hydroxyl on paromomycin from within the drug's canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation. PMID:26224058

  19. Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions

    NASA Astrophysics Data System (ADS)

    Wasserman, Michael R.; Pulk, Arto; Zhou, Zhou; Altman, Roger B.; Zinder, John C.; Green, Keith D.; Garneau-Tsodikova, Sylvie; Doudna Cate, Jamie H.; Blanchard, Scott C.

    2015-07-01

    Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin--paromomycin, ribostamycin and neamine--each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6'-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6'-substituent and the drug's net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6'-hydroxyl on paromomycin from within the drug's canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation.

  20. Differential distribution of calpain small subunit 1 and 2 in rat brain.

    PubMed

    Friedrich, Peter; Papp, Henrietta; Halasy, Katalin; Farkas, Attila; Farkas, Bence; Tompa, Peter; Kása, Peter

    2004-04-01

    Calpains, the Ca(2+)-dependent thiol proteases, are abundant in the nervous tissue. The ubiquitous enzyme forms in mammals are heterodimers consisting of a specific, micro or m, large (catalytic) subunit and, apparently, a common small (regulatory) subunit (CSS1). Recently, however, we described a second form of small subunit (CSS2), which is of restricted occurrence [Schád, E., Farkas, A., Jékely, G., Tompa, P. & Friedrich, P. (2002) Biochem. J., 362, 383-388]. Here we analysed the distribution of immunoreactivity in various parts of rat brain against two anti-CSS1 and two anti-CSS2 antibodies by correlated light and electron microscopy. Remarkably, the antibodies showed differential distribution in various parts of rat cortex: anti-CSS1 reacted mainly with perikarya and dendrites, whereas anti-CSS2 was more prominent in axons. In serial sections CSS2 and synaptophysin gave very similar patterns, i.e. these epitopes seem to colocalize. Electron microscopy confirmed that CSS1 was mainly localized postsynaptically in dendrites and somata, whereas CSS2 was found presynaptically. The hypothesis is advanced that these distinct distributions of calpain subunits may be related to the transport of these enzymes in nerve cells. PMID:15078555

  1. The catalytic and the RNA subunits of human telomerase are required to immortalize equid primary fibroblasts.

    PubMed

    Vidale, Pamela; Magnani, Elisa; Nergadze, Solomon G; Santagostino, Marco; Cristofari, Gael; Smirnova, Alexandra; Mondello, Chiara; Giulotto, Elena

    2012-10-01

    Many human primary somatic cells can be immortalized by inducing telomerase activity through the exogenous expression of the human telomerase catalytic subunit (hTERT). This approach has been extended to the immortalization of cell lines from several mammals. Here, we show that hTERT expression is not sufficient to immortalize primary fibroblasts from three equid species, namely donkey, Burchelli's zebra and Grevy's zebra. In vitro analysis of a reconstituted telomerase composed by hTERT and an equid RNA component of telomerase (TERC) revealed a low activity of this enzyme compared to human telomerase, suggesting a low compatibility of equid and human telomerase subunits. This conclusion was also strengthened by comparison of human and equid TERC sequences, which revealed nucleotide differences in key regions for TERC and TERT interaction. We then succeeded in immortalizing equid fibroblasts by expressing hTERT and hTERC concomitantly. Expression of both human telomerase subunits led to telomerase activity and telomere elongation, indicating that human telomerase is compatible with the other equid telomerase subunits and proteins involved in telomere metabolism. The immortalization procedure described herein could be extended to primary cells from other mammals. The availability of immortal cells from endangered species could be particularly useful for obtaining new information on the organization and function of their genomes, which is relevant for their preservation.

  2. Arrangement of subunits in microribbons from Giardia.

    PubMed

    Holberton, D V

    1981-02-01

    Ultrasound has been used to disperse the cytoplasm of Giardia muris and Giardia duodenalis trophozoites, releasing disk cytoskeletons for negative staining and study by electron microscopy. Sonication also breaks down the corss-bridges uniting microribbons in disks. Individual ribbons and small bundles of these structures, are found in these preparations and have been imaged both from their edges and in flat face view. The outer layers of ribbons are 2 sheets of regularly arranged globular subunits, held apart by a fibrous inner core. The axial repeat of the microribbon is 15 nm, which is also the distance separating cross-bridge sites along ribbons. Pronounced striping at this interval is a feature of ribbon faces where they are joined in bundles. Subunits in the outer layer are arranged in vertical protofilaments that are set orthogonally to the long axis of the ribbon. Protofilaments bind tannic acid and are seen clearly in sectioned ribbons. Three protofilaments fit into the 15-nm longitudinal spacing. Optical diffraction patterns from ribbon images are dominated by orders of the 15-nm periodicity, including the third-order reflexions expected from protofilaments spacings. Fourth-order reflexions indicate that the ribbon core may also be structured. Ribbon face images give rise to a strong 4-nm layer line, corresponding to the vertical spacing of subunits in protofilaments. Neighbouring protofilaments are staggered by about 0.67 nm. The lattices found in ribbons are consistent with studies of cytoskeleton composition.

  3. Type B Heterotrimeric G Protein γ-Subunit Regulates Auxin and ABA Signaling in Tomato[OPEN

    PubMed Central

    Subramaniam, Gayathery; Trusov, Yuri; Hayashi, Satomi; Batley, Jacqueline

    2016-01-01

    Heterotrimeric G proteins composed of α, β, and γ subunits are central signal transducers mediating the cellular response to multiple stimuli in most eukaryotes. Gγ subunits provide proper cellular localization and functional specificity to the heterotrimer complex. Plant Gγ subunits, divided into three structurally distinct types, are more diverse than their animal counterparts. Type B Gγ subunits, lacking a carboxyl-terminal isoprenylation motif, are found only in flowering plants. We present the functional characterization of type B Gγ subunit (SlGGB1) in tomato (Solanum lycopersicum). We show that SlGGB1 is the most abundant Gγ subunit in tomato and strongly interacts with the Gβ subunit. Importantly, the green fluorescent protein-SlGGB1 fusion protein as well as the carboxyl-terminal yellow fluorescent protein-SlGGB1/amino-terminal yellow fluorescent protein-Gβ heterodimer were localized in the plasma membrane, nucleus, and cytoplasm. RNA interference-mediated silencing of SlGGB1 resulted in smaller seeds, higher number of lateral roots, and pointy fruits. The silenced lines were hypersensitive to exogenous auxin, while levels of endogenous auxins were lower or similar to those of the wild type. SlGGB1-silenced plants also showed strong hyposensitivity to abscisic acid (ABA) during seed germination but not in other related assays. Transcriptome analysis of the transgenic seeds revealed abnormal expression of genes involved in ABA sensing, signaling, and response. We conclude that the type B Gγ subunit SlGGB1 mediates auxin and ABA signaling in tomato. PMID:26668332

  4. DNA sequences, recombinant DNA molecules and processes for producing the A and B subunits of cholera toxin and preparations containing so-obtained subunit or subunits

    SciTech Connect

    Harford, N.; De Wilde, M.

    1987-05-19

    A recombinant DNA molecule is described comprising at least a portion coding for subunits A and B of cholera toxin, or a fragment or derivative of the portion wherein the fragment or derivative codes for a polypeptide have an activity which can induce an immune response to subunit A; can induce an immune response to subunit A and cause epithelial cell penetration and the enzymatic effect leading to net loss of fluid into the gut lumen; can bind to the membrane receptor for the B subunit of cholera toxin; can induce an immune response to subunit B; can induce an immune response to subunit B and bind to the membrane receptor; or has a combination of the activities.

  5. Na+ Channel β Subunits: Overachievers of the Ion Channel Family

    PubMed Central

    Brackenbury, William J.; Isom, Lori L.

    2011-01-01

    Voltage-gated Na+ channels (VGSCs) in mammals contain a pore-forming α subunit and one or more β subunits. There are five mammalian β subunits in total: β1, β1B, β2, β3, and β4, encoded by four genes: SCN1B–SCN4B. With the exception of the SCN1B splice variant, β1B, the β subunits are type I topology transmembrane proteins. In contrast, β1B lacks a transmembrane domain and is a secreted protein. A growing body of work shows that VGSC β subunits are multifunctional. While they do not form the ion channel pore, β subunits alter gating, voltage-dependence, and kinetics of VGSCα subunits and thus regulate cellular excitability in vivo. In addition to their roles in channel modulation, β subunits are members of the immunoglobulin superfamily of cell adhesion molecules and regulate cell adhesion and migration. β subunits are also substrates for sequential proteolytic cleavage by secretases. An example of the multifunctional nature of β subunits is β1, encoded by SCN1B, that plays a critical role in neuronal migration and pathfinding during brain development, and whose function is dependent on Na+ current and γ-secretase activity. Functional deletion of SCN1B results in Dravet Syndrome, a severe and intractable pediatric epileptic encephalopathy. β subunits are emerging as key players in a wide variety of physiopathologies, including epilepsy, cardiac arrhythmia, multiple sclerosis, Huntington’s disease, neuropsychiatric disorders, neuropathic and inflammatory pain, and cancer. β subunits mediate multiple signaling pathways on different timescales, regulating electrical excitability, adhesion, migration, pathfinding, and transcription. Importantly, some β subunit functions may operate independently of α subunits. Thus, β subunits perform critical roles during development and disease. As such, they may prove useful in disease diagnosis and therapy. PMID:22007171

  6. Tropical species of Cladobotryum and Hypomyces producing red pigments

    PubMed Central

    Põldmaa, Kadri

    2011-01-01

    Twelve species of Hypomyces/Cladobotryum producing red pigments are reported growing in various tropical areas of the world. Ten of these are described as new, including teleomorphs for two previously known anamorphic species. In two species the teleomorph has been found in nature and in three others it was obtained in culture; only anamorphs are known for the rest. None of the studied tropical collections belongs to the common temperate species H. rosellus and H. odoratus to which the tropical teleomorphic collections had previously been assigned. Instead, taxa encountered in the tropics are genetically and morphologically distinct from the nine species of Hypomyces/Cladobotryum producing red pigments known from temperate regions. Besides observed host preferences, anamorphs of several species can spread fast on soft ephemeral agaricoid basidiomata but the slower developing teleomorphs are mostly found on polyporoid basidiomata or bark. While a majority of previous records from the tropics involve collections from Central America, this paper also reports the diversity of these fungi in the Paleotropics. Africa appears to hold a variety of taxa as five of the new species include material collected in scattered localities of this mostly unexplored continent. In examining distribution patterns, most of the taxa do not appear to be pantropical. Some species are known only from the Western Hemisphere, while others have a geographic range from southeastern Asia to Africa or Australia. The use of various morphological characters of anamorphs and teleomorphs as well as culture characteristics in species delimitation is evaluated. For detecting genetic segregation, partial sequences of the two largest subunits of the ribosomal polymerase perform the best in terms of providing informative sites and the number of well-supported groups recognised in the phylogenies. These are followed by the sequence data of the translation-elongation factor 1-alpha, while the ribosomal DNA

  7. Calcium activated K⁺ channels in the electroreceptor of the skate confirmed by cloning. Details of subunits and splicing.

    PubMed

    King, Benjamin L; Shi, Ling Fang; Kao, Peter; Clusin, William T

    2016-03-01

    Elasmobranchs detect small potentials using excitable cells of the ampulla of Lorenzini which have calcium-activated K(+) channels, first described in 1974. A distinctive feature of the outward current in voltage clamped ampullae is its apparent insensitivity to voltage. The sequence of a BK channel α isoform expressed in the ampulla of the skate was characterized. A signal peptide is present at the beginning of the gene. When compared to human isoform 1 (the canonical sequence), the largest difference was absence of a 59 amino acid region from the S8-S9 intra-cellular linker that contains the strex regulatory domain. The ampulla isoform was also compared with the isoform predicted in late skate embryos where strex was also absent. The BK voltage sensors were conserved in both skate isoforms. Differences between the skate and human BK channel included alternative splicing. Alternative splicing occurs at seven previously defined sites that are characteristic for BK channels in general and hair cells in particular. Skate BK sequences were highly similar to the Australian ghost shark and several other vertebrate species. Based on alignment of known BK sequences with the skate genome and transcriptome, there are at least two isoforms of Kcnma1α expressed in the skate. One of the β subunits (β4), which is known to decrease voltage sensitivity, was also identified in the skate genome and transcriptome and in the ampulla. These studies advance our knowledge of BK channels and suggest further studies in the ampulla and other excitable tissues. PMID:26687710

  8. Favourability: concept, distinctive characteristics and potential usefulness

    NASA Astrophysics Data System (ADS)

    Acevedo, Pelayo; Real, Raimundo

    2012-07-01

    The idea of analysing the general favourability for the occurrence of an event was presented in 2006 through a mathematical function. However, even when favourability has been used in species distribution modelling, the conceptual framework of this function is not yet well perceived among many researchers. The present paper is conceived for providing a wider and more in-depth presentation of the idea of favourability; concretely, we aimed to clarify both the concept and the main distinctive characteristics of the favourability function, especially in relation to probability and suitability, the most common outputs in species distribution modelling. As the capabilities of the favourability function go beyond species distribution modelling, we also illustrate its usefulness for different research disciplines for which this function remains unknown. In particular, we stressed that the favourability function has potential to be applied in all the cases where the probability of occurrence of an event is analysed, such as, for example, habitat selection or epidemiological studies.

  9. Molecular cloning and expression of a GABA receptor subunit from the crayfish Procambarus clarkii.

    PubMed

    Jiménez-Vázquez, Eric N; Díaz-Velásquez, Clara E; Uribe, R M; Arias, Juan M; García, Ubaldo

    2016-02-01

    Molecular cloning has introduced an unexpected, large diversity of neurotransmitter hetero- oligomeric receptors. Extensive research on the molecular structure of the γ-aminobutyric acid receptor (GABAR) has been of great significance for understanding how the nervous system works in both vertebrates and invertebrates. However, only two examples of functional homo-oligomeric GABA-activated Cl(-) channels have been reported. In the vertebrate retina, the GABAρ1 subunit of various species forms homo-oligomeric receptors; in invertebrates, a cDNA encoding a functional GABA-activated Cl(-) channel has been isolated from a Drosophila melanogaster head cDNA library. When expressed in Xenopus laevis oocytes, these subunits function efficiently as a homo-oligomeric complex. To investigate the structure-function of GABA channels from the crayfish Procambarus clarkii, we cloned a subunit and expressed it in human embryonic kidney cells. Electrophysiological recordings show that this subunit forms a homo-oligomeric ionotropic GABAR that gates a bicuculline-insensitive Cl(-) current. The order of potency of the agonists was GABA > trans-4-amino-crotonic acid = cis-4-aminocrotonic acid > muscimol. These data support the notion that X-organ sinus gland neurons express at least two GABA subunits responsible for the formation of hetero-oligomeric and homo-oligomeric receptors. In addition, by in situ hybridization studies we demonstrate that most X-organ neurons from crayfish eyestalk express the isolated pcGABAA β subunit. This study increases the knowledge of the genetics of the crayfish, furthers the understanding of this important neurotransmitter receptor family, and provides insight into the evolution of these genes among vertebrates and invertebrates.

  10. Peltaster cerophilus is a new species of the apple sooty blotch complex from Europe.

    PubMed

    Medjedović, Ajda; Frank, Jana; Schroers, Hans-Josef; Oertel, Bernd; Batzer, Jean Carlson

    2014-01-01

    Adopting the currently used concept for the genus Peltaster, the sooty blotch fungus Peltaster cerophilus is newly described from the cuticle of ripening or ripe apples. It forms a punctate phenotype consisting of superficially formed pycnothyria and a superficial mycelial mat consisting of a net of brown or brownish black hyphae. The pycnothyria are olivaceous brown to brown but have a spot in the center that is less strongly pigmented. Pycnothyria on the holotype of P. fructicola are homogeneously pigmented. On synthetic nutrient-poor agar, P. cerophilus is largely indistinguishable from P. fructicola. It forms delicate, spreading hyphae and intercalary conidiogenous cells with short, lateral, apically thick-walled conidiogenous necks forming blastic, unpigmented, one-celled conidia in basipetal succession. Conidia can swell and become one-septate. The species has microcyclical conidiation in proximate parts of colonies. DNA sequence analyses based on the ITS and the partial nuclear small and large subunit ribosomal RNA genes, the partial mitochondrial small subunit rRNA gene and the partial translation elongation factor 1-α gene support the distinction of the European P. cerophilus from P. fructicola, which is known from North America and Europe. The nuclear small ribosomal RNA subunit gene sequences of P. cerophilus contain two group I introns at locations known to accommodate introns in certain other, unrelated taxa. One of these, for which the code "SSU-1506 intron" was adopted, is 1459 base pairs long and located between the universal primer sites ITS5 and ITS1. Similar or positional-differing introns were encountered also in three currently undescribed Peltaster species. Representative strains of Peltaster fructicola did not accommodate introns in the nuclear small subunit ribosomal RNA gene.

  11. Botrytis californica, a new cryptic species in the B. cinerea species complex causing gray mold in blueberries and table grapes.

    PubMed

    Saito, S; Margosan, D; Michailides, T J; Xiao, C L

    2016-01-01

    The Botrytis cinerea species complex comprises two cryptic species, originally referred to Group I and Group II based on Bc-hch gene RFLP haplotyping. Group I was described as a new cryptic species B. pseudocinerea During a survey of Botrytis spp. causing gray mold in blueberries and table grapes in the Central Valley of California, six isolates, three from blueberries and three from table grapes, were placed in Group I but had a distinct morphological character with conidiophores significantly longer than those of B. cinerea and B. pseudocinerea We compared these with B. cinerea and B. pseudocinerea by examining morphological and physiological characters, sensitivity to fenhexamid and phylogenetic analysis inferred from sequences of three nuclear genes. Phylogenetic analysis with the three partial gene sequences encoding glyceraldehyde-3-phosate dehydrogenase (G3PDH), heat-shock protein 60 (HSP60) and DNA-dependent RNA polymerase subunit II (RPB2) supported the proposal of a new Botrytis species, B. californica, which is closely related genetically to B. cinerea, B. pseudocinerea and B. sinoviticola, all known as causal agents of gray mold of grapes. Botrytis californica caused decay on blueberry and table grape fruit inoculated with the fungus. This study suggests that B. californica is a cryptic species sympatric with B. cinerea on blueberries and table grapes in California. PMID:26740541

  12. Botrytis californica, a new cryptic species in the B. cinerea species complex causing gray mold in blueberries and table grapes.

    PubMed

    Saito, S; Margosan, D; Michailides, T J; Xiao, C L

    2016-01-01

    The Botrytis cinerea species complex comprises two cryptic species, originally referred to Group I and Group II based on Bc-hch gene RFLP haplotyping. Group I was described as a new cryptic species B. pseudocinerea During a survey of Botrytis spp. causing gray mold in blueberries and table grapes in the Central Valley of California, six isolates, three from blueberries and three from table grapes, were placed in Group I but had a distinct morphological character with conidiophores significantly longer than those of B. cinerea and B. pseudocinerea We compared these with B. cinerea and B. pseudocinerea by examining morphological and physiological characters, sensitivity to fenhexamid and phylogenetic analysis inferred from sequences of three nuclear genes. Phylogenetic analysis with the three partial gene sequences encoding glyceraldehyde-3-phosate dehydrogenase (G3PDH), heat-shock protein 60 (HSP60) and DNA-dependent RNA polymerase subunit II (RPB2) supported the proposal of a new Botrytis species, B. californica, which is closely related genetically to B. cinerea, B. pseudocinerea and B. sinoviticola, all known as causal agents of gray mold of grapes. Botrytis californica caused decay on blueberry and table grape fruit inoculated with the fungus. This study suggests that B. californica is a cryptic species sympatric with B. cinerea on blueberries and table grapes in California.

  13. PKA catalytic subunit mutations in adrenocortical Cushing's adenoma impair association with the regulatory subunit.

    PubMed

    Calebiro, Davide; Hannawacker, Annette; Lyga, Sandra; Bathon, Kerstin; Zabel, Ulrike; Ronchi, Cristina; Beuschlein, Felix; Reincke, Martin; Lorenz, Kristina; Allolio, Bruno; Kisker, Caroline; Fassnacht, Martin; Lohse, Martin J

    2014-01-01

    We recently identified a high prevalence of mutations affecting the catalytic (Cα) subunit of protein kinase A (PKA) in cortisol-secreting adrenocortical adenomas. The two identified mutations (Leu206Arg and Leu199_Cys200insTrp) are associated with increased PKA catalytic activity, but the underlying mechanisms are highly controversial. Here we utilize a combination of biochemical and optical assays, including fluorescence resonance energy transfer in living cells, to analyze the consequences of the two mutations with respect to the formation of the PKA holoenzyme and its regulation by cAMP. Our results indicate that neither mutant can form a stable PKA complex, due to the location of the mutations at the interface between the catalytic and the regulatory subunits. We conclude that the two mutations cause high basal catalytic activity and lack of regulation by cAMP through interference of complex formation between the regulatory and the catalytic subunits of PKA. PMID:25477193

  14. Counselor Identity: Conformity or Distinction?

    ERIC Educational Resources Information Center

    McLaughlin, Jerry E.; Boettcher, Kathryn

    2009-01-01

    The authors explore 3 debates in other disciplines similar to counseling's identity debate in order to learn about common themes and outcomes. Conformity, distinction, and cohesion emerged as common themes. They conclude that counselors should retain their distinctive, humanistic approach rather than conforming to the dominant, medical approach.

  15. Is Face Distinctiveness Gender Based?

    ERIC Educational Resources Information Center

    Baudouin, Jean-Yves; Gallay, Mathieu

    2006-01-01

    Two experiments were carried out to study the role of gender category in evaluations of face distinctiveness. In Experiment 1, participants had to evaluate the distinctiveness and the femininity-masculinity of real or artificial composite faces. The composite faces were created by blending either faces of the same gender (sexed composite faces,…

  16. Optimal Distinctiveness Signals Membership Trust.

    PubMed

    Leonardelli, Geoffrey J; Loyd, Denise Lewin

    2016-07-01

    According to optimal distinctiveness theory, sufficiently small minority groups are associated with greater membership trust, even among members otherwise unknown, because the groups are seen as optimally distinctive. This article elaborates on the prediction's motivational and cognitive processes and tests whether sufficiently small minorities (defined by relative size; for example, 20%) are associated with greater membership trust relative to mere minorities (45%), and whether such trust is a function of optimal distinctiveness. Two experiments, examining observers' perceptions of minority and majority groups and using minimal groups and (in Experiment 2) a trust game, revealed greater membership trust in minorities than majorities. In Experiment 2, participants also preferred joining minorities over more powerful majorities. Both effects occurred only when minorities were 20% rather than 45%. In both studies, perceptions of optimal distinctiveness mediated effects. Discussion focuses on the value of relative size and optimal distinctiveness, and when membership trust manifests. PMID:27140657

  17. Dynamic subunit stoichiometry confers a progressive continuum of pharmacological sensitivity by KCNQ potassium channels.

    PubMed

    Yu, Haibo; Lin, Zhihong; Mattmann, Margrith E; Zou, Beiyan; Terrenoire, Cecile; Zhang, Hongkang; Wu, Meng; McManus, Owen B; Kass, Robert S; Lindsley, Craig W; Hopkins, Corey R; Li, Min

    2013-05-21

    Voltage-gated KCNQ1 (Kv7.1) potassium channels are expressed abundantly in heart but they are also found in multiple other tissues. Differential coassembly with single transmembrane KCNE beta subunits in different cell types gives rise to a variety of biophysical properties, hence endowing distinct physiological roles for KCNQ1-KCNEx complexes. Mutations in either KCNQ1 or KCNE1 genes result in diseases in brain, heart, and the respiratory system. In addition to complexities arising from existence of five KCNE subunits, KCNE1 to KCNE5, recent studies in heterologous systems suggest unorthodox stoichiometric dynamics in subunit assembly is dependent on KCNE expression levels. The resultant KCNQ1-KCNE channel complexes may have a range of zero to two or even up to four KCNE subunits coassembling per KCNQ1 tetramer. These findings underscore the need to assess the selectivity of small-molecule KCNQ1 modulators on these different assemblies. Here we report a unique small-molecule gating modulator, ML277, that potentiates both homomultimeric KCNQ1 channels and unsaturated heteromultimeric (KCNQ1)4(KCNE1)n (n < 4) channels. Progressive increase of KCNE1 or KCNE3 expression reduces efficacy of ML277 and eventually abolishes ML277-mediated augmentation. In cardiomyocytes, the slowly activating delayed rectifier potassium current, or IKs, is believed to be a heteromultimeric combination of KCNQ1 and KCNE1, but it is not entirely clear whether IKs is mediated by KCNE-saturated KCNQ1 channels or by channels with intermediate stoichiometries. We found ML277 effectively augments IKs current of cultured human cardiomyocytes and shortens action potential duration. These data indicate that unsaturated heteromultimeric (KCNQ1)4(KCNE1)n channels are present as components of IKs and are pharmacologically distinct from KCNE-saturated KCNQ1-KCNE1 channels. PMID:23650380

  18. The RCN1-encoded A subunit of protein phosphatase 2A increases phosphatase activity in vivo

    NASA Technical Reports Server (NTRS)

    Deruere, J.; Jackson, K.; Garbers, C.; Soll, D.; Delong, A.; Evans, M. L. (Principal Investigator)

    1999-01-01

    Protein phosphatase 2A (PP2A), a heterotrimeric serine/threonine-specific protein phosphatase, comprises a catalytic C subunit and two distinct regulatory subunits, A and B. The RCN1 gene encodes one of three A regulatory subunits in Arabidopsis thaliana. A T-DNA insertion mutation at this locus impairs root curling, seedling organ elongation and apical hypocotyl hook formation. We have used in vivo and in vitro assays to gauge the impact of the rcn1 mutation on PP2A activity in seedlings. PP2A activity is decreased in extracts from rcn1 mutant seedlings, and this decrease is not due to a reduction in catalytic subunit expression. Roots of mutant seedlings exhibit increased sensitivity to the phosphatase inhibitors okadaic acid and cantharidin in organ elongation assays. Shoots of dark-grown, but not light-grown seedlings also show increased inhibitor sensitivity. Furthermore, cantharidin treatment of wild-type seedlings mimics the rcn1 defect in root curling, root waving and hypocotyl hook formation assays. In roots of wild-type seedlings, RCN1 mRNA is expressed at high levels in root tips, and accumulates to lower levels in the pericycle and lateral root primordia. In shoots, RCN1 is expressed in the apical hook and the basal, rapidly elongating cells in etiolated hypocotyls, and in the shoot meristem and leaf primordia of light-grown seedlings. Our results show that the wild-type RCN1-encoded A subunit functions as a positive regulator of the PP2A holoenzyme, increasing activity towards substrates involved in organ elongation and differential cell elongation responses such as root curling.

  19. Microbial community composition and in silico predicted metabolic potential reflect biogeochemical gradients between distinct peatland types.

    PubMed

    Urbanová, Zuzana; Bárta, Jiří

    2014-12-01

    It is not well understood how the ecological status and microbial community composition of spruce swamp forests (SSF) relate to those found in bogs and fens. To clarify this, we investigated biogeochemical parameters and microbial community composition in a bog, a fen and two SSF using high throughput barcoded sequencing of the small ribosomal subunit (SSU) variable region V4. The results demonstrated that the microbial community of SSF is positioned between those of bogs and fens, and this was confirmed by in silico predicted metabolic potentials. This corresponds well with the position of SSF on the trophic gradient and reflects distinct responses of microbial communities to environmental variables. Species richness and microbial diversity increased significantly from bog to fen, with SSF in between, reflecting the variation in pH, nutrient availability and peat decomposability. The archaeal community, dominated by hydrogenotrophic methanogens, was more similar in SSF and the bog compared with the fen. The composition of the bacterial community of SSF was intermediate between those of bog and fen. However, the production of CO2 (an indicator of peat decomposability) did not differ between SSF and bog, suggesting the limiting effect of low pH and poor litter quality on the functioning of the bacterial community in SSF. These results help to clarify the transitional position of SSF between bogs and fens and showed the strong effect of environmental conditions on microbial community composition and functioning.

  20. Kir3 channel ontogeny – the role of Gβγ subunits in channel assembly and trafficking

    PubMed Central

    Zylbergold, Peter; Sleno, Rory; Khan, Shahriar M.; Jacobi, Ashley M.; Belhke, Mark A.; Hébert, Terence E.

    2014-01-01

    The role of Gβγ subunits in Kir3 channel gating is well characterized. Here, we have studied the role of Gβγ dimers during their initial contact with Kir3 channels, prior to their insertion into the plasma membrane. We show that distinct Gβγ subunits play an important role in orchestrating and fine-tuning parts of the Kir3 channel life cycle. Gβ1γ2, apart from its role in channel opening that it shares with other Gβγ subunit combinations, may play a unique role in protecting maturing channels from degradation as they transit to the cell surface. Taken together, our data suggest that Gβ1γ2 prolongs the lifetime of the Kir3.1/Kir3.2 heterotetramer, although further studies would be required to shed more light on these early Gβγ effects on Kir3 maturation and trafficking. PMID:24782712

  1. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates.

    PubMed

    Folmer, O; Black, M; Hoeh, W; Lutz, R; Vrijenhoek, R

    1994-10-01

    We describe "universal" DNA primers for polymerase chain reaction (PCR) amplification of a 710-bp fragment of the mitochondrial cytochrome c oxidase subunit I gene (COI) from 11 invertebrate phyla: Echinodermata, Mollusca, Annelida, Pogonophora, Arthropoda, Nemertinea, Echiura, Sipuncula, Platyhelminthes, Tardigrada, and Coelenterata, as well as the putative phylum Vestimentifera. Preliminary comparisons revealed that these COI primers generate informative sequences for phylogenetic analyses at the species and higher taxonomic levels.

  2. [Nose surgical anatomy in six aesthetic subunits].

    PubMed

    Chaput, B; Lauwers, F; Lopez, R; Saboye, J; André, A; Grolleau, J-L; Chavoin, J-P

    2013-04-01

    The nose is a complex entity, combining aesthetic and functional roles. Descriptive anatomy is a fundamental science that it can be difficult to relate directly to our daily surgical activity. Reasoning in terms of aesthetic subunits to decide on his actions appeared to us so obvious. The aim of this paper is to resume the anatomical bases relevant to our daily practice in order to fully apprehend the restorative or cosmetic procedures. We discuss the limits of the systematization of these principles in nasal oncology.

  3. Molecular and morphological evidence indicates that Pseudorhabdosynochus lantauensis (Monogenea: Diplectanidae) represents two species.

    PubMed

    Wu, X Y; Chilton, N B; Zhu, X Q; Xie, M Q; Li, A X

    2005-06-01

    Sequences of the first internal transcribed spacer (ITS-1) and the D1-D3 domains of the large subunit (LSU) of the ribosomal DNA (rDNA) were determined for multiple specimens of 4 operational taxonomic units (OTUs) of the monogenean, Pseudorhabdosynochus lantauensis. OTUs were defined based on their collecting localities, host and/or morphological characteristics. All P. lantauensis specimens of one group (OTUs 1 and 3) differed in their sequences of the ITS-1 and partial LSU rDNA when compared with specimens of a second group (OTUs 2 and 4) by 12% and 2%, respectively. Results of the phylogenetic analyses of the LSU rDNA sequence data showed total (100%) bootstrap support for the separation of P. lantauensis into 2 distinct clades. At least 11 of the 18 nucleotide differences in the LSU sequence between the two P. lantauensis clades were derived (i.e. autapomorphic) characters when the morphologically distinct species, P. epinepheli and P. coioidesis, were used as outgroups. Furthermore, there were several autapomorphic character states for each P. lantauensis clade. This provides sufficient evidence to reject the null hypothesis that P. lantauensis represents a single species. Morphological and morphometric differences between these two clades provided additional strong support for the separation of P. lantauensis into two species. These two parasite species were found to co-exist on one of the two species of serranid fish (i.e. Epinephelus coioides) examined in the South China Sea (Guangdong Province, China).

  4. DNA Barcodes of Rosy Tetras and Allied Species (Characiformes: Characidae: Hyphessobrycon) from the Brazilian Amazon Basin

    PubMed Central

    Castro Paz, Francis Paola; Batista, Jacqueline da Silva; Porto, Jorge Ivan Rebelo

    2014-01-01

    DNA barcoding can be an effective tool for fast and accurate species-level identification based on sequencing of the mitochondrial cytochrome c oxidase subunit (COI) gene. The diversity of this fragment can be used to estimate the richness of the respective species. In this study, we explored the use of DNA barcoding in a group of ornamental freshwater fish of the genus Hyphessobrycon. We sequenced the COI from 10 species of Hyphessobrycon belonging to the “Rosy Tetra Clade” collected from the Amazon and Negro River basins and combined our results with published data. The average conspecific and congeneric Kimura 2-parameter distances were 2.3% and 19.3%, respectively. Six of the 10 species were easily distinguishable by DNA barcoding (H. bentosi, H. copelandi, H. eques, H. epicharis, H. pulchrippinis, and H. sweglesi), whereas the remaining species (H. erythrostigma, H. pyrrhonotus, H. rosaceus and H. socolofi) lacked reciprocal monophyly. Although the COI gene was not fully diagnostic, the discovery of distinct evolutionary units in certain Hyphessobrycon species under the same specific epithet as well as haplotype sharing between different species suggest that DNA barcoding is useful for species identification in this speciose genus. PMID:24878569

  5. DNA barcodes of Rosy Tetras and allied species (Characiformes: Characidae: Hyphessobrycon) from the Brazilian Amazon basin.

    PubMed

    Castro Paz, Francis Paola; Batista, Jacqueline da Silva; Porto, Jorge Ivan Rebelo

    2014-01-01

    DNA barcoding can be an effective tool for fast and accurate species-level identification based on sequencing of the mitochondrial cytochrome c oxidase subunit (COI) gene. The diversity of this fragment can be used to estimate the richness of the respective species. In this study, we explored the use of DNA barcoding in a group of ornamental freshwater fish of the genus Hyphessobrycon. We sequenced the COI from 10 species of Hyphessobrycon belonging to the "Rosy Tetra Clade" collected from the Amazon and Negro River basins and combined our results with published data. The average conspecific and congeneric Kimura 2-parameter distances were 2.3% and 19.3%, respectively. Six of the 10 species were easily distinguishable by DNA barcoding (H. bentosi, H. copelandi, H. eques, H. epicharis, H. pulchrippinis, and H. sweglesi), whereas the remaining species (H. erythrostigma, H. pyrrhonotus, H. rosaceus and H. socolofi) lacked reciprocal monophyly. Although the COI gene was not fully diagnostic, the discovery of distinct evolutionary units in certain Hyphessobrycon species under the same specific epithet as well as haplotype sharing between different species suggest that DNA barcoding is useful for species identification in this speciose genus.

  6. A multi-gene phylogeny for species of Mycosphaerella occurring on Eucalyptus leaves.

    PubMed

    Hunter, Gavin C; Wingfield, Brenda D; Crous, Pedro W; Wingfield, Michael J

    2006-01-01

    Species of the ascomycete genus Mycosphaerella are regarded as some of the most destructive leaf pathogens of a large number of economically important crop plants. Amongst these, approximately 60 Mycosphaerella spp. have been identified from various Eucalyptus spp. where they cause leaf diseases collectively known as Mycosphaerella Leaf Disease (MLD). Species concepts for this group of fungi remain confused, and hence their species identification is notoriously difficult. Thus, the introduction of DNA sequence comparisons has become the definitive characteristic used to distinguish species of Mycosphaerella. Sequences of the Internal Transcribed Spacer (ITS) region of the ribosomal RNA operon have most commonly been used to consider species boundaries in Mycosphaerella. However, sequences for this gene region do not always provide sufficient resolution for cryptic taxa. The aim of this study was, therefore, to use DNA sequences for three loci, ITS, Elongation factor 1-alpha (EF-1alpha) and Actin (ACT) to reconsider species boundaries for Mycosphaerella spp. from Eucalyptus. A further aim was to study the anamorph concepts and resolve the deeper nodes of Mycosphaerella, for which part of the Large Subunit (LSU) of the nuclear rRNA operon was sequenced. The ITS and EF-1alpha gene regions were found to be useful, but the ACT gene region did not provide species-level resolution in Mycosphaerella. A phylogeny of the combined DNA datasets showed that species of Mycosphaerella from Eucalyptus cluster in two distinct groups, which might ultimately represent discrete genera. PMID:18490976

  7. Reservoir floodplains support distinct fish assemblages

    USGS Publications Warehouse

    Miranda, Leandro E.; Wigen, S. L.; Dagel, Jonah D.

    2014-01-01

    Reservoirs constructed on floodplain rivers are unique because the upper reaches of the impoundment may include extensive floodplain environments. Moreover, reservoirs that experience large periodic water level fluctuations as part of their operational objectives seasonally inundate and dewater floodplains in their upper reaches, partly mimicking natural inundations of river floodplains. In four flood control reservoirs in Mississippi, USA, we explored the dynamics of connectivity between reservoirs and adjacent floodplains and the characteristics of fish assemblages that develop in reservoir floodplains relative to those that develop in reservoir bays. Although fish species richness in floodplains and bays were similar, species composition differed. Floodplains emphasized fish species largely associated with backwater shallow environments, often resistant to harsh environmental conditions. Conversely, dominant species in bays represented mainly generalists that benefit from the continuous connectivity between the bay and the main reservoir. Floodplains in the study reservoirs provided desirable vegetated habitats at lower water level elevations, earlier in the year, and more frequently than in bays. Inundating dense vegetation in bays requires raising reservoir water levels above the levels required to reach floodplains. Therefore, aside from promoting distinct fish assemblages within reservoirs and helping promote diversity in regulated rivers, reservoir floodplains are valued because they can provide suitable vegetated habitats for fish species at elevations below the normal pool, precluding the need to annually flood upland vegetation that would inevitably be impaired by regular flooding. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  8. Mass spectrometry reveals modularity and a complete subunit interaction map of the eukaryotic translation factor eIF3.

    PubMed

    Zhou, Min; Sandercock, Alan M; Fraser, Christopher S; Ridlova, Gabriela; Stephens, Elaine; Schenauer, Matthew R; Yokoi-Fong, Theresa; Barsky, Daniel; Leary, Julie A; Hershey, John W; Doudna, Jennifer A; Robinson, Carol V

    2008-11-25

    The eukaryotic initiation factor 3 (eIF3) plays an important role in translation initiation, acting as a docking site for several eIFs that assemble on the 40S ribosomal subunit. Here, we use mass spectrometry to probe the subunit interactions within the human eIF3 complex. Our results show that the 13-subunit complex can be maintained intact in the gas phase, enabling us to establish unambiguously its stoichiometry and its overall subunit architecture via tandem mass spectrometry and solution disruption experiments. Dissociation takes place as a function of ionic strength to form three stable modules eIF3(c:d:e:l:k), eIF3(f:h:m), and eIF3(a:b:i:g). These modules are linked by interactions between subunits eIF3b:c and eIF3c:h. We confirmed our interaction map with the homologous yeast eIF3 complex that contains the five core subunits found in the human eIF3 and supplemented our data with results from immunoprecipitation. These results, together with the 27 subcomplexes identified with increasing ionic strength, enable us to define a comprehensive interaction map for this 800-kDa species. Our interaction map allows comparison of free eIF3 with that bound to the hepatitis C virus internal ribosome entry site (HCV-IRES) RNA. We also compare our eIF3 interaction map with related complexes, containing evolutionarily conserved protein domains, and reveal the location of subunits containing RNA recognition motifs proximal to the decoding center of the 40S subunit of the ribosome.

  9. Subunit Arrangement and Function in NMDA Receptors

    SciTech Connect

    Furukawa,H.; Singh, S.; Mancusso, R.; Gouaux, E.

    2005-01-01

    Excitatory neurotransmission mediated by NMDA (N-methyl-D-aspartate) receptors is fundamental to the physiology of the mammalian central nervous system. These receptors are heteromeric ion channels that for activation require binding of glycine and glutamate to the NR1 and NR2 subunits, respectively. NMDA receptor function is characterized by slow channel opening and deactivation, and the resulting influx of cations initiates signal transduction cascades that are crucial to higher functions including learning and memory. Here we report crystal structures of the ligand-binding core of NR2A with glutamate and that of the NR1-NR2A heterodimer with glutamate and glycine. The NR2A-glutamate complex defines the determinants of glutamate and NMDA recognition, and the NR1-NR2A heterodimer suggests a mechanism for ligand-induced ion channel opening. Analysis of the heterodimer interface, together with biochemical and electrophysiological experiments, confirms that the NR1-NR2A heterodimer is the functional unit in tetrameric NMDA receptors and that tyrosine 535 of NR1, located in the subunit interface, modulates the rate of ion channel deactivation.

  10. Binary Toxin Subunits of Lysinibacillus sphaericus Are Monomeric and Form Heterodimers after In Vitro Activation.

    PubMed

    Surya, Wahyu; Chooduang, Sivadatch; Choong, Yeu Khai; Torres, Jaume; Boonserm, Panadda

    2016-01-01

    The binary toxin from Lysinibacillus sphaericus has been successfully used for controlling mosquito-transmitted diseases. An activation step shortens both subunits BinA and BinB before their interaction with membranes and internalization in midgut cells, but the precise role of this activation step is unknown. Herein, we show conclusively using three orthogonal biophysical techniques that protoxin subunits form only monomers in aqueous solution. However, in vitro activated toxins readily form heterodimers. This oligomeric state did not change after incubation of these heterodimers with detergent. These results are consistent with the evidence that maximal toxicity in mosquito larvae is achieved when the two subunits, BinA and BinB, are in a 1:1 molar ratio, and directly link proteolytic activation to heterodimerization. Formation of a heterodimer must thus be necessary for subsequent steps, e.g., interaction with membranes, or with a suitable receptor in susceptible mosquito species. Lastly, despite existing similarities between BinB C-terminal domain with domains 3 and 4 of pore-forming aerolysin, no aerolysin-like SDS-resistant heptameric oligomers were observed when the activated Bin subunits were incubated in the presence of detergents or lipidic membranes.

  11. The δ Subunit of RNA Polymerase Guides Promoter Selectivity and Virulence in Staphylococcus aureus

    PubMed Central

    Weiss, Andy; Ibarra, J. Antonio; Paoletti, Jessica; Carroll, Ronan K.

    2014-01-01

    In Gram-positive bacteria, and particularly the Firmicutes, the DNA-dependent RNA polymerase (RNAP) complex contains an additional subunit, termed the δ factor, or RpoE. This enigmatic protein has been studied for more than 30 years for various organisms, but its function is still not well understood. In this study, we investigated its role in the major human pathogen Staphylococcus aureus. We showed conservation of important structural regions of RpoE in S. aureus and other species and demonstrated binding to core RNAP that is mediated by the β and/or β′ subunits. To identify the impact of the δ subunit on transcription, we performed transcriptome sequencing (RNA-seq) analysis and observed 191 differentially expressed genes in the rpoE mutant. Ontological analysis revealed, quite strikingly, that many of the downregulated genes were known virulence factors, while several mobile genetic elements (SaPI5 and prophage ϕSA3usa) were strongly upregulated. Phenotypically, the rpoE mutant had decreased accumulation and/or activity of a number of key virulence factors, including alpha toxin, secreted proteases, and Panton-Valentine leukocidin (PVL). We further observed significantly decreased survival of the mutant in whole human blood, increased phagocytosis by human leukocytes, and impaired virulence in a murine model of infection. Collectively, our results demonstrate that the δ subunit of RNAP is a critical component of the S. aureus transcription machinery and plays an important role during infection. PMID:24491578

  12. Binary Toxin Subunits of Lysinibacillus sphaericus Are Monomeric and Form Heterodimers after In Vitro Activation

    PubMed Central

    Choong, Yeu Khai; Torres, Jaume; Boonserm, Panadda

    2016-01-01

    The binary toxin from Lysinibacillus sphaericus has been successfully used for controlling mosquito-transmitted diseases. An activation step shortens both subunits BinA and BinB before their interaction with membranes and internalization in midgut cells, but the precise role of this activation step is unknown. Herein, we show conclusively using three orthogonal biophysical techniques that protoxin subunits form only monomers in aqueous solution. However, in vitro activated toxins readily form heterodimers. This oligomeric state did not change after incubation of these heterodimers with detergent. These results are consistent with the evidence that maximal toxicity in mosquito larvae is achieved when the two subunits, BinA and BinB, are in a 1:1 molar ratio, and directly link proteolytic activation to heterodimerization. Formation of a heterodimer must thus be necessary for subsequent steps, e.g., interaction with membranes, or with a suitable receptor in susceptible mosquito species. Lastly, despite existing similarities between BinB C-terminal domain with domains 3 and 4 of pore-forming aerolysin, no aerolysin-like SDS-resistant heptameric oligomers were observed when the activated Bin subunits were incubated in the presence of detergents or lipidic membranes. PMID:27341696

  13. A human RNA polymerase II subunit is encoded by a recently generated multigene family

    PubMed Central

    Grandemange, Sylvie; Schaller, Sophie; Yamano, Shigeru; Du Manoir, Stanislas; Shpakovski, George V; Mattei, Marie-Geneviève; Kedinger, Claude; Vigneron, Marc

    2001-01-01

    Background The sequences encoding the yeast RNA polymerase II (RPB) subunits are single copy genes. Results While those characterized so far for the human (h) RPB are also unique, we show that hRPB subunit 11 (hRPB11) is encoded by a multigene family, mapping on chromosome 7 at loci p12, q11.23 and q22. We focused on two members of this family, hRPB11a and hRPB11b: the first encodes subunit hRPB11a, which represents the major RPB11 component of the mammalian RPB complex ; the second generates polypeptides hRPB11bα and hRPB11bβ through differential splicing of its transcript and shares homologies with components of the hPMS2L multigene family related to genes involved in mismatch-repair functions (MMR). Both hRPB11a and b genes are transcribed in all human tissues tested. Using an inter-species complementation assay, we show that only hRPB11bα is functional in yeast. In marked contrast, we found that the unique murine homolog of RPB11 gene maps on chromosome 5 (band G), and encodes a single polypeptide which is identical to subunit hRPB11a. Conclusions The type hRPB11b gene appears to result from recent genomic recombination events in the evolution of primates, involving sequence elements related to the MMR apparatus. PMID:11747469

  14. New Species of Boletellus Section Boletellus (Boletaceae, Boletales) from Japan, B. aurocontextus sp. nov. and B. areolatus sp. nov.

    PubMed Central

    Sato, Hirotoshi; Hattori, Tsutomu

    2015-01-01

    We describe and illustrate two new species of Boletellus section Boletellus, B. aurocontextus sp. nov. and B. areolatus sp. nov., which are generally assumed to be B. emodensis. In this study, we reconstructed separate molecular phylogenetic trees of section Boletellus using the nucleotide sequences of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA, the largest subunit (RPB1) and the second-largest subunit (RPB2) of nuclear RNA polymerase II gene and mitochondrial cytochrome oxidase subunit 3 (cox3) gene. We also examined the morphologies of B. emodensis sensu lato (s.l.) and other related species for comparison. The molecular phylogenetic tree inferred from the sequences of nuclear DNA (ITS, and combined dataset of RPB1 and RPB2) indicated that three genetically and phylogenetically well-separated lineages were present within B. emodensis s.l. These three lineages were also distinguished on the basis of the molecular phylogenetic tree constructed using the sequences of mitochondrial DNA (cox3), suggesting distinct cytonuclear disequilibria (i.e., evidence of reproductive isolation) among these lineages. Therefore, these three lineages can be treated as independent species: B. aurocontextus, B. areolatus, and B. emodensis. Boletellus aurocontextus and B. areolatus are also distinct from B. emodensis by the macro- and microscopic morphologies. Boletellus aurocontextus is characterized by a pileus with bright yellow to lemon yellow context, which can be observed through a gap in the scales, and basidiospores with relatively large length (mean spore length, 21.4 μm; quotient of spore length and width, 2.51). In contrast, B. areolatus is characterized by a pileus with floccose to appressed thin scaly patches, a stipe with pallid or pale cream color at the upper half, and basidiospores with relatively small length (mean spore length, 16.5 μm; quotient of spore length and width, 1.80). PMID:26083243

  15. The sodium channel auxiliary subunits beta1 and beta2 are differentially expressed in the spinal cord of neuropathic rats.

    PubMed

    Blackburn-Munro, G; Fleetwood-Walker, S M

    1999-04-01

    Neuropathic pain is thought to arise from ectopic discharges at the site of injury within the peripheral nervous system, and is manifest as a general increase in the level of neuronal excitability within primary afferent fibres and their synaptic contacts within the spinal cord. Voltage-activated Na+ channel blockers such as lamotrigine have been shown to be clinically effective in the treatment of neuropathic pain. Na+ channels are structurally diverse comprising a principal a subunit (of which there are variable isoforms) and two auxiliary subunits termed beta1 and beta2. Both beta subunits affect the rates of channel activation and inactivation, and can modify alpha subunit density within the plasma membrane. In addition, these subunits may interact with extracellular matrix molecules to affect growth and myelination of axons. Using in situ hybridization histochemistry we have shown that the expression of the beta1 and beta2 subunits within the dorsal horn of the spinal cord of neuropathic rats is differentially regulated by a chronic constrictive injury to the sciatic nerve. At days 12-15 post-neuropathy, beta1 messenger RNA levels had increased, whereas beta2 messenger RNA levels had decreased significantly within laminae I, II on the ipsilateral side of the cord relative to the contralateral side. Within laminae III-IV beta2 messenger RNA levels showed a small but significant decrease on the ipsilateral side relative to the contralateral side, whilst expression of beta1 messenger RNA remained unchanged. Thus, differential regulation of the individual beta subunit types may (through their distinct influences on Na+ channel function) contribute to altered excitability of central neurons after neuropathic injury.

  16. DNA barcoding of oomycetes with cytochrome c oxidase subunit I and internal transcribed spacer

    PubMed Central

    Robideau, Gregg P; de Cock, Arthur W A M; Coffey, Michael D; Voglmayr, Hermann; Brouwer, Henk; Bala, Kanak; Chitty, David W; Désaulniers, Nicole; Eggertson, Quinn A; Gachon, Claire M M; Hu, Chia-Hui; Küpper, Frithjof C; Rintoul, Tara L; Sarhan, Ehab; Verstappen, Els C P; Zhang, Yonghong; Bonants, Peter J M; Ristaino, Jean B; Lévesque, C André

    2011-01-01

    Oomycete species occupy many different environments and many ecological niches. The genera Phytophthora and Pythium for example, contain many plant pathogens which cause enormous damage to a wide range of plant species. Proper identification to the species level is a critical first step in any investigation of oomycetes, whether it is research driven or compelled by the need for rapid and accurate diagnostics during a pathogen outbreak. The use of DNA for oomycete species identification is well established, but DNA barcoding with cytochrome c oxidase subunit I (COI) is a relatively new approach that has yet to be assessed over a significant sample of oomycete genera. In this study we have sequenced COI, from 1205 isolates representing 23 genera. A comparison to internal transcribed spacer (ITS) sequences from the same isolates showed that COI identification is a practical option; complementary because it uses the mitochondrial genome instead of nuclear DNA. In some cases COI was more discriminative than ITS at the species level. This is in contrast to the large ribosomal subunit, which showed poor species resolution when sequenced from a subset of the isolates used in this study. The results described in this paper indicate that COI sequencing and the dataset generated are a valuable addition to the currently available oomycete taxonomy resources, and that both COI, the default DNA barcode supported by GenBank, and ITS, the de facto barcode accepted by the oomycete and mycology community, are acceptable and complementary DNA barcodes to be used for identification of oomycetes. PMID:21689384

  17. Spatial arrangement and functional role of α subunits of proteasome activator PA28 in hetero-oligomeric form

    SciTech Connect

    Sugiyama, Masaaki; Sahashi, Hiroki; Kurimoto, Eiji; Takata, Shin-ichi; Yagi, Hirokazu; Kanai, Keita; Sakata, Eri; Minami, Yasufumi; Tanaka, Keiji; Kato, Koichi

    2013-03-01

    Highlights: ► Homologous α and β subunits are alternatively arranged in the PA28 heptameric ring. ► The flexible loops of the three α subunits surround the site of substrate entry. ► The loops serve as gatekeepers that selectively hinder passage of longer peptides. - Abstract: A major form of proteasome activator PA28 is a heteroheptamer composed of interferon-γ-inducible α and β subunits, which share approximately 50% amino acid identity and possess distinct insert loops. This activator forms a complex with the 20S proteasome and thereby stimulates proteasomal degradation of peptides in an ATP-independent manner, giving rise to smaller antigenic peptides presented by major histocompatibility complex class I molecules. In this study, we performed biophysical and biochemical characterization of the structure and function of the PA28 hetero-oligomer. Deuteration-assisted small-angle neutron scattering demonstrated three α and four β subunits are alternately arranged in the heptameric ring. In this arrangement, PA28 loops surround the central pore of the heptameric ring (site for peptide entry). Activating the 20S proteasome with a PA28 mutant that lacked the α subunit loops cleaved model substrates longer than a nonapeptide with better efficiency when compared to wild-type PA28. Based on these data, we hypothesize that the flexible PA28 loops act as gatekeepers, which function to select the length of peptide substrates to be transported between the proteolytic chamber and the extra-proteasomal medium.

  18. GluN2A and GluN2B subunit-containing NMDA receptors in hippocampal plasticity

    PubMed Central

    Shipton, Olivia A.; Paulsen, Ole

    2014-01-01

    N-Methyl-d-aspartate receptor (NMDAR)-dependent synaptic plasticity is a strong candidate to mediate learning and memory processes that require the hippocampus. This plasticity is bidirectional, and how the same receptor can mediate opposite changes in synaptic weights remains a conundrum. It has been suggested that the NMDAR subunit composition could be involved. Specifically, one subunit composition of NMDARs would be responsible for the induction of long-term potentiation (LTP), whereas NMDARs with a different subunit composition would be engaged in the induction of long-term depression (LTD). Unfortunately, the results from studies that have investigated this hypothesis are contradictory, particularly in relation to LTD. Nevertheless, current evidence does suggest that the GluN2B subunit might be particularly important for plasticity and may make a synapse bidirectionally malleable. In particular, we conclude that the presence of GluN2B subunit-containing NMDARs at the postsynaptic density might be a necessary, though not a sufficient, condition for the strengthening of individual synapses. This is owing to the interaction of GluN2B with calcium/calmodulin-dependent protein kinase II (CaMKII) and is distinct from its contribution as an ion channel. PMID:24298164

  19. Identification and functional expression of a family of nicotinic acetylcholine receptor subunits in the central nervous system of the mollusc Lymnaea stagnalis.

    PubMed

    van Nierop, Pim; Bertrand, Sonia; Munno, David W; Gouwenberg, Yvonne; van Minnen, Jan; Spafford, J David; Syed, Naweed I; Bertrand, Daniel; Smit, August B

    2006-01-20

    We described a family of nicotinic acetylcholine receptor (nAChR) subunits underlying cholinergic transmission in the central nervous system (CNS) of the mollusc Lymnaea stagnalis. By using degenerate PCR cloning, we identified 12 subunits that display a high sequence similarity to nAChR subunits, of which 10 are of the alpha-type, 1 is of the beta-type, and 1 was not classified because of insufficient sequence information. Heterologous expression of identified subunits confirms their capacity to form functional receptors responding to acetylcholine. The alpha-type subunits can be divided into groups that appear to underlie cation-conducting (excitatory) and anion-conducting (inhibitory) channels involved in synaptic cholinergic transmission. The expression of the Lymnaea nAChR subunits, assessed by real time quantitative PCR and in situ hybridization, indicates that it is localized to neurons and widespread in the CNS, with the number and localization of expressing neurons differing considerably between subunit types. At least 10% of the CNS neurons showed detectable nAChR subunit expression. In addition, cholinergic neurons, as indicated by the expression of the vesicular ACh transporter, comprise approximately 10% of the neurons in all ganglia. Together, our data suggested a prominent role for fast cholinergic transmission in the Lymnaea CNS by using a number of neuronal nAChR subtypes comparable with vertebrate species but with a functional complexity that may be much higher.

  20. Strong cooperativity between subunits in voltage-gated proton channels

    PubMed Central

    Gonzalez, Carlos; Koch, Hans P.; Drum, Ben M.; Larsson, H. Peter

    2010-01-01

    Voltage-activated proton (HV) channels are essential components in the innate immune response. HV channels are dimeric proteins with one proton permeation pathway per subunit. It is not known how HV channels are activated by voltage and whether there is any cooperativity between subunits during voltage activation. Using cysteine accessibility measurements and voltage clamp fluorometry, we show data that are consistent with that the fourth transmembrane segment S4 functions as the voltage sensor in HV channels from Ciona intestinalis. Surprisingly, in a dimeric HV channel, S4 in both subunits have to move to activate the two proton permeation pathways. In contrast, if HV subunits are prevented from dimerizing, then the movement of a single S4 is sufficient to activate the proton permeation pathway in a subunit. These results suggest a strong cooperativity between subunits in dimeric HV channels. PMID:20023639

  1. Subunit stoichiometry of the chloroplast photosystem I complex

    SciTech Connect

    Bruce, B.D.; Malkin, R.

    1988-05-25

    A native photosystem I (PS I) complex and a PS I core complex depleted of antenna subunits has been isolated from the uniformly /sup 14/C-labeled aquatic higher plant, Lemna. These complexes have been analyzed for their subunit stoichiometry by quantitative sodium dodecyl sulfate-polyacrylamide gel electrophoresis methods. The results for both preparations indicate that one copy of each high molecular mass subunit is present per PS I complex and that a single copy of most low molecular mass subunits is also present. These results suggest that iron-sulfur center X, an early PS I electron acceptor proposed to bind to the high molecular mass subunits, contains a single (4Fe-4S) cluster which is bound to a dimeric structure of high molecular mass subunits, each providing 2 cysteine residues to coordinate this cluster.

  2. Cloning and characterization of homeologous cellulose synthase catalytic subunit 2 genes from allotetraploid cotton (Gossypium hirsutum L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cellulose synthase catalytic subunits (CesAs) are the catalytic sites within a multisubunit complex for cellulose biosynthesis in plants. CesAs have been extensively studied in diploid plants, but are not well characterized in polyploid plants. Gossypium hirsutum is an allotetraploid cotton specie...

  3. An atypical heterotrimeric G-protein γ-subunit is involved in guard cell K⁺-channel regulation and morphological development in Arabidopsis thaliana.

    PubMed

    Chakravorty, David; Trusov, Yuri; Zhang, Wei; Acharya, Biswa R; Sheahan, Michael B; McCurdy, David W; Assmann, Sarah M; Botella, José Ramón

    2011-09-01

    Currently, there are strong inconsistencies in our knowledge of plant heterotrimeric G-proteins that suggest the existence of additional members of the family. We have identified a new Arabidopsis G-protein γ-subunit (AGG3) that modulates morphological development and ABA-regulation of stomatal aperture. AGG3 strongly interacts with the Arabidopsis G-protein β-subunit in vivo and in vitro. Most importantly, AGG3-deficient mutants account for all but one of the 'orphan' phenotypes previously unexplained by the two known γ-subunits in Arabidopsis. AGG3 has unique characteristics never before observed in plant or animal systems, such as its size (more than twice that of canonical γ-subunits) and the presence of a C-terminal Cys-rich domain. AGG3 thus represent a novel class of G-protein γ-subunits, widely spread throughout the plant kingdom but not present in animals. Homologues of AGG3 in rice have been identified as important quantitative trait loci for grain size and yield, but due to the atypical nature of the proteins their identity as G-protein subunits was thus far unknown. Our work demonstrates a similar trend in seeds of Arabidopsis agg3 mutants, and implicates G-proteins in such a crucial agronomic trait. The discovery of this highly atypical subunit reinforces the emerging notion that plant and animal G-proteins have distinct as well as shared evolutionary pathways. PMID:21575088

  4. Subunit structure of the phycobiliproteins of blue-green algae.

    PubMed

    Glazer, A N; Cohen-Bazire, G

    1971-07-01

    The phycobiliproteins of the blue-green algae Synechococcus sp. and Aphanocapsu sp. were characterized with respect to homogeneity, isoelectric point, and subunit composition. Each of the biliproteins consisted of two different noncovalently associated subunits, with molecular weights of about 20,000 and 16,000 for phycocyanin, 17,500 and 15,500 for allophycocyanin, and 22,000 and 20,000 for phycoerythrin. Covalently bound chromophore was associated with each subunit.

  5. Conditional mutations occur predominantly in highly conserved residues of RNA polymerase II subunits.

    PubMed Central

    Scafe, C; Martin, C; Nonet, M; Podos, S; Okamura, S; Young, R A

    1990-01-01

    Conditional mutations in the Saccharomyces cerevisiae RNA polymerase II large subunit, RPB1, were obtained by introducing a mutagenized RPB1 plasmid into yeast cells, selecting for loss of the wild-type RPB1 gene, and screening the cells for heat or cold sensitivity. Sequence analysis of 10 conditional RPB1 mutations and 10 conditional RPB2 mutations revealed that the amino acid residues altered by these distinct mutations are nearly always invariant among eucaryotic RPB1 and RPB2 homologs. These results suggest that RNA polymerase mutants might be obtained in other eucaryotic organisms by alteration of these invariant residues. Images PMID:2406567

  6. Phylogenetic classification of the frog pathogen Amphibiothecum (Dermosporidium) penneri based on small ribosomal subunit sequencing

    USGS Publications Warehouse

    Feldman, S.H.; Wimsatt, J.H.; Green, D.E.

    2005-01-01

    We determined 1,600 base pairs of DNA sequence in the 18S small ribosomal subunit from two geographically distinct isolates of Dermosporidium penneri. Maximum likelihood and parsimony analysis of these sequences place D. penneri in the order Dermocystida of the class Mesomycetozoea. The 18S rRNA sequences from these two isolates only differ within a single region of 16 contiguous nucleotides. Based on the distant phylogenetic relationship of these organisms to Amphibiocystidium ranae and similarity to Sphaerothecum destruens we propose the organism be renamed Amphibiothecum penneri.

  7. Cellular identity of a novel small subunit rDNA sequence clade of apicomplexans: description of the marine parasite Rhytidocystis polygordiae n. sp. (host: Polygordius sp., Polychaeta).

    PubMed

    Leander, Brian S; Ramey, Patricia A

    2006-01-01

    A new species of Rhytidocystis (Apicomplexa) is characterized from North American waters of the Atlantic Ocean using electron microscopy and phylogenetic analyses of small subunit (SSU) rDNA sequences. Rhytidocystis polygordiae n. sp. is a parasite of the polychaete Polygordius sp. and becomes the fourth described species within this genus. The trophozoites of R. polygordiae were relatively small oblong cells (L=35-55 microm; W=20-25 microm) and distinctive in possessing subterminal indentations at both ends of the cell. The surface of the trophozoites had six to eight longitudinal series of small transverse folds and several micropores arranged in short linear rows. The trophozoites of R. polygordiae were positioned beneath the brush border of the intestinal epithelium but appeared to reside between the epithelial cells within the extracellular matrix rather than within the cells. The trophozoites possessed a uniform distribution of paraglycogen granules, putative apicoplasts, mitochondria with tubular cristae, and a centrally positioned nucleus. The trophozoites were non-motile and lacked a mucron and an apical complex. Intracellular sporozoites of R. polygordiae had a conoid, a few rhoptries, micronemes, dense granules, and a posteriorly positioned nucleus. Phylogenies inferred from SSU rDNA sequences demonstrated a close relationship between R. polygordiae and the poorly known parasite reported from the hemolymph of the giant clam Tridacna crocea. The rhytidocystid clade diverged early in the apicomplexan radiation and showed a weak affinity to a clade consisting of cryptosporidian parasites, monocystids, and neogregarines.

  8. Gingival Tissue Transcriptomes Identify Distinct Periodontitis Phenotypes

    PubMed Central

    Kebschull, M.; Demmer, R.T.; Grün, B.; Guarnieri, P.; Pavlidis, P.; Papapanou, P.N.

    2014-01-01

    The currently recognized principal forms of periodontitis—chronic and aggressive—lack an unequivocal, pathobiology-based foundation. We explored whether gingival tissue transcriptomes can serve as the basis for an alternative classification of periodontitis. We used cross-sectional whole-genome gene expression data from 241 gingival tissue biopsies obtained from sites with periodontal pathology in 120 systemically healthy nonsmokers with periodontitis, with available data on clinical periodontal status, subgingival microbial profiles, and serum IgG antibodies to periodontal microbiota. Adjusted model-based clustering of transcriptomic data using finite mixtures generated two distinct clusters of patients that did not align with the current classification of chronic and aggressive periodontitis. Differential expression profiles primarily related to cell proliferation in cluster 1 and to lymphocyte activation and unfolded protein responses in cluster 2. Patients in the two clusters did not differ with respect to age but presented with distinct phenotypes (statistically significantly different whole-mouth clinical measures of extent/severity, subgingival microbial burden by several species, and selected serum antibody responses). Patients in cluster 2 showed more extensive/severe disease and were more often male. The findings suggest that distinct gene expression signatures in pathologic gingival tissues translate into phenotypic differences and can provide a basis for a novel classification. PMID:24646639

  9. P. berghei telomerase subunit TERT is essential for parasite survival.

    PubMed

    Religa, Agnieszka A; Ramesar, Jai; Janse, Chris J; Scherf, Artur; Waters, Andrew P

    2014-01-01

    Telomeres define the ends of chromosomes protecting eukaryotic cells from chromosome instability and eventual cell death. The complex regulation of telomeres involves various proteins including telomerase, which is a specialized ribonucleoprotein responsible for telomere maintenance. Telomeres of chromosomes of malaria parasites are kept at a constant length during blood stage proliferation. The 7-bp telomere repeat sequence is universal across different Plasmodium species (GGGTTT/CA), though the average telomere length varies. The catalytic subunit of telomerase, telomerase reverse transcriptase (TERT), is present in all sequenced Plasmodium species and is approximately three times larger than other eukaryotic TERTs. The Plasmodium RNA component of TERT has recently been identified in silico. A strategy to delete the gene encoding TERT via double cross-over (DXO) homologous recombination was undertaken to study the telomerase function in P. berghei. Expression of both TERT and the RNA component (TR) in P. berghei blood stages was analysed by Western blotting and Northern analysis. Average telomere length was measured in several Plasmodium species using Telomere Restriction Fragment (TRF) analysis. TERT and TR were detected in blood stages and an average telomere length of ∼ 950 bp established. Deletion of the tert gene was performed using standard transfection methodologies and we show the presence of tert- mutants in the transfected parasite populations. Cloning of tert- mutants has been attempted multiple times without success. Thorough analysis of the transfected parasite populations and the parasite obtained from extensive parasite cloning from these populations provide evidence for a so called delayed death phenotype as observed in different organisms lacking TERT. The findings indicate that TERT is essential for P. berghei cell survival. The study extends our current knowledge on telomere biology in malaria parasites and validates further investigations to

  10. Compilation of small ribosomal subunit RNA structures.

    PubMed Central

    Neefs, J M; Van de Peer, Y; De Rijk, P; Chapelle, S; De Wachter, R

    1993-01-01

    The database on small ribosomal subunit RNA structure contained 1804 nucleotide sequences on April 23, 1993. This number comprises 365 eukaryotic, 65 archaeal, 1260 bacterial, 30 plastidial, and 84 mitochondrial sequences. These are stored in the form of an alignment in order to facilitate the use of the database as input for comparative studies on higher-order structure and for reconstruction of phylogenetic trees. The elements of the postulated secondary structure for each molecule are indicated by special symbols. The database is available on-line directly from the authors by ftp and can also be obtained from the EMBL nucleotide sequence library by electronic mail, ftp, and on CD ROM disk. PMID:8332525

  11. Descriptions of two azooxanthellate Palythoa species (Subclass Hexacorallia, Order Zoantharia) from the Ryukyu Archipelago, southern Japan.

    PubMed

    Irei, Yuka; Sinniger, Frederic; Reimer, James Davis

    2015-01-01

    Two new species of zoantharians (Hexacorallia, Zoantharia, Sphenopidae), Palythoamizigama sp. n. and Palythoaumbrosa sp. n., are described from the Ryukyu Archipelago, southern Japan. Unlike almost all other known Palythoa spp., both species are azooxanthellate and inhabit low-light environments such as floors or sides of caves, crevasses, or hollows of shallow coral reefs. The two species were initially considered to be the same species from their similar habitat environments and highly similar morphological features. However, phylogenetic analyses of nuclear internal transcribed spacer (ITS) ribosomal DNA, mitochondrial 16S ribosomal DNA, and cytochrome oxidase subunit I (COI) sequences revealed that these two species have a genetically distant relationship within the genus Palythoa. Morphological characteristics, including polyp size, tentacle number, external/internal coloration, and types and sizes of cnidae were examined in this study. As a result, only tentacle coloration was found to be useful for the morphological distinction between the two species. Palythoamizigama possesses white tentacles with black horizontal stripes while Palythoaumbrosa possesses white tentacles without any stripe patterns. Considering their distant phylogenetic relationship, it can be assumed that their unique yet similar morphological and ecological characteristics developed independently in each species as an example of parallel evolution. PMID:25685008

  12. DNA Barcodes for Species Identification in the Hyperdiverse Ant Genus Pheidole (Formicidae: Myrmicinae)

    PubMed Central

    Ng'endo, R.N.; Osiemo, Z.B.; Brandl, R.

    2013-01-01

    DNA sequencing is increasingly being used to assist in species identification in order to overcome taxonomic impediment. However, few studies attempt to compare the results of these molecular studies with a more traditional species delineation approach based on morphological characters. Mitochondrial DNA Cytochrome oxidase subunit 1 (CO1) gene was sequenced, measuring 636 base pairs, from 47 ants of the genus Pheidole (Formicidae: Myrmicinae) collected in the Brazilian Atlantic Forest to test whether the morphology-based assignment of individuals into species is supported by DNA-based species delimitation. Twenty morphospecies were identified, whereas the barcoding analysis identified 19 Molecular Operational Taxonomic Units (MOTUs). Fifteen out of the 19 DNA-based clusters allocated, using sequence divergence thresholds of 2% and 3%, matched with morphospecies. Both thresholds yielded the same number of MOTUs. Only one MOTU was successfully identified to species level using the CO1 sequences of Pheidole species already in the Genbank. The average pairwise sequence divergence for all 47 sequences was 19%, ranging between 0–25%. In some cases, however, morphology and molecular based methods differed in their assignment of individuals to morphospecies or MOTUs. The occurrence of distinct mitochondrial lineages within morphological species highlights groups for further detailed genetic and morphological studies, and therefore a pluralistic approach using several methods to understand the taxonomy of difficult lineages is advocated. PMID:23902257

  13. An integrative taxonomy on the locally endangered species of the Korean Scarabaeus (Coleoptera, Scarabaeidae).

    PubMed

    Han, Taeman; Kim, Jin Ill; Yi, Dae-Am; Jeong, Jongchel; An, Seung Lak; Park, In Gyun; Park, Haechul

    2016-01-01

    The ball-rolling dung beetles of the genus Scarabaeus are very ecologically important for the recycling of feces of large herbivores and the related nature management. There has been a significant decline, however, in the numbers of many species at the population and individual levels. S. typhon is currently thought to be the sole member of Scarabaeus distributed in Korea; however, that species underwent serious local extinctions in the 1970s. Before planning a full-scale species recovery, it is important to have an understanding of the exact species diversity and genetic structures of the focal species. We therefore attempted an integrative taxonomy focused on the Korean population of S. typhon and also on S. pius and S. sacer, which were once thought to be distributed in Korea, using both morphological and molecular approaches. The results of both approaches reveal the Korean species of Scarabaeus to be S. typhon and S. pius. In particular, our molecular results inferred from cytochrome c oxidase subunit I genetic analysis show that S. typhon should be considered a single species despite having various haplotypes throughout its wide geographical range from Europe to Korea. We identified two distinct lineages of S. pius (groups A and B) across a wide distributional range. We conclude that the Korean specimens of S. pius belong to group A and that S. pius is new to Korea under the current taxonomic treatment. PMID:27470822

  14. An integrative taxonomy on the locally endangered species of the Korean Scarabaeus (Coleoptera, Scarabaeidae).

    PubMed

    Han, Taeman; Kim, Jin Ill; Yi, Dae-Am; Jeong, Jongchel; An, Seung Lak; Park, In Gyun; Park, Haechul

    2016-07-22

    The ball-rolling dung beetles of the genus Scarabaeus are very ecologically important for the recycling of feces of large herbivores and the related nature management. There has been a significant decline, however, in the numbers of many species at the population and individual levels. S. typhon is currently thought to be the sole member of Scarabaeus distributed in Korea; however, that species underwent serious local extinctions in the 1970s. Before planning a full-scale species recovery, it is important to have an understanding of the exact species diversity and genetic structures of the focal species. We therefore attempted an integrative taxonomy focused on the Korean population of S. typhon and also on S. pius and S. sacer, which were once thought to be distributed in Korea, using both morphological and molecular approaches. The results of both approaches reveal the Korean species of Scarabaeus to be S. typhon and S. pius. In particular, our molecular results inferred from cytochrome c oxidase subunit I genetic analysis show that S. typhon should be considered a single species despite having various haplotypes throughout its wide geographical range from Europe to Korea. We identified two distinct lineages of S. pius (groups A and B) across a wide distributional range. We conclude that the Korean specimens of S. pius belong to group A and that S. pius is new to Korea under the current taxonomic treatment.

  15. Malic enzymes of Trichomonas vaginalis: two enzyme families, two distinct origins.

    PubMed

    Dolezal, Pavel; Vanácová, Stepánka; Tachezy, Jan; Hrdý, Ivan

    2004-03-31

    The cytosolic malic enzyme of the amitochondriate protist Trichomonas vaginalis was purified to homogeneity and characterized. The corresponding gene was sequenced and compared with its hydrogenosomal homologue from the same organism. The enzymes were found to differ in coenzyme specificity, molecular mass and physiological role. The cytosolic malic enzyme is a dimer consisting of two 42-kDa subunits with strict specificity for nicotinamide adenine dinucleotide phosphate (NADP(+)), and has a presumed function of pyruvate and NADPH production. The hydrogenosomal malic enzyme is a tetramer of 60-kDa subunits that preferentially utilizes nicotinamide adenine dinucleotide (NAD(+)) to NADP(+). The hydrogenosomal enzyme supplies the hydrogenosome with pyruvate for further catabolic processes linked with substrate-level phosphorylation. Phylogenetic analysis of malic enzymes showed the existence of two distinct families of these enzymes in nature, which differ in subunit size. The trichomonad cytosolic malic enzyme belongs to the small subunit-type family that occurs almost exclusively in prokaryotes. In contrast, the hydrogenosomal malic enzyme displays a close relationship with the large subunit-type family of the enzyme, which is found in mitochondria, plastids and the cytosol of eukaryotes. The eubacterial origin of trichomonad cytosolic malic enzyme suggests an occurrence of horizontal gene transfer from a eubacterium to the ancestor of T. vaginalis. The presence of both prokaryotic and eukaryotic type of malic enzyme in different compartments of a single eukaryotic cell appears to be unique in nature. PMID:15033531

  16. Cloning of eight Rhopalosiphum padi (Hemiptera: Aphididae) nAChR subunit genes and mutation detection of the β1 subunit in field samples from China.

    PubMed

    Zhang, Meng; Qiao, Xianfeng; Li, Yuting; Fang, Bing; Zuo, Yayun; Chen, Maohua

    2016-09-01

    The bird cherry-oat aphid, Rhopalosiphum padi (L.), is one of the most important wheat pests. This aphid damages through direct feeding and by transmitting the Barley yellow dwarf virus (BYDV). Both types of damage significantly reduce the quality and yield of wheat crops globally. Insecticides are the primary method of controlling the bird cherry-oat aphid in China, yet this aphid species has developed resistance to different types of insecticides, especially organophosphates and carbamates. In the last decade, control of R. padi depends primarily on the spray of neonicotinoid insecticides, however, research on the resistance of R. padi to neonicotinoids has been limited. In this study, the full lengths of seven α-subunit (Rpα1, Rpα2, Rpα3, Rpα4, Rpα5, Rpα7-1, and Rpα7-2) and one β-subunit (Rpβ1) genes from R. padi were obtained with RT-PCR and RACE techniques. Sequence analysis showed that these genes had all the characteristics of the nAChR gene family and were highly homologous with the reported nAChR genes from other insects, and alternative splicing was detected in Rpα3 and Rpα5 subunits. Analysis of the cDNA sequence of the extracellular region of the nicotinic acetylcholine receptor β1 subunit gene from 120 R. padi field samples collected in 11 Provinces revealed 17 single nucleotides polymorphism (SNP) sites, of which seven were amino acid polymorphism sites (V53I, V53G, N54T, A60T, F61L, W79C, and V83I) and two were in the loop D region (W79C and V83I). The current study will facilitate further studies on the molecular mechanisms of targeted resistance of the aphid to neonicotinoid insecticides. PMID:27521918

  17. Regulatory–auxiliary subunits of CLC chloride channel–transport proteins

    PubMed Central

    Barrallo-Gimeno, Alejandro; Gradogna, Antonella; Zanardi, Ilaria; Pusch, Michael; Estévez, Raúl

    2015-01-01

    Abstract The CLC family of chloride channels and transporters is composed by nine members, but only three of them, ClC-Ka/b, ClC-7 and ClC-2, have been found so far associated with auxiliary subunits. These CLC regulatory subunits are small proteins that present few common characteristics among them, both structurally and functionally, and their effects on the corresponding CLC protein are different. Barttin, a protein with two transmembrane domains, is essential for the membrane localization of ClC-K proteins and their activity in the kidney and inner ear. Ostm1 is a protein with a single transmembrane domain and a highly glycosylated N-terminus. Unlike the other two CLC auxiliary subunits, Ostm1 shows a reciprocal relationship with ClC-7 for their stability. The subcellular localization of Ostm1 depends on ClC-7 and not the other way around. ClC-2 is active on its own, but GlialCAM, a transmembrane cell adhesion molecule with two extracellular immunoglobulin (Ig)-like domains, regulates its subcellular localization and activity in glial cells. The common theme for these three proteins is their requirement for a proper homeostasis, since their malfunction leads to distinct diseases. We will review here their properties and their role in normal chloride physiology and the pathological consequences of their improper function. PMID:25762128

  18. Specific inhibition of NF-Y subunits triggers different cell proliferation defects

    PubMed Central

    Benatti, Paolo; Dolfini, Diletta; Viganò, Alessandra; Ravo, Maria; Weisz, Alessandro; Imbriano, Carol

    2011-01-01

    Regulated gene expression is essential for a proper progression through the cell cycle. The transcription factor NF-Y has a fundamental function in transcriptional regulation of cell cycle genes, particularly of G2/M genes. In order to investigate common and distinct functions of NF-Y subunits in cell cycle regulation, NF-YA, NF-YB and NF-YC have been silenced by shRNAs in HCT116 cells. NF-YA loss led to a delay in S-phase progression, DNA damage and apoptosis: we showed the activation of the replication checkpoint, through the recruitment of Δp53 and of the replication proteins PCNA and Mcm7 to chromatin. Differently, NF-YB depletion impaired cells from exiting G2/M, but did not interfere with S-phase progression. Gene expression analysis of NF-YA and NF-YB inactivated cells highlighted a common set of hit genes, as well as a plethora of uncommon genes, unveiling a different effect of NF-Y subunits loss on NF-Y binding to its target genes. Chromatin extracts and ChIP analysis showed that NF-YA depletion was more effective than NF-YB in hitting NF-Y recruitment to CCAAT-promoters. Our data suggest a critical role of NF-Y expression, highlighting that the lack of the single subunits are differently perceived by the cells, which activate diverse cell cycle blocks and signaling pathways. PMID:21415014

  19. Regulation of the mammalian elongation cycle by subunit rolling: a eukaryotic-specific ribosome rearrangement

    PubMed Central

    Budkevich, Tatyana V.; Giesebrecht, Jan; Behrmann, Elmar; Loerke, Justus; Ramrath, David J.F.; Mielke, Thorsten; Ismer, Jochen; Hildebrand, Peter W.; Tung, Chang-Shung; Nierhaus, Knud H.; Sanbonmatsu, Karissa Y.; Spahn, Christian M.T.

    2014-01-01

    SUMMARY The extent to which bacterial ribosomes and the significantly larger eukaryotic ribosomes share the same mechanisms of ribosomal elongation is unknown. Here, we present sub-nanometer resolution cryo-electron microscopy maps of the mammalian 80S ribosome in the post-translocational state and in complex with the eukaryotic eEF1A•Val-tRNA•GMPPNP ternary complex, revealing significant differences in the elongation mechanism between bacteria and mammals. Surprisingly, and in contrast to bacterial ribosomes, a rot