CD147 regulates the expression of MCT1 and lactate export in multiple myeloma cells

PubMed Central

Walters, Denise K; Arendt, Bonnie K; Jelinek, Diane F

2013-01-01

Increased use of the glycolytic pathway, even in the presence of oxygen, has recently been recognized as a key characteristic of malignant cells. However, the glycolytic phenotype results in increased lactic acid production and, in order to prevent cellular acidosis, tumor cells must increase proton efflux via upregulation of pH regulators such as proton-pumps, sodium-proton exchangers, and/or monocarboxylate transporters (MCT) (e.g., MCT1, MCT4). Interestingly, expression of MCT1 and MCT4 has been previously shown to be dependent upon expression of the transmembrane glycoprotein CD147. Recently, we demonstrated that primary patient multiple myeloma (MM) cells and human MM cell lines (HMCLs) overexpress CD147. Therefore, the goal of the current study was to specifically determine if MCT1 and MCT4 were also overexpressed in MM cells. RT-PCR analysis demonstrated both primary patient MM cells and HMCLs overexpress MCT1 and MCT4 mRNA. Notably, primary MM cells or HMCLs were found to express variable levels of MCT1 and/or MCT4 at the protein level despite CD147 expression. In those HMCLs positive for MCT1 and/or MCT4 protein expression, MCT1 and/or MCT4 were found to be associated with CD147. Specific siRNA-mediated downregulation of MCT1 but not MCT4 resulted in decreased HMCL proliferation, decreased lactate export, and increased cellular media pH. However, western blot analysis revealed that downregulation of MCT1 also downregulated CD147 and vice versa despite no effect on mRNA levels. Taken together, these data demonstrate the association between MCT1 and CD147 proteins in MM cells and importance of their association for lactate export and proliferation in MM cells. PMID:24013424

CD147 regulates the expression of MCT1 and lactate export in multiple myeloma cells.

PubMed

Walters, Denise K; Arendt, Bonnie K; Jelinek, Diane F

2013-10-01

Increased use of the glycolytic pathway, even in the presence of oxygen, has recently been recognized as a key characteristic of malignant cells. However, the glycolytic phenotype results in increased lactic acid production and, in order to prevent cellular acidosis, tumor cells must increase proton efflux via upregulation of pH regulators such as proton-pumps, sodium-proton exchangers, and/or monocarboxylate transporters (MCT) (e.g., MCT1, MCT4). Interestingly, expression of MCT1 and MCT4 has been previously shown to be dependent upon expression of the transmembrane glycoprotein CD147. Recently, we demonstrated that primary patient multiple myeloma (MM) cells and human MM cell lines (HMCLs) overexpress CD147. Therefore, the goal of the current study was to specifically determine if MCT1 and MCT4 were also overexpressed in MM cells. RT-PCR analysis demonstrated both primary patient MM cells and HMCLs overexpress MCT1 and MCT4 mRNA. Notably, primary MM cells or HMCLs were found to express variable levels of MCT1 and/or MCT4 at the protein level despite CD147 expression. In those HMCLs positive for MCT1 and/or MCT4 protein expression, MCT1 and/or MCT4 were found to be associated with CD147. Specific siRNA-mediated downregulation of MCT1 but not MCT4 resulted in decreased HMCL proliferation, decreased lactate export, and increased cellular media pH. However, western blot analysis revealed that downregulation of MCT1 also downregulated CD147 and vice versa despite no effect on mRNA levels. Taken together, these data demonstrate the association between MCT1 and CD147 proteins in MM cells and importance of their association for lactate export and proliferation in MM cells.

Structures of the TetR-like simocyclinone efflux pump repressor, SimR, and the mechanism of ligand-mediated derepression.

PubMed

Le, Tung B K; Stevenson, Clare E M; Fiedler, Hans-Peter; Maxwell, Anthony; Lawson, David M; Buttner, Mark J

2011-04-22

Simocyclinone D8 (SD8), a potent DNA gyrase inhibitor made by Streptomyces antibioticus, is exported from the producing organism by the SimX efflux pump. The expression of simX is under the control of SimR, a member of the TetR family of transcriptional regulators. SimR represses simX transcription by binding to operators in the intergenic region between simR and simX. Previously, we have shown that the mature antibiotic SD8 or its biosynthetic intermediate, simocyclinone C4, can dissociate SimR from its operators, leading to derepression of simX and export of SD8 from the cell. This provides a mechanism that couples the biosynthesis of the antibiotic to its export. Here, we report the crystal structures of SimR alone and in complex with either SD8 or simocyclinone C4. The ligand-binding pocket is unusual compared to those of other characterized TetR-family transcriptional regulators: the structures show an extensive ligand-binding pocket spanning both monomers in the functional dimeric unit, with the aminocoumarin moiety of SD8 buried in the protein core, while the angucyclic polyketide moiety is partially exposed to bulk solvent. Through comparisons of the structures, we postulate a derepression mechanism for SimR that invokes rigid-body motions of the subunits relative to one another, coupled with a putative locking mechanism to restrict further conformational change. Copyright © 2011 Elsevier Ltd. All rights reserved.

Wild Horse 69-kV transmission line environmental assessment

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

NONE

1996-12-01

Hill County Electric Cooperative Inc. (Hill County) proposes to construct and operate a 69-kV transmission line from its North Gildford Substation in Montana north to the Canadian border. A vicinity project area map is enclosed as a figure. TransCanada Power Corporation (TCP), a Canadian power-marketing company, will own and construct the connecting 69-kV line from the international border to Express Pipeline`s pump station at Wild Horse, Alberta. This Environmental Assessment is prepared for the Department of Energy (DOE) as lead federal agency to comply with the requirements of the National Environmental Policy Act (NEPA), as part of DOE`s review andmore » approval process of the applications filed by Hill County for a DOE Presidential Permit and License to Export Electricity to a foreign country. The purpose of the proposed line is to supply electric energy to a crude oil pump station in Canada, owned by Express Pipeline Ltd. (Express). The pipeline would transport Canadian-produced oil from Hardisty, Alberta, Canada, to Caster, Wyoming. The Express Pipeline is scheduled to be constructed in 1996--97 and will supply crude oil to refineries in Wyoming and the midwest.« less

Short- and medium-chain fatty acids enhance the cell surface expression and transport capacity of the bile salt export pump (BSEP/ABCB11).

PubMed

Kato, Takuya; Hayashi, Hisamitsu; Sugiyama, Yuichi

2010-09-01

The reduced expression of the bile salt export pump (BSEP/ABCB11) at the canalicular membrane is associated with cholestasis-induced hepatotoxicity due to the accumulation of bile acids in hepatocytes. We previously reported that 4-phenylbutyrate (4PBA), an approved drug for urea cycle disorders, is a promising agent for intrahepatic cholestasis because it increases both the cell surface expression and the transport capacity of BSEP. In the present study, we searched for effective compounds other than 4PBA by focusing on short- and medium-chain fatty acids, which have similar characteristics to 4PBA such as their low-molecular-weight and a carboxyl group. In transcellular transport studies using Madin-Darby canine kidney (MDCK) II cells, all short- and medium-chain fatty acids tested except for formate, acetate, and hexanoic acid showed more potent effects on wild type (WT) BSEP-mediated [3H]taurocholate transport than did 4PBA. The increase in WT BSEP transport with butyrate and octanoic acid treatment correlated with an increase in its expression at the cell surface. Two PFIC2-type variants, E297G and D482G BSEP, were similarly affected with both compounds treatment. The prolonged half-life of cell surface-resident WT BSEP was responsible for this increased octanoic acid-stimulated transport, but not for that of butyrate. In conclusion, short- and medium-chain fatty acids have potent effects on the increase in WT and PFIC2-type BSEP-mediated transport in MDCK II cells. Although both short- and medium-chain fatty acids enhance the transport capacity of WT and PFIC2-type BSEP by inducing those expressions at the cell surface, the underlying mechanism seems to differ between fatty acids. 2010 Elsevier B.V. All rights reserved.

The dynamic ocean biological pump: Insights from a global compilation of particulate organic carbon, CaCO3, and opal concentration profiles from the mesopelagic

NASA Astrophysics Data System (ADS)

Lam, Phoebe J.; Doney, Scott C.; Bishop, James K. B.

2011-09-01

We have compiled a global data set of 62 open ocean profiles of particulate organic carbon (POC), CaCO3, and opal concentrations collected by large volume in situ filtration in the upper 1000 m over the last 30 years. We define concentration-based metrics for the strength (POC concentration at depth) and efficiency (attenuation of POC with depth in the mesopelagic) of the biological pump. We show that the strength and efficiency of the biological pump are dynamic and are characterized by a regime of constant and high transfer efficiency at low to moderate surface POC and a bloom regime where the height of the bloom is characterized by a weak deep biological pump and low transfer efficiency. The variability in POC attenuation length scale manifests in a clear decoupling between the strength of the shallow biological pump (e.g., POC at the export depth) and the strength of the deep biological pump (POC at 500 m). We suggest that the paradigm of diatom-driven export production is driven by a too restrictive perspective on upper mesopelagic dynamics. Indeed, our full mesopelagic analysis suggests that large, blooming diatoms have low transfer efficiency and thus may not export substantially to depth; rather, our analysis suggests that ecosystems characterized by smaller cells and moderately high %CaCO3 have a high mesopelagic transfer efficiency and can have higher POC concentrations in the deep mesopelagic even with relatively low surface or near-surface POC. This has negative implications for the carbon sequestration prospects of deliberate iron fertilization.

76 FR 16842 - Request for a License To Export Reactor Components

Federal Register 2010, 2011, 2012, 2013, 2014

2011-03-25

... NUCLEAR REGULATORY COMMISSION Request for a License To Export Reactor Components Pursuant to 10.... Mechanical Corporation. coolant pump 1000 (design) maintenance, and systems, related reactors. operation of AP- equipment, and 1000 (design) spare parts. nuclear reactors. February 10, 2011 February 23, 2011...

Characterization and regulation of the resistance-nodulation-cell division-type multidrug efflux pumps MdtABC and MdtUVW from the fire blight pathogen Erwinia amylovora.

PubMed

Pletzer, Daniel; Weingart, Helge

2014-07-11

The Gram-negative bacterium Erwinia amylovora is the causal agent of the devastating disease fire blight in rosaceous plants such as apple, pear, quince, raspberry, and cotoneaster. In order to survive and multiply in a host, microbes must be able to circumvent the toxic effects of antimicrobial plant compounds, such as flavonoids and tannins. E. amylovora uses multidrug efflux transporters that recognize and actively export toxic compounds out of the cells. Here, two heterotrimeric resistance-nodulation-cell division (RND)-type multidrug efflux pumps, MdtABC and MdtUVW, from E. amylovora were identified. These RND systems are unusual in that they contain two different RND proteins forming a functional pump. To find the substrate specificities of the two efflux systems, we overexpressed the transporters in a hypersensitive mutant lacking the major RND pump AcrB. Both transporters mediated resistance to several flavonoids, fusidic acid and novobiocin. Additionally, MdtABC mediated resistance towards josamycin, bile salts and silver nitrate, and MdtUVW towards clotrimazole. The ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock was reduced. Quantitative RT-PCR analyses revealed that the expression of the transporter genes was induced during infection of apple rootstock. The polyphenolic plant compound tannin, as well as the heavy metal salt tungstate was found to induce the expression of mdtABC. Finally, the expression of the mdtABC genes was shown to be regulated by BaeR, the response regulator of the two-component system BaeSR, a cell envelope stress response system that controls the adaptive responses to changes in the environment. The expression of MdtABC and MdtUVW is induced during growth of E. amylovora in planta. We identified the plant polyphenol tannin as inducer of mdtABC expression. The reduced ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock suggests that the efflux pumps are involved in resistance to plant antimicrobials, maybe including flavonoids, which were identified as substrates of both pumps. Furthermore, we found that the mdtABC operon belongs to the regulon of the two-component regulator BaeR suggesting a role of this RND transporter in the cell envelope stress response of E. amylovora.

Characterization and regulation of the Resistance-Nodulation-Cell Division-type multidrug efflux pumps MdtABC and MdtUVW from the fire blight pathogen Erwinia amylovora

PubMed Central

2014-01-01

Background The Gram-negative bacterium Erwinia amylovora is the causal agent of the devastating disease fire blight in rosaceous plants such as apple, pear, quince, raspberry, and cotoneaster. In order to survive and multiply in a host, microbes must be able to circumvent the toxic effects of antimicrobial plant compounds, such as flavonoids and tannins. E. amylovora uses multidrug efflux transporters that recognize and actively export toxic compounds out of the cells. Here, two heterotrimeric resistance-nodulation-cell division (RND)-type multidrug efflux pumps, MdtABC and MdtUVW, from E. amylovora were identified. These RND systems are unusual in that they contain two different RND proteins forming a functional pump. Results To find the substrate specificities of the two efflux systems, we overexpressed the transporters in a hypersensitive mutant lacking the major RND pump AcrB. Both transporters mediated resistance to several flavonoids, fusidic acid and novobiocin. Additionally, MdtABC mediated resistance towards josamycin, bile salts and silver nitrate, and MdtUVW towards clotrimazole. The ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock was reduced. Quantitative RT-PCR analyses revealed that the expression of the transporter genes was induced during infection of apple rootstock. The polyphenolic plant compound tannin, as well as the heavy metal salt tungstate was found to induce the expression of mdtABC. Finally, the expression of the mdtABC genes was shown to be regulated by BaeR, the response regulator of the two-component system BaeSR, a cell envelope stress response system that controls the adaptive responses to changes in the environment. Conclusions The expression of MdtABC and MdtUVW is induced during growth of E. amylovora in planta. We identified the plant polyphenol tannin as inducer of mdtABC expression. The reduced ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock suggests that the efflux pumps are involved in resistance to plant antimicrobials, maybe including flavonoids, which were identified as substrates of both pumps. Furthermore, we found that the mdtABC operon belongs to the regulon of the two-component regulator BaeR suggesting a role of this RND transporter in the cell envelope stress response of E. amylovora. PMID:25012600

Antibiotic Resistance Mediated by the MacB ABC Transporter Family: A Structural and Functional Perspective.

PubMed

Greene, Nicholas P; Kaplan, Elise; Crow, Allister; Koronakis, Vassilis

2018-01-01

The MacB ABC transporter forms a tripartite efflux pump with the MacA adaptor protein and TolC outer membrane exit duct to expel antibiotics and export virulence factors from Gram-negative bacteria. Here, we review recent structural and functional data on MacB and its homologs. MacB has a fold that is distinct from other structurally characterized ABC transporters and uses a unique molecular mechanism termed mechanotransmission. Unlike other bacterial ABC transporters, MacB does not transport substrates across the inner membrane in which it is based, but instead couples cytoplasmic ATP hydrolysis with transmembrane conformational changes that are used to perform work in the extra-cytoplasmic space. In the MacAB-TolC tripartite pump, mechanotransmission drives efflux of antibiotics and export of a protein toxin from the periplasmic space via the TolC exit duct. Homologous tripartite systems from pathogenic bacteria similarly export protein-like signaling molecules, virulence factors and siderophores. In addition, many MacB-like ABC transporters do not form tripartite pumps, but instead operate in diverse cellular processes including antibiotic sensing, cell division and lipoprotein trafficking.

Global patterns of organic carbon export and sequestration in the ocean (Arne Richter Award for Outstanding Young Scientists)

NASA Astrophysics Data System (ADS)

Henson, S.; Sanders, R.; Madsen, E.; Le Moigne, F.; Quartly, G.

2012-04-01

A major term in the global carbon cycle is the ocean's biological carbon pump which is dominated by sinking of small organic particles from the surface ocean to its interior. Here we examine global patterns in particle export efficiency (PEeff), the proportion of primary production that is exported from the surface ocean, and transfer efficiency (Teff), the fraction of exported organic matter that reaches the deep ocean. This is achieved through extrapolating from in situ estimates of particulate organic carbon export to the global scale using satellite-derived data. Global scale estimates derived from satellite data show, in keeping with earlier studies, that PEeff is high at high latitudes and low at low latitudes, but that Teff is low at high latitudes and high at low latitudes. However, in contrast to the relationship observed for deep biomineral fluxes in previous studies, we find that Teff is strongly negatively correlated with opal export flux from the upper ocean, but uncorrelated with calcium carbonate export flux. We hypothesise that the underlying factor governing the spatial patterns observed in Teff is ecosystem function, specifically the degree of recycling occurring in the upper ocean, rather than the availability of calcium carbonate for ballasting. Finally, our estimate of global integrated carbon export is only 50% of previous estimates. The lack of consensus amongst different methodologies on the strength of the biological carbon pump emphasises that our knowledge of a major planetary carbon flux remains incomplete.

Impact of the natural Fe-fertilization on the magnitude, stoichiometry and efficiency of particulate biogenic silica, nitrogen and iron export fluxes

NASA Astrophysics Data System (ADS)

Lemaitre, N.; Planquette, H.; Dehairs, F.; van der Merwe, P.; Bowie, A. R.; Trull, T. W.; Laurenceau-Cornec, E. C.; Davies, D.; Bollinger, C.; Le Goff, M.; Grossteffan, E.; Planchon, F.

2016-11-01

The Kerguelen Plateau is characterized by a naturally Fe-fertilized phytoplankton bloom that extends more than 1000 km downstream in the Antarctic Circumpolar Current. During the KEOPS2 study, in austral spring, we measured particulate nitrogen (PN), biogenic silica (BSi) and particulate iron (PFe) export fluxes in order to investigate how the natural fertilization impacts the stoichiometry and the magnitude of export fluxes and therefore the efficiency of the biological carbon pump. At 9 stations, we estimated elemental export fluxes based on element concentration to 234Th activity ratios for particulate material collected with in-situ pumps and 234Th export fluxes (Planchon et al., 2015). This study revealed that the natural Fe-fertilization increased export fluxes but to variable degrees. Export fluxes for the bloom impacted area were compared with those of a high-nutrient, low-chlorophyll (HNLC), low-productive reference site located to the south-west of Kerguelen and which had the lowest BSi and PFe export fluxes (2.55 mmol BSi m-2 d-1 and 1.92 μmol PFem-2 d-1) and amongst the lowest PN export flux (0.73 mmol PN m-2 d-1). The impact of the Fe fertilization was the greatest within a meander of the polar front (PF), to the east of Kerguelen, with fluxes reaching 1.26 mmol PN m-2 d-1; 20.4 mmol BSi m-2 d-1 and 22.4 μmol PFe m-2 d-1. A highly productive site above the Kerguelen Plateau, on the contrary, was less impacted by the fertilization with export fluxes reaching 0.72 mmol PN m-2 d-1; 4.50 mmol BSi m-2 d-1 and 21.4 μmol PFe m-2 d-1. Our results suggest that ecosystem features (i.e. type of diatom community) could play an important role in setting the magnitude of export fluxes of these elements. Indeed, for the PF meander, the moderate productivity was sustained by the presence of large and strongly silicified diatom species while at the higher productivity sites, smaller and slightly silicified diatoms dominated. Interestingly, our results suggest that PFe export fluxes can be driven by the lithogenic pool of particles, especially over the Plateau where such inputs from the sediments are important. Finally, for the Plateau and the PF meander, the comparison between PFe export and the particulate PFe stock integrated over the mixed layer depth revealed an efficient PFe export out of the mixed layer at these sites. Export efficiencies (i.e. the ratio between export and uptake) exhibit a very efficient silica pump especially at the HNLC reference station where heavily silicified diatoms were present. On the contrary, the increase with depth of the C:N ratio and the low nitrogen export efficiencies support the idea of a strong remineralization and nitrification activity.

77 FR 39521 - Application for a License To Export Nuclear Reactor Major Components and Equipment

Federal Register 2010, 2011, 2012, 2013, 2014

2012-07-03

... LLC reactor coolant equipment for four constructing four plant May 14, 2012 pumps with motors, APR1400... Emirates. XR176 monitoring and plant in Braka. 110060011 control equipment, auxiliary equipment and... NUCLEAR REGULATORY COMMISSION Application for a License To Export Nuclear Reactor Major Components...

Stress Introduction Rate Alters the Benefit of AcrAB-TolC Efflux Pumps.

PubMed

Langevin, Ariel M; Dunlop, Mary J

2018-01-01

Stress tolerance studies are typically conducted in an all-or-none fashion. However, in realistic settings-such as in clinical or metabolic engineering applications-cells may encounter stresses at different rates. Therefore, how cells tolerate stress may depend on its rate of appearance. To address this, we studied how the rate of stress introduction affects bacterial stress tolerance by focusing on a key stress response mechanism. Efflux pumps, such as AcrAB-TolC of Escherichia coli , are membrane transporters well known for the ability to export a wide variety of substrates, including antibiotics, signaling molecules, and biofuels. Although efflux pumps improve stress tolerance, pump overexpression can result in a substantial fitness cost to the cells. We hypothesized that the ideal pump expression level would involve a rate-dependent trade-off between the benefit of pumps and the cost of their expression. To test this, we evaluated the benefit of the AcrAB-TolC pump under different rates of stress introduction, including a step, a fast ramp, and a gradual ramp. Using two chemically diverse stresses, the antibiotic chloramphenicol and the jet biofuel precursor pinene, we assessed the benefit provided by the pumps. A mathematical model describing these effects predicted the benefit as a function of the rate of stress introduction. Our findings demonstrate that as the rate of introduction is lowered, stress response mechanisms provide a disproportionate benefit to pump-containing strains, allowing cells to survive beyond the original inhibitory concentrations. IMPORTANCE Efflux pumps are ubiquitous in nature and provide stress tolerance in the cells of species ranging from bacteria to mammals. Understanding how pumps provide tolerance has far-reaching implications for diverse fields, from medicine to biotechnology. Here, we investigated how the rate of stressor appearance impacts tolerance. We focused on two distinct substrates of AcrAB-TolC efflux pumps, the antibiotic chloramphenicol and the biofuel precursor pinene. Interestingly, tolerance is highly dependent on the rate of stress introduction. Therefore, it is important to consider not only the total quantity of a stressor but also the rate at which it is applied. The implications of this work are significant because environments are rarely static; antibiotic concentrations change during dosing, and metabolic engineering processes change with time. Copyright © 2017 American Society for Microbiology.

Antibiotic Resistance Mediated by the MacB ABC Transporter Family: A Structural and Functional Perspective

PubMed Central

Greene, Nicholas P.; Kaplan, Elise; Crow, Allister; Koronakis, Vassilis

2018-01-01

The MacB ABC transporter forms a tripartite efflux pump with the MacA adaptor protein and TolC outer membrane exit duct to expel antibiotics and export virulence factors from Gram-negative bacteria. Here, we review recent structural and functional data on MacB and its homologs. MacB has a fold that is distinct from other structurally characterized ABC transporters and uses a unique molecular mechanism termed mechanotransmission. Unlike other bacterial ABC transporters, MacB does not transport substrates across the inner membrane in which it is based, but instead couples cytoplasmic ATP hydrolysis with transmembrane conformational changes that are used to perform work in the extra-cytoplasmic space. In the MacAB-TolC tripartite pump, mechanotransmission drives efflux of antibiotics and export of a protein toxin from the periplasmic space via the TolC exit duct. Homologous tripartite systems from pathogenic bacteria similarly export protein-like signaling molecules, virulence factors and siderophores. In addition, many MacB-like ABC transporters do not form tripartite pumps, but instead operate in diverse cellular processes including antibiotic sensing, cell division and lipoprotein trafficking. PMID:29892271

JPRS Report, Near East & South Asia

DTIC Science & Technology

1988-01-28

Israel’s peace in first instance. Having guar - anteed the Jewish state the separate peace it wanted with Egypt, the United States paid no attention to...plant gums , marine prod- ucts, wool and yarn have decreased. This decrease has been compensated by the great increase in the export of dried goods...mentioned session, the requests of the "Tuli Pers" company to export 3,000 tons of powdered detergent and the Pars pump company to export 1,500

Carbon fluxes in the Arabian Sea: Export versus recycling

NASA Astrophysics Data System (ADS)

Rixen, Tim; Gaye, Birgit; Ramaswamy, Venkitasubramani

2016-04-01

The organic carbon pump strongly influences the exchange of carbon between the ocean and the atmosphere. It is known that it responds to global change but the magnitude and the direction of change are still unpredictable. Sediment trap experiments carried out at various sites in the Arabian Sea between 1986 and 1998 have shown differences in the functioning of the organic carbon pump (OCP). An OCP driven by eukaryotic phytoplankton operated in the upwelling region off Oman and during the spring bloom in the northern Arabian Sea. Cyanobacteria capable of fixing nitrogen seem to dominate the phytoplankton community during all other seasons. The export driven by cyanobacteria was much lower than the export driven by eukaryotic phytoplankton. Productivity and nutrient availability seems to be a main factor controlling fluxes during blooms of eukaryotic phytoplankton. The ballast effect caused by inputs of dust into the ocean and its incorporation into sinking particles seems to be the main factor controlling the export during times when cyanobacteria dominate the phytoplankton community. C/N ratios of organic matter exported from blooms dominated by nitrogen fixing cyanobacteria are enhanced and, furthermore, indicate a more efficient recycling of nutrients at shallower water depth. This implies that the bacterial-driven OCP operates more in a recycling mode that keeps nutrients closer to the euphotic zone whereas the OCP driven by eukaryotic phytoplankton reduces the recycling of nutrients by exporting them into greater water-depth.

Short-chain ubiquitination is associated with the degradation rate of a cell-surface-resident bile salt export pump (BSEP/ABCB11).

PubMed

Hayashi, Hisamitsu; Sugiyama, Yuichi

2009-01-01

The reduced expression of the bile salt export pump (BSEP/ABCB11) at the canalicular membrane is associated with cholestasis-induced hepatotoxicity due to the accumulation of bile acids in hepatocytes. We demonstrated previously that 4-phenylbutyrate (4PBA) treatment, a U.S. Food and Drug Administration-approved drug for the treatment of urea cycle disorders, induces the cell-surface expression of BSEP by prolonging the degradation rate of cell-surface-resident BSEP. On the other hand, BSEP mutations, E297G and D482G, found in progressive familial intrahepatic cholestasis type 2 (PFIC2), reduced it by shortening the degradation rate of cell-surface-resident BSEP. Therefore, to help the development of the medical treatment of cholestasis, we investigated the underlying mechanism by which 4PBA and PFIC2-type mutations affect the BSEP degradation from cell surface, focusing on short-chain ubiquitination. In Madin-Darby canine kidney II (MDCK II) cells expressing BSEP and rat canalicular membrane vesicles, the molecular mass of the mature form of BSEP/Bsep shifted from 170 to 190 kDa after ubiquitin modification (molecular mass, 8 kDa). Ubiquitination susceptibility of BSEP/Bsep was reduced in vitro and in vivo by 4PBA treatment and, conversely, was enhanced by BSEP mutations E297G and D482G. Moreover, biotin-labeling studies using MDCK II cells demonstrated that the degradation of cell-surface-resident chimeric protein fusing ubiquitin to BSEP was faster than that of BSEP itself. In conclusion, BSEP/Bsep is modified with two to three ubiquitins, and its ubiquitination is modulated by 4PBA treatment and PFIC2-type mutations. Modulation of short-chain ubiquitination can regulate the change in the degradation rate of cell-surface-resident BSEP by 4PBA treatment and PFIC2-type mutations.

78 FR 42753 - Materials Technical Advisory Committee; Notice of Partially Closed Meeting

Federal Register 2010, 2011, 2012, 2013, 2014

2013-07-17

... BIS senior management and export control reform update including the proposed Routed Export....'' 4. Report of Composite Working Group and discussion on (1) feasibility of universal test for modulus... values for 1C008-10,1C210, and 1C990. 5. Report of Biological and Pump/Valves Working Group. 6. Report on...

Spatial variability of upper ocean POC export in the Bay of Bengal and the Indian Ocean determined using particle-reactive 234Th

NASA Astrophysics Data System (ADS)

Subha Anand, S.; Rengarajan, R.; Sarma, V. V. S. S.; Sudheer, A. K.; Bhushan, R.; Singh, S. K.

2017-05-01

The northern Indian Ocean is globally significant for its seasonally reversing winds, upwelled nutrients, high biological production, and expanding oxygen minimum zones. The region acts as sink and source for atmospheric CO2. However, the efficiency of the biological carbon pump to sequester atmospheric CO2 and export particulate organic carbon from the surface is not well known. To quantify the upper ocean carbon export flux and to estimate the efficiency of biological carbon pump in the Bay of Bengal and the Indian Ocean, seawater profiles of total 234Th were measured from surface to 300 m depth at 13 stations from 19.9°N to 25.3°S in a transect along 87°E, during spring intermonsoon period (March-April 2014). Results showed enhanced in situ primary production in the equatorial Indian Ocean and the central Bay of Bengal and varied from 13.2 to 173.8 mmol C m-2 d-1. POC export flux in this region varied from 0 to 7.7 mmol C m-2 d-1. Though high carbon export flux was found in the equatorial region, remineralization of organic carbon in the surface and subsurface waters considerably reduced organic carbon export in the Bay of Bengal. Annually recurring anticyclonic eddies enhanced organic carbon utilization and heterotrophy. Oxygen minimum zone developed due to stratification and poor ventilation was intensified by subsurface remineralization. 234Th-based carbon export fluxes were not comparable with empirical statistical model estimates based on primary production and temperature. Region-specific refinement of model parameters is required to accurately predict POC export fluxes.

Biosynthetic Approaches to Isotope Enrichment for Applications in Neutron Scattering and High Field NMR Spectroscopy: Methylotrophic

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

Mary E. lidstrom

Limitations in current isotopic labeling methods present a substantial bottleneck for the application of advanced structural techniques to many important biochemical problems. New tools are required to efficiently produce the necessary labeling patterns in biochemical precursors and incorporate them into protein molecules for structural studies. This project proposed involved one aspect of this problem, the development of expression vectors for a methylotrophic bacterium, Methylobacterium extorquens AM1. If high-level, efficient expression could be obtained in such a bacterium, it would be possible to use low-cost {sup 2}H- and/or {sup 13}C-labeled substrates such as methanol to label proteins. The Lidstrom laboratory atmore » the University of Washington worked closely with the collaborators at Los Alamos National Laboratories in the development and use of these vectors. (1) Overexpression of a target gene, bacterial dehalogenase--This enzyme was expressed in Methylobacterium extorquens AM1 using a high level methanol-inducible promoter, the mxaF promoter. High expression was achieved, but most was in an insoluble form. They expressed this protein in a mutant lacking polybetahydroxybutyrate granules, and high expression was achieved, up to 10% of the total soluble protein. The recombinant protein was purified and shown to be active, with characteristics similar to the enzyme produced in E. coli. (2) Development of regulated expression systems--A number of regulated promoters were tested in M. extorquens AM1, the most promising of which appeared to be the E. coli lac promoter coupled to the Laciq regulator. The repressor was shown to be active and a chromosomal insertion construct was generated that repressed the low-level lac promoter activity in M. extorquens AM1. However, IPTG induced this system only poorly. A number of studies were carried out leading to the conclusion that IPTG entered the cell but was exported by one or more export pumps. Target genes for such pumps were mutated but none of these showed increased induction. A number of methods were used to permeabilize the cell, and a 2-fold increase in induction was obtained with one of these. The activity of the lac promoter was increased by inserting a recently-identified M. extorquens AM1 enhancer element upstream. The promoter increased in activity 5-6 fold with this addition. In summary, they have developed a suite of expression tools and host mutant strains for expressing a variety of heterologous proteins in this methylotroph. These are now available for testing by the LANL collaborators in labeling reactors to obtain labeled proteins of interest.« less

Programmed cell death in the marine cyanobacterium Trichodesmium mediates carbon and nitrogen export

PubMed Central

Bar-Zeev, Edo; Avishay, Itamar; Bidle, Kay D; Berman-Frank, Ilana

2013-01-01

The extent of carbon (C) and nitrogen (N) export to the deep ocean depends upon the efficacy of the biological pump that transports primary production to depth, thereby preventing its recycling in the upper photic zone. The dinitrogen-fixing (diazotrophic) Trichodesmium spp. contributes significantly to oceanic C and N cycling by forming extensive blooms in nutrient-poor tropical and subtropical regions. These massive blooms generally collapse several days after forming, but the cellular mechanism responsible, along with the magnitude of associated C and N export processes, are as yet unknown. Here, we used a custom-made, 2-m high water column to simulate a natural bloom and to specifically test and quantify whether the programmed cell death (PCD) of Trichodesmium mechanistically regulates increased vertical flux of C and N. Our findings demonstrate that extremely rapid development and abrupt, PCD-induced demise (within 2–3 days) of Trichodesmium blooms lead to greatly elevated excretions of transparent exopolymers and a massive downward pulse of particulate organic matter. Our results mechanistically link autocatalytic PCD and bloom collapse to quantitative C and N export fluxes, suggesting that PCD may have an impact on the biological pump efficiency in the oceans. PMID:23887173

Value of pH regulators in the diagnosis, prognosis and treatment of cancer.

PubMed

Granja, Sara; Tavares-Valente, Diana; Queirós, Odília; Baltazar, Fátima

2017-04-01

Altered metabolism, associated with acidification of the extracellular milieu, is one of the major features of cancer. As pH regulation is crucial for the maintenance of all biological functions, cancer cells rely on the activity of lactate exporters and proton transporters to regulate their intracellular pH. The major players in cancer pH regulation are proton pump ATPases, sodium-proton exchangers (NHEs), monocarboxylate transporters (MCTs), carbonic anhydrases (CAs) and anion exchangers (AEs), which have been shown to be upregulated in several human malignancies. Thanks to the activity of the proton pumps and transporters, tumours acidify their microenvironment, becoming more aggressive and resistant to therapy. Thus, targeting tumour pH may contribute to more effective anticancer strategies for controlling tumour progression and therapeutic resistance. In the present study, we review the role of the main pH regulators expressed in human cancer cells, including their diagnostic and prognostic value, as well as their usefulness as therapeutic targets. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Herpesvirus Protein Selectively Inhibits Cellular mRNA Nuclear Export.

PubMed

Gong, Danyang; Kim, Yong Hoon; Xiao, Yuchen; Du, Yushen; Xie, Yafang; Lee, Kevin K; Feng, Jun; Farhat, Nisar; Zhao, Dawei; Shu, Sara; Dai, Xinghong; Chanda, Sumit K; Rana, Tariq M; Krogan, Nevan J; Sun, Ren; Wu, Ting-Ting

2016-11-09

Nuclear mRNA export is highly regulated to ensure accurate cellular gene expression. Viral inhibition of cellular mRNA export can enhance viral access to the cellular translation machinery and prevent anti-viral protein production but is generally thought to be nonselective. We report that ORF10 of Kaposi's sarcoma-associated herpesvirus (KSHV), a nuclear DNA virus, inhibits mRNA export in a transcript-selective manner to control cellular gene expression. Nuclear export inhibition by ORF10 requires an interaction with an RNA export factor, Rae1. Genome-wide analysis reveals a subset of cellular mRNAs whose nuclear export is blocked by ORF10 with the 3' UTRs of ORF10-targeted transcripts conferring sensitivity to export inhibition. The ORF10-Rae1 interaction is important for the virus to express viral genes and produce infectious virions. These results suggest that a nuclear DNA virus can selectively interfere with RNA export to restrict host gene expression for optimal replication. Published by Elsevier Inc.

Efflux pumps as antimicrobial resistance mechanisms.

PubMed

Poole, Keith

2007-01-01

Antibiotic resistance continues to hamper antimicrobial chemotherapy of infectious disease, and while biocide resistance outside of the laboratory is as yet unrealized, in vitro and in vivo episodes of reduced biocide susceptibility are not uncommon. Efflux mechanisms, both drug-specific and multidrug, are important determinants of intrinsic and/or acquired resistance to these antimicrobials in important human pathogens. Multidrug efflux mechanisms are generally chromosome-encoded, with their expression typically resultant from mutations in regulatory genes, while drug-specific efflux mechanisms are encoded by mobile genetic elements whose acquisition is sufficient for resistance. While it has been suggested that drug-specific efflux systems originated from efflux determinants of self-protection in antibiotic-producing Actinomycetes, chromosomal multidrug efflux determinants, at least in Gram-negative bacteria, are appreciated as having an intended housekeeping function unrelated to drug export and resistance. Thus, it will be important to elucidate the intended natural function of these efflux mechanisms in order, for example, to anticipate environmental conditions or circumstances that might promote their expression and, so, compromise antimicrobial chemotherapy. Given the clinical significance of antimicrobial exporters, it is clear that efflux must be considered in formulating strategies for treatment of drug-resistant infections, both in the development of new agents, for example, less impacted by efflux or in targeting efflux directly with efflux inhibitors.

Quantification of the lithogenic carbon pump following a dust deposition event

NASA Astrophysics Data System (ADS)

Bressac, M.; Guieu, C.; Doxaran, D.; Bourrin, F.; Desboeufs, K.; Leblond, N.; Ridame, C.

2013-08-01

Lithogenic particles, such as desert dust, have been postulated to influence particulate organic carbon (POC) export to the deep ocean by acting as mineral ballasts. However, an accurate understanding and quantification of the POC-dust association that occurs within the upper ocean is required in order to affine the "ballast hypothesis". In the framework of the DUNE project, two artificial seedings were performed seven days apart within large mesocosms. A suite of optical and biogeochemical measurements were used to quantify surface POC export following simulated dust events within a low-nutrient low-chlorophyll ecosystem. The two successive seedings led to a 2.3-6.7 fold higher POC flux as compared to the POC flux observed in controlled mesocosms. A simple linear regression analysis revealed that the lithogenic fluxes explained more than 85% of the variance in POC fluxes. At the scale of a dust deposition event, we estimated that 42-50% of POC fluxes were strictly associated with lithogenic particles through an aggregation process. Lithogenic ballasting also likely impacted the remaining POC fraction which resulted from the fertilization effect. The observations support the "ballast hypothesis" and provide a quantitative estimation of the surface POC export abiotically triggered by dust deposition. In this work, we demonstrate that the strength of such a "lithogenic carbon pump" depends on the biogeochemical conditions of the water column at the time of deposition. Based on these observations, we suggest that this "lithogenic carbon pump" could represent a major component of the biological pump in oceanic areas subjected to intense atmospheric forcing.

Selective nuclear export of specific classes of mRNA from mammalian nuclei is promoted by GANP

PubMed Central

Wickramasinghe, Vihandha O.; Andrews, Robert; Ellis, Peter; Langford, Cordelia; Gurdon, John B.; Stewart, Murray; Venkitaraman, Ashok R.; Laskey, Ronald A.

2014-01-01

The nuclear phase of the gene expression pathway culminates in the export of mature messenger RNAs (mRNAs) to the cytoplasm through nuclear pore complexes. GANP (germinal- centre associated nuclear protein) promotes the transfer of mRNAs bound to the transport factor NXF1 to nuclear pore complexes. Here, we demonstrate that GANP, subunit of the TRanscription-EXport-2 (TREX-2) mRNA export complex, promotes selective nuclear export of a specific subset of mRNAs whose transport depends on NXF1. Genome-wide gene expression profiling showed that half of the transcripts whose nuclear export was impaired following NXF1 depletion also showed reduced export when GANP was depleted. GANP-dependent transcripts were highly expressed, yet short-lived, and were highly enriched in those encoding central components of the gene expression machinery such as RNA synthesis and processing factors. After injection into Xenopus oocyte nuclei, representative GANP-dependent transcripts showed faster nuclear export kinetics than representative transcripts that were not influenced by GANP depletion. We propose that GANP promotes the nuclear export of specific classes of mRNAs that may facilitate rapid changes in gene expression. PMID:24510098

Regulation of the Expression of Bacterial Multidrug Exporters by Two-Component Signal Transduction Systems.

PubMed

Nishino, Kunihiko

2018-01-01

Bacterial multidrug exporters confer resistance to a wide range of antibiotics, dyes, and biocides. Recent studies have shown that there are many multidrug exporters encoded in bacterial genome. For example, it was experimentally identified that E. coli has at least 20 multidrug exporters. Because many of these multidrug exporters have overlapping substrate spectra, it is intriguing that bacteria, with their economically organized genomes, harbor such large sets of multidrug exporter genes. The key to understanding how bacteria utilize these multiple exporters lies in the regulation of exporter expression. Bacteria have developed signaling systems for eliciting a variety of adaptive responses to their environments. These adaptive responses are often mediated by two-component regulatory systems. In this chapter, the method to identify response regulators that affect expression of multidrug exporters is described.

A feedback regulatory model for RifQ-mediated repression of rifamycin export in Amycolatopsis mediterranei.

PubMed

Lei, Chao; Wang, Jingzhi; Liu, Yuanyuan; Liu, Xinqiang; Zhao, Guoping; Wang, Jin

2018-01-29

Due to the important role of rifamycin in curing tuberculosis infection, the study on rifamycin has never been stopped. Although RifZ, which locates within the rifamycin biosynthetic cluster, has recently been characterized as a pathway-specific regulator for rifamycin biosynthesis, little is known about the regulation of rifamycin export. In this work, we proved that the expression of the rifamycin efflux pump (RifP) was regulated by RifQ, a TetR-family transcriptional regulator. Deletion of rifQ had little impact on bacterial growth, but resulted in improved rifamycin production, which was consistent with the reverse transcription PCR results that RifQ negatively regulated rifP's transcription. With electrophoretic mobility shift assay and DNase I Footprinting assay, RifQ was found to directly bind to the promoter region of rifP, and a typical inverted repeat was identified within the RifQ-protected sequences. The transcription initiation site of rifP was further characterized and found to be upstream of the RifQ binding sites, well explaining the RifQ-mediated repression of rifP's transcription in vivo. Moreover, rifamycin B (the end product of rifamycin biosynthesis) remarkably decreased the DNA binding affinity of RifQ, which led to derepression of rifamycin export, reducing the intracellular concentration of rifamycin B as well as its toxicity against the host. Here, we proved that the export of rifamycin B was repressed by RifQ in Amycolatopsis mediterranei, and the RifQ-mediated repression could be specifically relieved by rifamycin B, the end product of rifamycin biosynthesis, based on which a feedback model was proposed for regulation of rifamycin export. With the findings here, one could improve the antibiotic yield by simply inactivating the negative regulator of the antibiotic transporter.

All0809/8/7 is a DevBCA-like ABC-type efflux pump required for diazotrophic growth in Anabaena sp. PCC 7120.

PubMed

Staron, Peter; Maldener, Iris

2012-10-01

Efflux pumps export a wide variety of proteinaceous and non-proteinaceous substrates across the Gram-negative cell wall. For the filamentous cyanobacterium Anabaena sp. strain PCC 7120, the ATP-driven glycolipid efflux pump DevBCA-TolC has been shown to be crucial for the differentiation of N(2)-fixing heterocysts from photosynthetically active vegetative cells. In this study, a homologous system was described. All0809/8/7-TolC form a typical ATP-driven efflux pump as shown by surface plasmon resonance. This putative exporter is also involved in diazotrophic growth of Anabaena sp. PCC 7120. A mutant in all0809 encoding the periplasmic membrane fusion protein of the pump was not able to grow without combined nitrogen. Although heterocysts of this mutant were not distinguishable from those of the wild-type in light and electron micrographs, they were impaired in providing the microoxic environment necessary for N(2) fixation. RT-PCR of all0809 transcripts and localization studies on All0807-GFP revealed that All0809/8/7 was initially downregulated during heterocyst maturation and upregulated at later stages of heterocyst formation in all cells of the filament. A substrate of the efflux pump could not be identified in ATP hydrolysis assays. We discuss a role for All0809/8/7-TolC in maintaining the continuous periplasm and how this would be of special importance for heterocyst differentiation.

Influence of diatom diversity on the ocean biological carbon pump

NASA Astrophysics Data System (ADS)

Tréguer, Paul; Bowler, Chris; Moriceau, Brivaela; Dutkiewicz, Stephanie; Gehlen, Marion; Aumont, Olivier; Bittner, Lucie; Dugdale, Richard; Finkel, Zoe; Iudicone, Daniele; Jahn, Oliver; Guidi, Lionel; Lasbleiz, Marine; Leblanc, Karine; Levy, Marina; Pondaven, Philippe

2018-01-01

Diatoms sustain the marine food web and contribute to the export of carbon from the surface ocean to depth. They account for about 40% of marine primary productivity and particulate carbon exported to depth as part of the biological pump. Diatoms have long been known to be abundant in turbulent, nutrient-rich waters, but observations and simulations indicate that they are dominant also in meso- and submesoscale structures such as fronts and filaments, and in the deep chlorophyll maximum. Diatoms vary widely in size, morphology and elemental composition, all of which control the quality, quantity and sinking speed of biogenic matter to depth. In particular, their silica shells provide ballast to marine snow and faecal pellets, and can help transport carbon to both the mesopelagic layer and deep ocean. Herein we show that the extent to which diatoms contribute to the export of carbon varies by diatom type, with carbon transfer modulated by the Si/C ratio of diatom cells, the thickness of the shells and their life strategies; for instance, the tendency to form aggregates or resting spores. Model simulations project a decline in the contribution of diatoms to primary production everywhere outside of the Southern Ocean. We argue that we need to understand changes in diatom diversity, life cycle and plankton interactions in a warmer and more acidic ocean in much more detail to fully assess any changes in their contribution to the biological pump.

Inhibition of pyrimidine synthesis reverses viral virulence factor-mediated block of mRNA nuclear export

PubMed Central

Zhang, Liang; Das, Priyabrata; Schmolke, Mirco; Manicassamy, Balaji; Wang, Yaming; Deng, Xiaoyi; Cai, Ling; Tu, Benjamin P.; Forst, Christian V.; Roth, Michael G.; Levy, David E.; García-Sastre, Adolfo; de Brabander, Jef; Phillips, Margaret A.

2012-01-01

The NS1 protein of influenza virus is a major virulence factor essential for virus replication, as it redirects the host cell to promote viral protein expression. NS1 inhibits cellular messenger ribonucleic acid (mRNA) processing and export, down-regulating host gene expression and enhancing viral gene expression. We report in this paper the identification of a nontoxic quinoline carboxylic acid that reverts the inhibition of mRNA nuclear export by NS1, in the absence or presence of the virus. This quinoline carboxylic acid directly inhibited dihydroorotate dehydrogenase (DHODH), a host enzyme required for de novo pyrimidine biosynthesis, and partially reduced pyrimidine levels. This effect induced NXF1 expression, which promoted mRNA nuclear export in the presence of NS1. The release of NS1-mediated mRNA export block by DHODH inhibition also occurred in the presence of vesicular stomatitis virus M (matrix) protein, another viral inhibitor of mRNA export. This reversal of mRNA export block allowed expression of antiviral factors. Thus, pyrimidines play a necessary role in the inhibition of mRNA nuclear export by virulence factors. PMID:22312003

Quantification of the lithogenic carbon pump following a simulated dust-deposition event in large mesocosms

NASA Astrophysics Data System (ADS)

Bressac, M.; Guieu, C.; Doxaran, D.; Bourrin, F.; Desboeufs, K.; Leblond, N.; Ridame, C.

2014-02-01

Lithogenic particles, such as desert dust, have been postulated to influence particulate organic carbon (POC) export to the deep ocean by acting as mineral ballasts. However, an accurate understanding and quantification of the POC-dust association that occurs within the upper ocean is required in order to refine the "ballast hypothesis". In the framework of the DUNE (a DUst experiment in a low-Nutrient, low-chlorophyll Ecosystem) project, two artificial seedings were performed seven days apart within large mesocosms. A suite of optical and biogeochemical measurements were used to quantify surface POC export following simulated dust events within a low-nutrient, low-chlorophyll ecosystem. The two successive seedings led to a 2.3-6.7-fold higher POC flux than the POC flux observed in controlled mesocosms. A simple linear regression analysis revealed that the lithogenic fluxes explained more than 85% of the variance in POC fluxes. On the scale of a dust-deposition event, we estimated that 42-50% of POC fluxes were strictly associated with lithogenic particles (through aggregation and most probably sorption processes). Lithogenic ballasting also likely impacted the remaining POC fraction which resulted from the fertilization effect. The observations support the "ballast hypothesis" and provide a quantitative estimation of the surface POC export abiotically triggered by dust deposition. In this work, we demonstrate that the strength of such a "lithogenic carbon pump" depends on the biogeochemical conditions of the water column at the time of deposition. Based on these observations, we suggest that this lithogenic carbon pump could represent a major component of the biological pump in oceanic areas subjected to intense atmospheric forcing.

Global assessment of ocean carbon export by combining satellite observations and food-web models

NASA Astrophysics Data System (ADS)

Siegel, D. A.; Buesseler, K. O.; Doney, S. C.; Sailley, S. F.; Behrenfeld, M. J.; Boyd, P. W.

2014-03-01

The export of organic carbon from the surface ocean by sinking particles is an important, yet highly uncertain, component of the global carbon cycle. Here we introduce a mechanistic assessment of the global ocean carbon export using satellite observations, including determinations of net primary production and the slope of the particle size spectrum, to drive a food-web model that estimates the production of sinking zooplankton feces and algal aggregates comprising the sinking particle flux at the base of the euphotic zone. The synthesis of observations and models reveals fundamentally different and ecologically consistent regional-scale patterns in export and export efficiency not found in previous global carbon export assessments. The model reproduces regional-scale particle export field observations and predicts a climatological mean global carbon export from the euphotic zone of 6 Pg C yr-1. Global export estimates show small variation (typically < 10%) to factor of 2 changes in model parameter values. The model is also robust to the choices of the satellite data products used and enables interannual changes to be quantified. The present synthesis of observations and models provides a path for quantifying the ocean's biological pump.

Expression of multidrug resistance efflux pump gene norA is iron responsive in Staphylococcus aureus.

PubMed

Deng, Xin; Sun, Fei; Ji, Quanjiang; Liang, Haihua; Missiakas, Dominique; Lan, Lefu; He, Chuan

2012-04-01

Staphylococcus aureus utilizes efflux transporter NorA to pump out a wide range of structurally dissimilar drugs, conferring low-level multidrug resistance. The regulation of norA expression has yet to be fully understood although past studies have revealed that this gene is under the control of the global transcriptional regulator MgrA and the two-component system ArlRS. To identify additional regulators of norA, we screened a transposon library in strain Newman expressing the transcriptional fusion norA-lacZ for altered β-galactosidase activity. We identify a transposon insertion in fhuB, a gene that encodes a ferric hydroxamate uptake system permease, and propose that the norA transcription is iron responsive. In agreement with this observation, addition of FeCl(3) repressed the induction of norA-lacZ, suggesting that bacterial iron uptake plays an important role in regulating norA transcription. In addition, a fur (ferric uptake regulator) deletion exhibited compromised norA transcription and reduced resistance to quinolone compared to the wild-type strain, indicating that fur functions as a positive regulator of norA. A putative Fur box identified in the promoter region of norA was confirmed by electrophoretic mobility shift and DNase I footprint assays. Finally, by employing a siderophore secretion assay, we reveal that NorA may contribute to the export of siderophores. Collectively, our experiments uncover some novel interactions between cellular iron level and norA regulation in S. aureus.

Expression of Multidrug Resistance Efflux Pump Gene norA Is Iron Responsive in Staphylococcus aureus

PubMed Central

Deng, Xin; Sun, Fei; Ji, Quanjiang; Liang, Haihua; Missiakas, Dominique; Lan, Lefu

2012-01-01

Staphylococcus aureus utilizes efflux transporter NorA to pump out a wide range of structurally dissimilar drugs, conferring low-level multidrug resistance. The regulation of norA expression has yet to be fully understood although past studies have revealed that this gene is under the control of the global transcriptional regulator MgrA and the two-component system ArlRS. To identify additional regulators of norA, we screened a transposon library in strain Newman expressing the transcriptional fusion norA-lacZ for altered β-galactosidase activity. We identify a transposon insertion in fhuB, a gene that encodes a ferric hydroxamate uptake system permease, and propose that the norA transcription is iron responsive. In agreement with this observation, addition of FeCl3 repressed the induction of norA-lacZ, suggesting that bacterial iron uptake plays an important role in regulating norA transcription. In addition, a fur (ferric uptake regulator) deletion exhibited compromised norA transcription and reduced resistance to quinolone compared to the wild-type strain, indicating that fur functions as a positive regulator of norA. A putative Fur box identified in the promoter region of norA was confirmed by electrophoretic mobility shift and DNase I footprint assays. Finally, by employing a siderophore secretion assay, we reveal that NorA may contribute to the export of siderophores. Collectively, our experiments uncover some novel interactions between cellular iron level and norA regulation in S. aureus. PMID:22267518

Improved expression of halorhodopsin for light-induced silencing of neuronal activity

PubMed Central

Zhao, Shengli; Cunha, Catarina; Zhang, Feng; Liu, Qun; Gloss, Bernd; Deisseroth, Karl; Augustine, George J.; Feng, Guoping

2011-01-01

The ability to control and manipulate neuronal activity within an intact mammalian brain is of key importance for mapping functional connectivity and for dissecting the neural circuitry underlying behaviors. We have previously generated transgenic mice that express channelrhodopsin-2 for light-induced activation of neurons and mapping of neural circuits. Here we describe transgenic mice that express halorhodopsin (NpHR), a light-driven chloride pump, that can be used to silence neuronal activity via light. Using the Thy-1 promoter to target NpHR expression to neurons, we found that neurons in these mice expressed high levels of NpHR-YFP and that illumination of cortical pyramidal neurons expressing NpHR-YFP led to rapid, reversible photoinhibition of action potential firing in these cells. However, NpHR-YFP expression led to the formation of numerous intracellular blebs, which may disrupt neuronal function. Labeling of various subcellular markers indicated that the blebs arise from retention of NpHR-YFP in the endoplasmic reticulum. By improving the signal peptide sequence and adding an ER export signal to NpHR-YFP, we eliminated the formation of blebs and dramatically increased the membrane expression of NpHR-YFP. Thus, the improved version of NpHR should serve as an excellent tool for neuronal silencing in vitro and in vivo. PMID:18931914

Regulation of hepatic bile acid transporters Ntcp and Bsep expression.

PubMed

Cheng, Xingguo; Buckley, David; Klaassen, Curtis D

2007-12-03

Sodium-taurocholate cotransporting polypeptide (Ntcp) and bile salt export pump (Bsep) are two key transporters for hepatic bile acid uptake and excretion. Alterations in Ntcp and Bsep expression have been reported in pathophysiological conditions. In the present study, the effects of age, gender, and various chemicals on the regulation of these two transporters were characterized in mice. Ntcp and Bsep mRNA levels in mouse liver were low in the fetus, but increased to its highest expression at parturition. After birth, mouse Ntcp and Bsep mRNA decreased by more than 50%, and then gradually increased to adult levels by day 30. Expression of mouse Ntcp mRNA and protein exhibit higher levels in female than male livers. No gender difference exists in BSEP/Bsep expression in human and mouse livers. Hormone replacements conducted in gonadectomized, hypophysectomized, and lit/lit mice indicate that female-predominant Ntcp expression in mouse liver is due to the inhibitory effect of male-pattern GH secretion, but not sex hormones. Ntcp and Bsep expression are in general resistant to induction by a large battery of microsomal enzyme inducers. Administration of cholestyramine increased Ntcp, whereas chenodeoxycholic acid (CDCA) increased Bsep mRNA expression. In conclusion, mouse Ntcp and Bsep are regulated by age, gender, cholestyramine, and bile acid, but resistant to induction by most microsomal enzyme inducers.

Measured flow and tracer-dye data for spring 1996 and 1997 for the south Sacramento-San Joaquin Delta, California

USGS Publications Warehouse

Oltmann, Richard N.

1999-01-01

During the spring of years when the flow of the San Joaquin River is less than 7,000 cubic feet per second (ft3/s) a temporary rock barrier is installed by the California Department of Water Resources (DWR) at the head of Old River (HOR) in the south Sacramento-San Joaquin Delta to prevent out migrating salmon in the San Joaquin River from entering Old River and being drawn to the State and federal pumping facilities (Figure 1). The export rate of the pumping facilities also is reduced during these migration periods to minimize the draw of fish to the export facilities through the other channels connected to the San Joaquin River north of the HOR such as Turner Cut, Columbia Cut, and Middle River.

The impact on atmospheric CO2 of iron fertilization induced changes in the ocean's biological pump

NASA Astrophysics Data System (ADS)

Jin, X.; Gruber, N.; Frenzel, H.; Doney, S. C.; McWilliams, J. C.

2008-03-01

Using numerical simulations, we quantify the impact of changes in the ocean's biological pump on the air-sea balance of CO2 by fertilizing a small surface patch in the high-nutrient, low-chlorophyll region of the eastern tropical Pacific with iron. Decade-long fertilization experiments are conducted in a basin-scale, eddy-permitting coupled physical/biogeochemical/ecological model. In contrast to previous studies, we find that most of the dissolved inorganic carbon (DIC) removed from the euphotic zone by the enhanced biological export is replaced by uptake of CO2 from the atmosphere. Atmospheric uptake efficiencies, the ratio of the perturbation in air-sea CO2 flux to the perturbation in export flux across 100 m, integrated over 10 years, are 0.75 to 0.93 in our patch size-scale experiments. The atmospheric uptake efficiency is insensitive to the duration of the experiment. The primary factor controlling the atmospheric uptake efficiency is the vertical distribution of the enhanced biological production and export. Iron fertilization at the surface tends to induce production anomalies primarily near the surface, leading to high efficiencies. In contrast, mechanisms that induce deep production anomalies (e.g. altered light availability) tend to have a low uptake efficiency, since most of the removed DIC is replaced by lateral and vertical transport and mixing. Despite high atmospheric uptake efficiencies, patch-scale iron fertilization of the ocean's biological pump tends to remove little CO2 from the atmosphere over the decadal timescale considered here.

Influence of a Decaying Cyclonic Eddy on Biogenic Silica and Particulate Organic Carbon in the Tropical South China Sea Based on 234Th-238U Disequilibrium.

PubMed

Yang, Weifeng; Chen, Min; Zheng, Minfang; He, Zhigang; Zhang, Xinxing; Qiu, Yusheng; Xu, Wangbin; Ma, Lili; Lin, Zhiyu; Hu, Wangjiang; Zeng, Jian

2015-01-01

Eddies play a critical role in regulating the biological pump by pumping new nutrients to the euphotic zone. However, the effects of cyclonic eddies on particle export are not well understood. Here, biogenic silica (BSi) and particulate organic carbon (POC) exports were examined inside and outside a decaying cyclonic eddy using 234Th-238U disequilibria in the tropical South China Sea. For the eddy and outside stations, the average concentrations of BSi in the euphotic zone were 0.17±0.09 μmol L-1 (mean±sd, n = 20) and 0.21±0.06 μmol L-1 (n = 34). The POC concentrations were 1.42±0.56 μmol L-1 (n = 34) and 1.30±0.46 μmol L-1 (n = 51). Both BSi and POC abundances did not show change at the 95% confidence level. Based on the 234Th-238U model, BSi export fluxes in the eddy averaged 0.18±0.15 mmol Si m-2 d-1, which was comparable with the 0.40±0.20 mmol Si m-2 d-1 outside the eddy. Similarly, the average POC export fluxes were 1.5±1.4 mmol C m-2 d-1 and 1.9±1.3 mmol C m-2 d-1 for the eddy and outside stations. From these results we concluded that cyclonic eddies in their decaying phase have little effect on the abundance and export of biogenic particles.

Influence of a Decaying Cyclonic Eddy on Biogenic Silica and Particulate Organic Carbon in the Tropical South China Sea Based on 234Th-238U Disequilibrium

PubMed Central

Yang, Weifeng; Chen, Min; Zheng, Minfang; He, Zhigang; Zhang, Xinxing; Qiu, Yusheng; Xu, Wangbin; Ma, Lili; Lin, Zhiyu; Hu, Wangjiang; Zeng, Jian

2015-01-01

Eddies play a critical role in regulating the biological pump by pumping new nutrients to the euphotic zone. However, the effects of cyclonic eddies on particle export are not well understood. Here, biogenic silica (BSi) and particulate organic carbon (POC) exports were examined inside and outside a decaying cyclonic eddy using 234Th-238U disequilibria in the tropical South China Sea. For the eddy and outside stations, the average concentrations of BSi in the euphotic zone were 0.17±0.09 μmol L-1 (mean±sd, n = 20) and 0.21±0.06 μmol L-1 (n = 34). The POC concentrations were 1.42±0.56 μmol L-1 (n = 34) and 1.30±0.46 μmol L-1 (n = 51). Both BSi and POC abundances did not show change at the 95% confidence level. Based on the 234Th-238U model, BSi export fluxes in the eddy averaged 0.18±0.15 mmol Si m-2 d-1, which was comparable with the 0.40±0.20 mmol Si m-2 d-1 outside the eddy. Similarly, the average POC export fluxes were 1.5±1.4 mmol C m-2 d-1 and 1.9±1.3 mmol C m-2 d-1 for the eddy and outside stations. From these results we concluded that cyclonic eddies in their decaying phase have little effect on the abundance and export of biogenic particles. PMID:26317555

Aiding and Abetting Cancer: mRNA export and the nuclear pore

PubMed Central

Culjkovic-Kraljacic, Biljana; Borden, Katherine L.B

2013-01-01

mRNA export is a critical step in gene expression. Export of transcripts can be modulated in response to cellular signaling or stress. Consistently, mRNA export is dysregulated in primary human specimens derived from many different forms of cancer. Aberrant expression of export factors can alter export of specific transcripts encoding proteins involved in proliferation, survival and oncogenesis. These specific factors, which are not used for bulk mRNA export, are obvious therapeutic targets. Indeed, given the emerging role of mRNA export in cancer, it is not surprising that efforts to target different aspects of this pathway have reached the clinical trial stage. Thus, like transcription and translation, mRNA export may also play a critical role in cancer genesis and maintenance. PMID:23582887

27 CFR 28.40 - Evidence of exportation: distilled spirits and wine.

Code of Federal Regulations, 2013 CFR

2013-04-01

... exportation of any shipment of distilled spirits or wine may be evidenced by: (a) A copy of the export bill of lading (§ 28.250); or (b) A copy of the railway express receipt (§ 28.251); or (c) A copy of the air express receipt (§ 28.252); or (d) A copy of the through bill of lading where exportation is to a...

27 CFR 28.40 - Evidence of exportation: distilled spirits and wine.

Code of Federal Regulations, 2014 CFR

2014-04-01

... exportation of any shipment of distilled spirits or wine may be evidenced by: (a) A copy of the export bill of lading (§ 28.250); or (b) A copy of the railway express receipt (§ 28.251); or (c) A copy of the air express receipt (§ 28.252); or (d) A copy of the through bill of lading where exportation is to a...

27 CFR 28.40 - Evidence of exportation: distilled spirits and wine.

Code of Federal Regulations, 2011 CFR

2011-04-01

... exportation of any shipment of distilled spirits or wine may be evidenced by: (a) A copy of the export bill of lading (§ 28.250); or (b) A copy of the railway express receipt (§ 28.251); or (c) A copy of the air express receipt (§ 28.252); or (d) A copy of the through bill of lading where exportation is to a...

27 CFR 28.40 - Evidence of exportation: distilled spirits and wine.

Code of Federal Regulations, 2012 CFR

2012-04-01

... exportation of any shipment of distilled spirits or wine may be evidenced by: (a) A copy of the export bill of lading (§ 28.250); or (b) A copy of the railway express receipt (§ 28.251); or (c) A copy of the air express receipt (§ 28.252); or (d) A copy of the through bill of lading where exportation is to a...

Ocean Biological Pump Sensitivities and Implications for Climate Change Impacts

NASA Technical Reports Server (NTRS)

Romanou, Anastasia

2013-01-01

The ocean is one of the principal reservoirs of CO2, a greenhouse gas, and therefore plays a crucial role in regulating Earth's climate. Currently, the ocean sequesters about a third of anthropogenic CO2 emissions, mitigating the human impact on climate. At the same time, the deeper ocean represents the largest carbon pool in the Earth System and processes that describe the transfer of carbon from the surface of the ocean to depth are intimately linked to the effectiveness of carbon sequestration.The ocean biological pump (OBP), which involves several biogeochemical processes, is a major pathway for transfer of carbon from the surface mixed layer into the ocean interior. About 75 of the carbon vertical gradient is due to the carbon pump with only 25 attributed to the solubility pump. However, the relative importance and role of the two pumps is poorly constrained. OBP is further divided to the organic carbon pump (soft tissue pump) and the carbonate pump, with the former exporting about 10 times more carbon than the latter through processes like remineralization.Major uncertainties about OBP, and hence in the carbon uptake and sequestration, stem from uncertainties in processes involved in OBP such as particulate organicinorganic carbon sinkingsettling, remineralization, microbial degradation of DOC and uptakegrowth rate changes of the ocean biology. The deep ocean is a major sink of atmospheric CO2 in scales of hundreds to thousands of years, but how the export efficiency (i.e. the fraction of total carbon fixation at the surface that is transported at depth) is affected by climate change remains largely undetermined. These processes affect the ocean chemistry (alkalinity, pH, DIC, particulate and dissolved organic carbon) as well as the ecology (biodiversity, functional groups and their interactions) in the ocean. It is important to have a rigorous, quantitative understanding of the uncertainties involved in the observational measurements, the models and the projections of future changes.

Role of eddy pumping in enhancing primary production in the ocean

NASA Technical Reports Server (NTRS)

Falkowski, Paul G.; Kolber, Zbigniew; Ziemann, David; Bienfang, Paul K.

1991-01-01

Eddy pumping is considered to explain the disparity between geochemical estimates and biological measurements of exported production. Episodic nutrient injections from the ocean into the photic zone can be generated by eddy pumping, which biological measurements cannot sample accurately. The enhancement of production is studied with respect to a cyclonic eddy in the subtropical Pacific. A pump-and-probe fluorimeter generates continuous vertical profiles of primary productivity from which the contributions of photochemical and nonphotochemical processes to fluorescence are derived. A significant correlation is observed between the fluorescence measurements and radiocarbon measurements. The results indicate that eddy pumping has an important effect on phytoplankton production and that this production is near the maximum relative specific growth rates. Based on the production enhancement observed in this case, eddy pumping increases total primary production by only 20 percent and does not account for all enhancement.

Section 7 reactor incident file general information from 1945

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

Not Available

1969-01-10

At 0308 on January 10, 1966, both B and C Reactors ``scrammed`` due to an electrical fault on Line C2-L8 caused by a raccoon coming in contact with the 13-8 KV line on top of transformer No. 2 at 182-B Building. Line C2-L8 relayed out at the 151-B Building. Details of the occurrence at 151-B are covered in the attachment. C-Reactor scrammed due to reduced voltage on the pressure monitor system. The reduction in voltage caused the auxiliary relays of the pressure monitor ground detector to open, de-energizing the end result relays PSR and PSRA. The safety circuit trip identificationmore » system displayed ``Pressure Monitor`` and ``Ground Detector.`` B-Reactor scrammed by a power failure signal from 190-B Building. The power failure relays for pump numbers 1 and 3 opened due to these pumps contributing power to the fault. The power failure relays at 190-B remained open long enough for the end result relays PF and PFA to open. Since these relays are timed delayed, 0.26 seconds, the power failure relays must have remained open at least that long. At the 190-B Building the steam turbines started due to the power failure relays for pump numbers 1 and 3 opening. The main process pumps remained stable and continued to supply normal flow to the reactor. Pumps were tripped from the line at 182-B and 183-B Buildings. The surge suppressors cycled normally and the turbine export pumps started as a result of low export line pressure. No power equipment was affected in C Area.« less

78 FR 65265 - Materials Technical Advisory Committee; Notice of Open Meeting

Federal Register 2010, 2011, 2012, 2013, 2014

2013-10-31

... BIS senior management and export control reform update. 3. Presentation from DuPont on the challenges... friendly. 4. Report of Composite Working Group. 5. Report of Biological and Pump/Valves Working Group. 6...

Bipolar and Related Disorders Induced by Sodium 4-Phenylbutyrate in a Male Adolescent with Bile Salt Export Pump Deficiency Disease.

PubMed

Vitale, Giovanni; Simonetti, Giulia; Pirillo, Martina; Taruschio, Gianfranco; Andreone, Pietro

2016-09-01

Bile Salt Export Pump (BSEP) Deficiency disease, including Progressive Familial Intrahepatic Cholestasis type 2 (PFIC2), is a rare disease, usually leading within the first ten years to portal hypertension, liver failure, hepatocellular carcinoma. Often liver transplantation is needed. Sodium 4-phenylbutyrate (4-PB) seems to be a potential therapeutic compound for PFIC2. Psychiatric side effects in the adolescent population are little known and little studied since the drug used to treat children and infants. So we described a case of Caucasian boy, suffering from a late onset PFIC2, listed for a liver transplant when he was sixteen and treated with 4-FB (200 mg per kilogram of body weight per day). The drug was discontinued for the onset of bipolar and related disorders. This case illustrates possible psychiatric side effects of the drug.

Preliminary study on Emodin alleviating alpha-naphthylisothiocyanate-induced intrahepatic cholestasis by regulation of liver farnesoid X receptor pathway.

PubMed

Ding, Yan; Xiong, Xiao-Li; Zhou, Li-Shan; Yan, Su-Qi; Qin, Huan; Li, Hua-Rong; Zhang, Ling-Ling; Chen, Peng; Yao, Cong; Jiang, Zhi-Xia; Zhao, Lei

2016-12-01

The aim of this study is to investigate Emodin on alleviating intrahepatic cholestasis by regulation of liver farnesoid X receptor (FXR) pathway. Cell and animal models of intrahepatic cholestatis were established. Biochemical tests and histomorphology were performed. The messenger RNA (mRNA) and protein expression of FXR, small heterodimer partner (SHP), uridine diphosphate glucuronosyltransferase 2 family polypeptide B4 (UGT2B4), and bile salt export pump (BSEP) was detected. As a result, compared with the model group, the serum levels of biochemical test were significantly lower in the Emodin group (P <0.01). The histopathological changes were remitted significantly by Emodin treatment. In the model group, the mRNA and protein expression of FXR, SHP, UGT2B4, and BSEP was significantly lower than in the normal group in cell models (P <0.05). With Emodin intervention, the expression of FXR, SHP, UGT2B4, and BSEP was notably increased (P <0.05). In conclusion, Emodin plays a protective role in intrahepatic cholestasis by promoting FXR signal pathways. © The Author(s) 2016.

Annual Cycles of Deep-ocean, Biogeochemical Export Fluxes and Biological Pump Processes in Subtropical and Subantarctic Waters, Southwest Pacific Ocean

NASA Astrophysics Data System (ADS)

Nodder, S.; Chiswell, S.; Northcote, L.

2016-02-01

One of the key aspects of the global carbon cycle is the efficiency and spatio-temporal variability of the biological pump. In this paper, the annual cycles of particle fluxes, derived from moored sediment trap data collected from 2000-12 in subtropical (STW) and subantarctic waters (SAW), east of New Zealand, are presented. These observations are the most comprehensive export flux time-series from temperate Southern Hemisphere latitudes to date. With high levels of variability, fluxes in SAW were markedly lower than in STW, reflecting the picophytoplankton-dominated communities in the iron-limited, high nutrient-low chlorophyll SAW. Austral spring chlorophyll blooms in surface STW were near-synchronous with elevated fluxes of bio-siliceous, carbonate and organic carbon-rich materials to the deep ocean, probably facilitated by diatom sedimentation. Lithogenic fluxes were also high in STW, compared to SAW, reflecting proximity to the New Zealand landmass. In contrast, the highest biogenic fluxes in SAW occurred in spring when surface chlorophyll concentrations were low, while highest annual chlorophyll concentrations were in summer with no associated flux increase. We hypothesize that the high spring export in SAW occurs from subsurface chlorophyll accumulations that are not evident from remote-sensing satellites. This material was also rich in biogenic silica, perhaps related to the preferential export of diatoms and other silica-producing organisms, such as silicoflagellates and radiolarians. Particle fluxes in STW are similar to that of other mesotrophic to oligotrophic waters ( 6-7 mgC m-2 d-1), whereas export from SAW is below global averages ( 3 mgC m-2 d-1), and is characterized by carbonate-dominated and prominent bio-siliceous deposition.

Planktic foraminifer and coccolith contribution to carbonate export fluxes over the central Kerguelen Plateau

NASA Astrophysics Data System (ADS)

Rembauville, M.; Meilland, J.; Ziveri, P.; Schiebel, R.; Blain, S.; Salter, I.

2016-05-01

We report the contribution of planktic foraminifers and coccoliths to the particulate inorganic carbon (PIC) export fluxes collected over an annual cycle (October 2011/September 2012) on the central Kerguelen Plateau in the Antarctic Zone (AAZ) south of the Polar Front (PF). The seasonality of PIC flux was decoupled from surface chlorophyll a concentration and particulate organic carbon (POC) fluxes and was characterized by a late summer (February) maximum. This peak was concomitant with the highest satellite-derived sea surface PIC and corresponded to a Emiliania huxleyi coccoliths export event that accounted for 85% of the annual PIC export. The foraminifer contribution to the annual PIC flux was much lower (15%) and dominated by Turborotalita quinqueloba and Neogloboquadrina pachyderma. Foraminifer export fluxes were closely related to the surface chlorophyll a concentration, suggesting food availability as an important factor regulating the foraminifer's biomass. We compared size-normalized test weight (SNW) of the foraminifers with previously published SNW from the Crozet Islands using the same methodology and found no significant difference in SNW between sites for a given species. However, the SNW was significantly species-specific with a threefold increase from T. quinqueloba to Globigerina bulloides. The annual PIC:POC molar ratio of 0.07 was close to the mean ratio for the global ocean and lead to a low carbonate counter pump effect (~5%) compared to a previous study north of the PF (6-32%). We suggest that lowers counter pump effect south of the PF despite similar productivity levels is due to a dominance of coccoliths in the PIC fluxes and a difference in the foraminifers species assemblage with a predominance of polar species with lower SNW.

Low hypoxia inducible factor-1α (HIF-1α) expression in testicular germ cell tumors - a major reason for enhanced chemosensitivity?

PubMed

Shenoy, Niraj; Dronca, Roxana; Quevedo, Fernando; Boorjian, Stephen A; Cheville, John; Costello, Brian; Kohli, Manish; Witzig, Thomas; Pagliaro, Lance

2017-08-01

The molecular basis for enhanced chemosensitivity of testicular germ cell tumors (GCT) has been an area of great interest, as it could potentially give us therapeutic leads in other resistant malignancies. Thus far, however, the increased sensitivity of GCT has been variously attributed to multiple factors - an inability to detoxify cisplatin, a lack of export pumps, an inability to repair the DNA damage, an intact apoptotic cascade and lack of p53 mutation; but a unifying underlying etiology leading to the aforementioned processes and having a translational implication has so far been elusive. Herein, we offer evidence to support a potential significant role for the previously demonstrated low hypoxia inducible factor-1α (HIF-1α) expression in mediating the general exquisite chemosensitivity of testicular GCT, through the aforementioned processes. This molecular mechanism based hypothesis could have a significant translational implication in platinum refractory GCT as well as other platinum resistant malignancies.

The Novel Helicobacter pylori CznABC Metal Efflux Pump Is Required for Cadmium, Zinc, and Nickel Resistance, Urease Modulation, and Gastric Colonization

PubMed Central

Stähler, Frank Nils; Odenbreit, Stefan; Haas, Rainer; Wilrich, Julia; Vliet, Arnoud H. M. Van; Kusters, Johannes G.; Kist, Manfred; Bereswill, Stefan

2006-01-01

Maintaining metal homeostasis is crucial for the adaptation of Helicobacter pylori to the gastric environment. Iron, copper, and nickel homeostasis has recently been demonstrated to be required for the establishment of H. pylori infection in animal models. Here we demonstrate that the HP0969-0971 gene cluster encoding the Czc-type metal export pump homologs HP0969, HP0970, and the H. pylori-specific protein HP0971 forms part of a novel H. pylori metal resistance determinant, which is required for gastric colonization and for the modulation of urease activity. Insertional mutagenesis of the HP0971, HP0970, or HP0969 genes in H. pylori reference strain 26695 resulted in increased sensitivity to cadmium, zinc, and nickel (czn), suggesting that the encoded proteins constitute a metal-specific export pump. Accordingly, the genes were designated cznC (HP0971), cznB (HP0970), and cznA (HP0969). The CznC and CznA proteins play a predominant role in nickel homeostasis, since only the cznC and cznA mutants but not the cznB mutant displayed an 8- to 10-fold increase in urease activity. Nickel-specific affinity chromatography demonstrated that recombinant versions of CznC and CznB can bind to nickel and that the purified CznB protein interacted with cadmium and zinc, since both metals competitively inhibited nickel binding. Finally, single cznA, cznB, and cznC mutants did not colonize the stomach in a Mongolian gerbil-based animal model. This demonstrates that the metal export functions of H. pylori cznABC are essential for gastric colonization and underlines the extraordinary importance of metal ion homeostasis for the survival of H. pylori in the gastric environment. PMID:16790756

Replumbing of the Biological Pump caused by Millennial Climate Variability

NASA Astrophysics Data System (ADS)

Galbraith, E.; Sarmiento, J.

2008-12-01

It has been hypothesized that millennial-timescale variability in the biological pump was a critical instigator of glacial-interglacial cycles. However, even in the absence of changes in ecosystem function (e.g. due to iron fertilization), determining the mechanisms by which physical climate variability alters the biological pump is not simple. Changes in upper ocean circulation and deep water formation have previously been shown to alter both the downward flux of organic matter and the mass of respired carbon in the ocean interior, often in non- intuitive ways. For example, a reduced upward flux of nutrients at the global scale will decrease the global rate of export production, but it could either increase or decrease the respired carbon content of the ocean interior, depending on where the reduced upward flux of nutrients occurs. Furthermore, viable candidates for physical climate forcing are numerous, including changes in the westerly winds, changes in the depth of the thermocline, and changes in the formation rate of North Atlantic Deep Water, among others. We use a simple, prognostic, light-and temperature-dependent model of biogeochemical cycling within a state-of-the- art global coupled ocean-atmosphere model to examine the response of the biological pump to changes in the coupled Earth system over multiple centuries. The biogeochemical model explicitly distinguishes respired carbon from preformed and saturation carbon, allowing the activity of the biological pump to be clearly quantified. Changes are forced in the model by altering the background climate state, and by manipulating the flux of freshwater to the North Atlantic region. We show how these changes in the physical state of the coupled ocean-atmosphere system impact the distribution and mass of respired carbon in the ocean interior, and the relationship these changes bear to global patterns of export production via the redistribution of nutrients.

Revealing and analyzing networks of environmental systems

NASA Astrophysics Data System (ADS)

Eveillard, D.; Bittner, L.; Chaffron, S.; Guidi, L.; Raes, J.; Karsenti, E.; Bowler, C.; Gorsky, G.

2015-12-01

Understanding the interactions between microbial communities and their environment well enough to be able to predict diversity on the basis of physicochemical parameters is a fundamental pursuit of microbial ecology that still eludes us. However, modeling microbial communities is a complicated task, because (i) communities are complex, (ii) most are described qualitatively, and (iii) quantitative understanding of the way communities interacts with their surroundings remains incomplete. Within this seminar, we will illustrate two complementary approaches that aim to overcome these points in different manners. First, we will present a network analysis that focus on the biological carbon pump in the global ocean. The biological carbon pump is the process by which photosynthesis transforms CO2 to organic carbon sinking to the deep-ocean as particles where it is sequestered. While the intensity of the pump correlate to plankton community composition, the underlying ecosystem structure and interactions driving this process remain largely uncharacterized Here we use environmental and metagenomic data gathered during the Tara Oceans expedition to improve understanding of these drivers. We show that specific plankton communities correlate with carbon export and highlight unexpected and overlooked taxa such as Radiolaria, alveolate parasites and bacterial pathogens, as well as Synechococcus and their phages, as key players in the biological pump. Additionally, we show that the abundances of just a few bacterial and viral genes predict most of the global ocean carbon export's variability. Together these findings help elucidate ecosystem drivers of the biological carbon pump and present a case study for scaling from genes-to-ecosystems. Second, we will show preliminary results on a probabilistic modeling that predicts microbial community structure across observed physicochemical data, from a putative network and partial quantitative knowledge. This modeling shows that, despite distinct quantitative environmental perturbations, the constraints on community structure could remain stable.

HIV-1 Rev protein specifies the viral RNA export pathway by suppressing TAP/NXF1 recruitment

PubMed Central

Taniguchi, Ichiro; Mabuchi, Naoto; Ohno, Mutsuhito

2014-01-01

Nuclear RNA export pathways in eukaryotes are often linked to the fate of a given RNA. Therefore, the choice of export pathway should be well-controlled to avoid an unfavorable effect on gene expression. Although some RNAs could be exported by more than one pathway, little is known about how the choice is regulated. This issue is highlighted when the human immunodeficiency virus type 1 (HIV-1) Rev protein induces the export of singly spliced and unspliced HIV-1 transcripts. How these RNAs are exported is not well understood because such transcripts should have the possibility of utilizing CRM1-dependent export via Rev or cellular TAP/NXF1-dependent export via the transcription/export (TREX) complex, or both. Here we found that Rev suppressed TAP/NXF1-dependent export of model RNA substrates that recapitulated viral transcripts. In this effect, Rev interacted with the cap-binding complex and inhibited the recruitment of the TREX complex. Thus, Rev controls the identity of the factor occupying the cap-proximal region that determines the RNA export pathway. This ribonucleoprotein remodeling activity of Rev may favor viral gene expression. PMID:24753416

Carbonate counter pump stimulated by natural iron fertilization in the Polar Frontal Zone

NASA Astrophysics Data System (ADS)

Salter, Ian; Schiebel, Ralf; Ziveri, Patrizia; Movellan, Aurore; Lampitt, Richard; Wolff, George A.

2014-12-01

The production of organic carbon in the ocean's surface and its subsequent downward export transfers carbon dioxide to the deep ocean. This CO2 drawdown is countered by the biological precipitation of carbonate, followed by sinking of particulate inorganic carbon, which is a source of carbon dioxide to the surface ocean, and hence the atmosphere over 100-1,000 year timescales. The net transfer of CO2 to the deep ocean is therefore dependent on the relative amount of organic and inorganic carbon in sinking particles. In the Southern Ocean, iron fertilization has been shown to increase the export of organic carbon, but it is unclear to what degree this effect is compensated by the export of inorganic carbon. Here we assess the composition of sinking particles collected from sediment traps located in the Polar Frontal Zone of the Southern Ocean. We find that in high-nutrient, low-chlorophyll regions that are characterized by naturally high iron concentrations, fluxes of both organic and inorganic carbon are higher than in regions with no iron fertilization. However, the excess flux of inorganic carbon is greater than that of organic carbon. We estimate that the production and flux of carbonate in naturally iron-fertilized waters reduces the overall amount of CO2 transferred to the deep ocean by 6-32%, compared to 1-4% at the non-fertilized site. We suggest that an increased export of organic carbon, stimulated by iron availability in the glacial sub-Antarctic oceans, may have been accompanied by a strengthened carbonate counter pump.

Randomised trial comparing hand expression with breast pumping for mothers of term newborns feeding poorly.

PubMed

Flaherman, Valerie J; Gay, Barbara; Scott, Cheryl; Avins, Andrew; Lee, Kathryn A; Newman, Thomas B

2012-01-01

Breast pumping or hand expression may be recommended when newborns latch or suck poorly. A recent trial found worse outcomes among mothers who used a breast pump in the early postpartum period. The objective of this study was to compare bilateral electric breast pumping to hand expression among mothers of healthy term infants feeding poorly at 12-36 h after birth. Randomised controlled trial. Well-baby nursery and postpartum unit. 68 mothers of newborns 12-36 h old who were latching or sucking poorly were randomly assigned to either 15 min of bilateral electric pumping or 15 min of hand expression. Milk transfer, maternal pain, breastfeeding confidence and breast milk expression experience (BMEE) immediately after the intervention, and breastfeeding rates at 2 months after birth. The median volume of expressed milk (range) was 0.5 (0-5) ml for hand expressing mothers and 1 (0-40) ml for pumping mothers (p=0.07). Maternal pain, breastfeeding confidence and BMEE did not differ by intervention. At 2 months, mothers assigned to hand expression were more likely to be breastfeeding (96.1%) than mothers assigned to breast pumping (72.7%) (p=0.02). Hand expression in the early postpartum period appears to improve eventual breastfeeding rates at 2 months after birth compared with breast pumping, but further research is needed to confirm this. However, in circumstances where either pumping or hand expression would be appropriate for healthy term infants 12-36 h old feeding poorly, providers should consider recommending hand expression.

Methods of milk expression for lactating women.

PubMed

Becker, Genevieve E; Smith, Hazel A; Cooney, Fionnuala

2016-09-29

Breastfeeding is important, however not all infants can feed at the breast and methods of expressing milk need evaluation. To assess acceptability, effectiveness, safety, effect on milk composition, contamination and costs of methods of milk expression. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register (21 March 2016), handsearched relevant journals and conference proceedings, and contacted experts in the field to seek additional published or unpublished studies. We also examined reference lists of all relevant retrieved papers. Randomised and quasi-randomised trials comparing methods at any time after birth. Three review authors independently assessed trials for inclusion and risk of bias, extracted data and checked them for accuracy. This updated review includes 41 trials involving 2293 participants, with 22 trials involving 1339 participants contributing data for analysis. Twenty-six of the trials referred to mothers of infants in neonatal units (n = 1547) and 14 to mothers of healthy infants at home (n = 730), with one trial containing mothers of both neonatal and healthy older infants (n = 16). Eleven trials compared one or more types of pump versus hand expression and 14 studies compared one type of pump versus another type of pump, with three of these studies comparing both hand expression and pump types. Twenty studies compared a specific protocol or adjunct behaviour including sequential versus simultaneous pumping protocols, pumping frequency, provision of an education and support intervention, relaxation, breast massage, combining hand expression with pumping and a breast cleansing protocol.Due to heterogeneity in participants, interventions, and outcomes measured or reported, we were unable to pool findings for most of the specified outcomes. It was not possible therefore to produce a 'Summary of findings' table in this update. Most of the included results were derived from single studies. Trials took place in 14 countries under a variety of circumstances and were published from 1982 to 2015. Sixteen of the 30 trials that evaluated pumps or products had support from the manufacturers. The risk of bias of the included studies was variable. Primary outcomesOnly one of the 17 studies examining maternal satisfaction/acceptability with the method or adjunct behaviour provided data suitable for analysis. In this study, self-efficacy was assessed by asking mothers if they agreed or disagreed with the following statement: 'I don't want anyone to see me (hand expressing/pumping)'. The study found that mothers who were using the electric pump were more likely to agree with the statement compared to mothers hand expressing, (mean difference (MD) 0.70, 95% confidence interval (CI) 0.15 to 1.25; P = 0.01, participants = 68). Mothers who were hand expressing reported that the instructions for expression were clearer compared to the electric pump, (MD -0.40, 95% CI -0.75 to -0.05; P = 0.02, participants = 68). Descriptive reporting of satisfaction in the other studies varied in the measures used, did not indicate a clear preference for one pump type, although there was satisfaction with some relaxation and support interventions.We found no clinically significant differences between methods related to contamination of the milk that compared any type of pump to hand expression (risk ratio (RR) 1.13, 95% CI 0.79 to 1.61; P = 0.51, participants = 28), manual pump compared to hand expression, (MD 0.20, 95% CI -0.18 to 0.58; P = 0.30, participants = 142) a large electric pump compared to hand expression (MD 0.10, 95% CI -0.29 to 0.49; P = 0.61, participants = 123), or a large electric pump compared to a manual pump (MD -0.10, 95% CI -0.46 to 0.26; P = 0.59, participants = 141).The level of maternal breast or nipple pain or damage was similar in comparisons of a large electric pump to hand expression (MD 0.02, 95% CI -0.67 to 0.71; P = 0.96, participants = 68). A study comparing a manual and large electric pump, reported sore nipples in 7% for both groups and engorgement in 4% using a manual pump versus 6% using an electric pump; and in one study no nipple damage was reported in the hand-expression group, and one case of nipple damage in each of the manual pump and the large electric pump groups.One study examined adverse effects on infants, however as the infants did not all receive their mothers' expressed milk, we have not included the results. Secondary outcomesThe quantity of expressed milk obtained was increased, in some studies by a clinically significant amount, in interventions involving relaxation, music, warmth, massage, initiation of pumping, increased frequency of pumping and suitable breast shield size. Support programmes and simultaneous compared to sequential pumping did not show a difference in milk obtained. No pump consistently increased the milk volume obtained significantly.In relation to nutrient quality, hand expression or a large electric pump were found to provide higher protein than a manual pump, and hand expression provided higher sodium and lower potassium compared to a large electric pump or a manual pump. Fat content was higher with breast massage when pumping; no evidence of difference was found for energy content between methods.No consistent effect was found related to prolactin change or effect on oxytocin release with pump type or method. Economic aspects were not reported. The most suitable method for milk expression may depend on the time since birth, purpose of expression and the individual mother and infant. Low-cost interventions including initiation of milk expression sooner after birth when not feeding at the breast, relaxation, massage, warming the breasts, hand expression and lower cost pumps may be as effective, or more effective, than large electric pumps for some outcomes. Variation in nutrient content across methods may be relevant to some infants. Small sample sizes, large standard deviations, and the diversity of the interventions argue caution in applying these results beyond the specific method tested in the specific settings. Independently funded research is needed for more trials on hand expression, relaxation and other techniques that do not have a commercial potential.

MCT1 modulates cancer cell pyruvate export and growth of tumors that co-express MCT1 and MCT4

PubMed Central

Hong, Candice Sun; Graham, Nicholas A.; Gu, Wen; Camacho, Carolina Espindola; Mah, Vei; Maresh, Erin L.; Alavi, Mohammed; Bagryanova, Lora; Krotee, Pascal A. L.; Gardner, Brian K.; Behbahan, Iman Saramipoor; Horvath, Steve; Chia, David; Mellinghoff, Ingo K.; Hurvitz, Sara A.; Dubinett, Steven M.; Critchlow, Susan E.; Kurdistani, Siavash K.; Goodglick, Lee; Braas, Daniel; Graeber, Thomas G.; Christofk, Heather R.

2016-01-01

SUMMARY Monocarboxylate Transporter 1 (MCT1) inhibition is thought to block tumor growth through disruption of lactate transport and glycolysis. Here we show MCT1 inhibition impairs proliferation of glycolytic breast cancer cells co-expressing MCT1 and MCT4 via disruption of pyruvate rather than lactate export. MCT1 expression is elevated in glycolytic breast tumors, and high MCT1 expression predicts poor prognosis in breast and lung cancer patients. Acute MCT1 inhibition reduces pyruvate export but does not consistently alter lactate transport or glycolytic flux in breast cancer cells that co-express MCT1 and MCT4. Despite the lack of glycolysis impairment, MCT1 loss-of-function decreases breast cancer cell proliferation and blocks growth of mammary fat pad xenograft tumors. Our data suggest MCT1 expression is elevated in glycolytic cancers to promote pyruvate export, which when inhibited enhances oxidative metabolism and reduces proliferation. This study presents an alternative molecular consequence of MCT1 inhibitors further supporting their use as anti-cancer therapeutics. PMID:26876179

MCT1 Modulates Cancer Cell Pyruvate Export and Growth of Tumors that Co-express MCT1 and MCT4.

PubMed

Hong, Candice Sun; Graham, Nicholas A; Gu, Wen; Espindola Camacho, Carolina; Mah, Vei; Maresh, Erin L; Alavi, Mohammed; Bagryanova, Lora; Krotee, Pascal A L; Gardner, Brian K; Behbahan, Iman Saramipoor; Horvath, Steve; Chia, David; Mellinghoff, Ingo K; Hurvitz, Sara A; Dubinett, Steven M; Critchlow, Susan E; Kurdistani, Siavash K; Goodglick, Lee; Braas, Daniel; Graeber, Thomas G; Christofk, Heather R

2016-02-23

Monocarboxylate transporter 1 (MCT1) inhibition is thought to block tumor growth through disruption of lactate transport and glycolysis. Here, we show MCT1 inhibition impairs proliferation of glycolytic breast cancer cells co-expressing MCT1 and MCT4 via disruption of pyruvate rather than lactate export. MCT1 expression is elevated in glycolytic breast tumors, and high MCT1 expression predicts poor prognosis in breast and lung cancer patients. Acute MCT1 inhibition reduces pyruvate export but does not consistently alter lactate transport or glycolytic flux in breast cancer cells that co-express MCT1 and MCT4. Despite the lack of glycolysis impairment, MCT1 loss-of-function decreases breast cancer cell proliferation and blocks growth of mammary fat pad xenograft tumors. Our data suggest MCT1 expression is elevated in glycolytic cancers to promote pyruvate export that when inhibited, enhances oxidative metabolism and reduces proliferation. This study presents an alternative molecular consequence of MCT1 inhibitors, further supporting their use as anti-cancer therapeutics. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

In Vitro Comparison of Adipokine Export Signals.

PubMed

Sharafi, Parisa; Kocaefe, Y Çetin

2016-01-01

Mammalian cells are widely used for recombinant protein production in research and biotechnology. Utilization of export signals significantly facilitates production and purification processes. 35 years after the discovery of the mammalian export machinery, there still are obscurities regarding the efficiency of the export signals. The aim of this study was the comparative evaluation of the efficiency of selected export signals using adipocytes as a cell model. Adipocytes have a large capacity for protein secretion including several enzymes, adipokines, and other signaling molecules, providing a valid system for a quantitative evaluation. Constructs that expressed N-terminal fusion export signals were generated to express Enhanced Green Fluorescence Protein (EGFP) as a reporter for quantitative and qualitative evaluation. Furthermore, fluorescent microscopy was used to trace the intracellular traffic of the reporter. The export efficiency of six selected proteins secreted from adipocytes was evaluated. Quantitative comparison of intracellular and exported fractions of the recombinant constructs demonstrated a similar efficiency among the studied sequences with minor variations. The export signal of Retinol Binding Protein (RBP4) exhibited the highest efficiency. This study presents the first quantitative data showing variations among export signals, in adipocytes which will help optimization of recombinant protein distribution.

Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

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

Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto

2009-12-18

The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins aremore » known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.« less

Analysis of Zinc-Exporters Expression in Prostate Cancer.

PubMed

Singh, Chandra K; Malas, Kareem M; Tydrick, Caitlin; Siddiqui, Imtiaz A; Iczkowski, Kenneth A; Ahmad, Nihal

2016-11-11

Maintaining optimal intracellular zinc (Zn) concentration is crucial for critical cellular functions. Depleted Zn has been associated with prostate cancer (PCa) progression. Solute carrier family 30 (SLC30A) proteins maintain cytoplasmic Zn balance by exporting Zn out to the extracellular space or by sequestering cytoplasmic Zn into intracellular compartments. In this study, we determined the involvement of Zn-exporters, SLC30A 1-10 in PCa, in the context of racial health disparity in human PCa samples obtained from European-American (EA) and African-American (AA) populations. We also analyzed the levels of Zn-exporters in a panel of PCa cells derived from EA and AA populations. We further explored the expression profile of Zn-exporters in PCa using Oncomine database. Zn-exporters were found to be differentially expressed at the mRNA level, with a significant upregulation of SLC30A1, SLC30A9 and SLC30A10, and downregulation of SLC30A5 and SLC30A6 in PCa, compared to benign prostate. Moreover, Ingenuity Pathway analysis revealed several interactions of Zn-exporters with certain tumor suppressor and promoter proteins known to be modulated in PCa. Our study provides an insight regarding Zn-exporters in PCa, which may open new avenues for future studies aimed at enhancing the levels of Zn by modulating Zn-transporters via pharmacological means.

Analysis of Zinc-Exporters Expression in Prostate Cancer

PubMed Central

Singh, Chandra K.; Malas, Kareem M.; Tydrick, Caitlin; Siddiqui, Imtiaz A.; Iczkowski, Kenneth A.; Ahmad, Nihal

2016-01-01

Maintaining optimal intracellular zinc (Zn) concentration is crucial for critical cellular functions. Depleted Zn has been associated with prostate cancer (PCa) progression. Solute carrier family 30 (SLC30A) proteins maintain cytoplasmic Zn balance by exporting Zn out to the extracellular space or by sequestering cytoplasmic Zn into intracellular compartments. In this study, we determined the involvement of Zn-exporters, SLC30A 1–10 in PCa, in the context of racial health disparity in human PCa samples obtained from European-American (EA) and African-American (AA) populations. We also analyzed the levels of Zn-exporters in a panel of PCa cells derived from EA and AA populations. We further explored the expression profile of Zn-exporters in PCa using Oncomine database. Zn-exporters were found to be differentially expressed at the mRNA level, with a significant upregulation of SLC30A1, SLC30A9 and SLC30A10, and downregulation of SLC30A5 and SLC30A6 in PCa, compared to benign prostate. Moreover, Ingenuity Pathway analysis revealed several interactions of Zn-exporters with certain tumor suppressor and promoter proteins known to be modulated in PCa. Our study provides an insight regarding Zn-exporters in PCa, which may open new avenues for future studies aimed at enhancing the levels of Zn by modulating Zn-transporters via pharmacological means. PMID:27833104

Types of Breast Pumps

MedlinePlus

... nipple (the areola). b. Pump: creates the gentle vacuum that expresses milk. The pump may be attached to the breast-shield or ... out of a larger tube to create a vacuum that expresses milk and collects it in an attached container. Another type of manual pump, called a bicycle horn pump, consists of a ...

Modelling the regulation effects of lowland polder with pumping station on hydrological processes and phosphorus loads.

PubMed

Yan, Renhua; Li, Lingling; Gao, Junfeng

2018-05-08

Exploring the hydrological regulation of a lowland polder is essential for increasing knowledge regarding the role of polders associated with pumping stations in lowlands. In this study, the Lowland Polder Hydrology and Phosphorus modelling System (PHPS) was applied to the Jianwei polder as a case study for quantifying the regulation effects of a lowland polder with pumping on discharge and phosphorus loads. The results indicate that the polder significantly affected the temporal distribution and annual amount of catchment discharge. Compared with a no-pumping scenario, an agricultural polder with pumping stations generated a sharper discharge hydrograph with higher peak-values and lower minimum-values, as well as an 8.6% reduction in average annual discharge. It also decreased the phosphorus export to downstream water bodies by 5.33 kg/hm 2 /yr because of widespread ditches and ponds, a lower hydraulic gradient, and increased retention times of surface water in ponds. Copyright © 2018 Elsevier B.V. All rights reserved.

The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA.

PubMed

Booth, David S; Cheng, Yifan; Frankel, Alan D

2014-12-08

The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-Ran(GTP) nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

Multidrug efflux pumps: the structures of prokaryotic ATP-binding cassette transporter efflux pumps and implications for our understanding of eukaryotic P-glycoproteins and homologues.

PubMed

Kerr, Ian D; Jones, Peter M; George, Anthony M

2010-02-01

One of the Holy Grails of ATP-binding cassette transporter research is a structural understanding of drug binding and transport in a eukaryotic multidrug resistance pump. These transporters are front-line mediators of drug resistance in cancers and represent an important therapeutic target in future chemotherapy. Although there has been intensive biochemical research into the human multidrug pumps, their 3D structure at atomic resolution remains unknown. The recent determination of the structure of a mouse P-glycoprotein at subatomic resolution is complemented by structures for a number of prokaryotic homologues. These structures have provided advances into our knowledge of the ATP-binding cassette exporter structure and mechanism, and have provided the template data for a number of homology modelling studies designed to reconcile biochemical data on these clinically important proteins.

Molecular mechanisms of gravity-dependent signaling in human melanocytic cells involve cyclic GMP

NASA Astrophysics Data System (ADS)

Ivanova, Krassimira; Lambers, Britta; Block, Ingrid; Bromeis, Birgit; Das, Pranab K.; Gerzer, Rupert

2005-08-01

Gravity alteration (micro- and hypergravity) is known to influence cell functions. As guanosine 3',5'-cyclic monophosphate (cGMP) is an important messenger in melanocyte signaling we have compared the regulation of cGMP levels in human melanocytes and melanoma cells with different metastatic potential under hypergravity conditions. We were able to demonstrate that long-term exposure to hypergravity stimulates cGMP efflux in cultured human melanocytes and non- metastatic melanoma cells, whereas highly metastatic melanoma cells appear to be insensitive to hypergravity, most probably, due to an up-regulated cGMP efflux at 1g. Here we report that these effects are associated with the expression of the multidrug resistance proteins 4 and 5 known to act as selective export pumps for amphiphilic anions like cGMP. Thus, an altered gravity vector may induce cGMP-dependent signaling events in melanocytic cells that could be important for malignant transformation.

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

Smirnova, Anna S.; Morgun, Andrey; Shulzhenko, Natalia

Two transcript variants (TV) of the T cell immune regulator gene 1 (TCIRG1) have already been characterized. TV1 encodes a subunit of the osteoclast vacuolar proton pump and TV2 encodes a T cell inhibitory receptor. Based on the search in dbEST, we validated by RT-PCR six new alternative splice events in TCIRG1 in most of the 28 human tissues studied. In addition, we observed that transcripts using the TV1 transcription start site and two splice forms previously described in a patient with infantile malignant osteopetrosis are also expressed in various tissues of healthy individuals. Studies of these nine splice formsmore » in cytoplasmic RNA of peripheral blood mononuclear cells showed that at least six of them could be efficiently exported from the nucleus. Since various products with nearly ubiquitous tissue distribution are generated from TCIRG1, this gene may be involved in other processes besides immune response and bone resorption.« less

The effect of acetaminophen on the expression of BCRP in trophoblast cells impairs the placental barrier to bile acids during maternal cholestasis

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

Blazquez, Alba G., E-mail: albamgb@usal.es; CIBERehd, Instituto de Salud Carlos III, Madrid; Briz, Oscar, E-mail: obriz@usal.es

Acetaminophen is used as first-choice drug for pain relief during pregnancy. Here we have investigated the effect of acetaminophen at subtoxic doses on the expression of ABC export pumps in trophoblast cells and its functional repercussion on the placental barrier during maternal cholestasis. The incubation of human choriocarcinoma cells (JAr, JEG-3 and BeWo) with acetaminophen for 48 h resulted in no significant changes in the expression and/or activity of MDR1 and MRPs. In contrast, in JEG-3 cells, BCRP mRNA, protein, and transport activity were reduced. In rat placenta, collected at term, acetaminophen administration for the last three days of pregnancymore » resulted in enhanced mRNA, but not protein, levels of Mrp1 and Bcrp. In fact, a decrease in Bcrp protein was found. Using in situ perfused rat placenta, a reduction in the Bcrp-dependent fetal-to-maternal bile acid transport after treating the dams with acetaminophen was found. Complete biliary obstruction in pregnant rats induced a significant bile acid accumulation in fetal serum and tissues, which was further enhanced when the mothers were treated with acetaminophen. This drug induced increased ROS production in JEG-3 cells and decreased the total glutathione content in rat placenta. Moreover, the NRF2 pathway was activated in JEG-3 cells as shown by an increase in nuclear NRF2 levels and an up-regulation of NRF2 target genes, NQO1 and HMOX-1, which was not observed in rat placenta. In conclusion, acetaminophen induces in placenta oxidative stress and a down-regulation of BCRP/Bcrp, which may impair the placental barrier to bile acids during maternal cholestasis. - Highlights: • Acetaminophen induces changes in placental BCRP expression in vitro. • This drug reduces the ability of placental cells to export BCRP substrates. • Acetaminophen induces changes in Bcrp expression in rat placenta. • Placental barrier to bile acids is impaired in rats treated with this drug.« less

Enhanced open ocean storage of CO2 from shelf sea pumping.

PubMed

Thomas, Helmuth; Bozec, Yann; Elkalay, Khalid; de Baar, Hein J W

2004-05-14

Seasonal field observations show that the North Sea, a Northern European shelf sea, is highly efficient in pumping carbon dioxide from the atmosphere to the North Atlantic Ocean. The bottom topography-controlled stratification separates production and respiration processes in the North Sea, causing a carbon dioxide increase in the subsurface layer that is ultimately exported to the North Atlantic Ocean. Globally extrapolated, the net uptake of carbon dioxide by coastal and marginal seas is about 20% of the world ocean's uptake of anthropogenic carbon dioxide, thus enhancing substantially the open ocean carbon dioxide storage.

Menadione metabolism to thiodione in hepatoblastoma by scanning electrochemical microscopy

PubMed Central

Mauzeroll, Janine; Bard, Allen J.; Owhadian, Omeed; Monks, Terrence J.

2004-01-01

The cytotoxicity of menadione on hepatocytes was studied by using the substrate generation/tip collection mode of scanning electrochemical microscopy by exposing the cells to menadione and detecting the menadione-S-glutathione conjugate (thiodione) that is formed during the cellular detoxication process and is exported from the cell by an ATP-dependent pump. This efflux was electrochemically detected and allowed scanning electrochemical microscopy monitoring and imaging of single cells and groups of highly confluent live cells. Based on a constant flux model, ≈6 × 106 molecules of thiodione per cell per second are exported from monolayer cultures of Hep G2 cells. PMID:15601769

Temporal and spatial regulation of mRNA export: Single particle RNA-imaging provides new tools and insights

PubMed Central

Heinrich, Stephanie; Derrer, Carina Patrizia; Lari, Azra; Weis, Karsten; Montpetit, Ben

2017-01-01

The transport of messenger RNAs (mRNAs) from the nucleus to cytoplasm is an essential step in the gene expression program of all eukaryotes. Recent technological advances in the areas of RNA-labeling, microscopy, and sequencing are leading to novel insights about mRNA biogenesis and export. This includes quantitative single molecule imaging (SMI) of RNA molecules in live cells, which is providing knowledge of the spatial and temporal dynamics of the export process. As this information becomes available, it leads to new questions, the reinterpretation of previous findings, and revised models of mRNA export. In this review, we will briefly highlight some of these recent findings and discuss how live cell SMI approaches may be used to further our current understanding of mRNA export and gene expression. PMID:28052353

Efflux systems in bacteria and their metabolic engineering applications.

PubMed

Jones, Christopher M; Hernández Lozada, Néstor J; Pfleger, Brian F

2015-11-01

The production of valuable chemicals from metabolically engineered microbes can be limited by excretion from the cell. Efflux is often overlooked as a bottleneck in metabolic pathways, despite its impact on alleviating feedback inhibition and product toxicity. In the past, it has been assumed that endogenous efflux pumps and membrane porins can accommodate product efflux rates; however, there are an increasing number of examples wherein overexpressing efflux systems is required to improve metabolite production. In this review, we highlight specific examples from the literature where metabolite export has been studied to identify unknown transporters, increase tolerance to metabolites, and improve the production capabilities of engineered bacteria. The review focuses on the export of a broad spectrum of valuable chemicals including amino acids, sugars, flavins, biofuels, and solvents. The combined set of examples supports the hypothesis that efflux systems can be identified and engineered to confer export capabilities on industrially relevant microbes.

Role of the mRNA export factor Sus1 in oxidative stress tolerance in Candida albicans.

PubMed

Xiao, Chenpeng; Yu, Qilin; Zhang, Bing; Li, Jianrong; Zhang, Dan; Li, Mingchun

2018-02-05

In eukaryotes, the nuclear export of mRNAs is essential for gene expression. However, little is known about the role of mRNA nuclear export in the important fungal pathogen, Candida albicans. In this study, we identified C. albicans Sus1, a nucleus-localized protein that is required for mRNA export. Interestingly, the sus1Δ/Δ displayed hyper-sensitivity to extracellular oxidative stress, enhanced ROS accumulation and severe oxidative stress-related cell death. More strikingly, although the mutant exhibited normal activation of the expression of oxidative stress response (OSR) genes, it had attenuated activity of ROS scavenging system, which may be attributed to the defect in OSR mRNA export in this mutant. In addition, the virulence of the sus1Δ/Δ was seriously attenuated. Taken together, our findings provide evidence that the mRNA export factor Sus1 plays an important role in oxidative stress tolerance and pathogenesis. Copyright © 2018 Elsevier Inc. All rights reserved.

Quantifying export production in the Southern Ocean: Implications for the Baxs proxy

NASA Astrophysics Data System (ADS)

Hernandez-Sanchez, Maria T.; Mills, Rachel A.; Planquette, HéLèNe; Pancost, Richard D.; Hepburn, Laura; Salter, Ian; Fitzgeorge-Balfour, Tania

2011-12-01

The water column and sedimentary Baxs distribution around the Crozet Plateau is used to decipher the controls and timing of barite formation and to evaluate how export production signals are recorded in sediments underlying a region of natural Fe fertilization within the Fe limited Southern Ocean. Export production estimated from preserved, vertical sedimentary Baxs accumulation rates are compared with published export fluxes assessed from an integrated study of the biological carbon pump to determine the validity of Baxs as a quantitative proxy under different Fe supply conditions typical of the Southern Ocean. Detailed assessment of the geochemical partitioning of Ba in sediments and the lithogenic end-member allows appropriate correction of the bulk Ba content and determination of the Baxs content of sediments and suspended particles. The upper water column distribution of Baxs is extremely heterogeneous spatially and temporally. Organic carbon/Baxs ratios in deep traps from the Fe fertilized region are similar to other oceanic settings allowing quantification of the inferred carbon export based on established algorithms. There appears to be some decoupling of POC and Ba export in the Fe limited region south of the Plateau. The export production across the Crozet Plateau inferred from the Baxs sedimentary proxy indicates that the Fe fertilized area to the north of the Plateau experiences enhanced export relative to equivalent Southern Ocean settings throughout the Holocene and that this influence may also have impacted the site to the south for significant periods. This interpretation is corroborated by alternative productivity proxies (opal accumulation, 231Paxs/230Thxs). Baxs can be used to quantify export production in complex settings such as naturally Fe-fertilized (volcanoclastic) areas, providing appropriate lithogenic correction is undertaken, and sediment focusing is corrected for along with evaluation of barite preservation.

Tap and Dbp5, but not Gag, are involved in DR-mediated nuclear export of unspliced Rous sarcoma virus RNA

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

LeBlanc, Jason J.; Uddowla, Sabena; Abraham, Benjamin

2007-07-05

All retroviruses must circumvent cellular restrictions on the export of unspliced RNAs from the nucleus. While the unspliced RNA export pathways for HIV and Mason-Pfizer monkey virus are well characterized, that of Rous sarcoma virus (RSV) is not. We have previously reported that the RSV direct repeat (DR) elements are involved in the cytoplasmic accumulation of unspliced viral RNA. Here, using fluorescent in situ hybridization (FISH), we demonstrate that unspliced viral RNAs bearing a single point mutation (G8863C) in the DR exhibit a restricted cellular localization in and around the nucleus. In contrast, wild type unspliced viral RNA had amore » diffuse localization throughout the nucleus and cytoplasm. Since the RSV Gag protein has a transient localization in the nucleus, we examined the effect of Gag over-expression on a DR-mediated reporter construct. While Gag did not enhance DR-mediated nuclear export, the dominant-negative expression of two cellular export factors, Tap and Dbp5, inhibited expression of the same reporter construct. Furthermore, FISH studies using the dominant-negative Dbp5 demonstrated that unspliced wild type RSV RNA was retained within the nucleus. Taken together, these results further implicate the DR in nuclear RNA export through interactions with Tap and Dbp5.« less

Expression of Iron-Related Proteins at the Neurovascular Unit Supports Reduction and Reoxidation of Iron for Transport Through the Blood-Brain Barrier.

PubMed

Burkhart, Annette; Skjørringe, Tina; Johnsen, Kasper Bendix; Siupka, Piotr; Thomsen, Louiza Bohn; Nielsen, Morten Schallburg; Thomsen, Lars Lykke; Moos, Torben

2016-12-01

The mechanisms for iron transport through the blood-brain barrier (BBB) remain a controversy. We analyzed for expression of mRNA and proteins involved in oxidation and transport of iron in isolated brain capillaries from dietary normal, iron-deficient, and iron-reverted rats. The expression was also investigated in isolated rat brain endothelial cells (RBECs) and in immortalized rat brain endothelial (RBE4) cells grown as monoculture or in hanging culture inserts with defined BBB properties. Transferrin receptor 1, ferrireductases Steap 2 and 3, divalent metal transporter 1 (DMT1), ferroportin, soluble and glycosylphosphatidylinositol (GPI)-anchored ceruloplasmin, and hephaestin were all expressed in brain capillaries in vivo and in isolated RBECs and RBE4 cells. Gene expression of DMT1, ferroportin, and soluble and GPI-anchored ceruloplasmin were significantly higher in isolated RBECs with induced BBB properties. Primary pericytes and astrocytes both expressed ceruloplasmin and hephaestin, and RBECs, pericytes, and astrocytes all exhibited ferrous oxidase activity. The coherent protein expression of these genes was demonstrated by immunocytochemistry. The data show that brain endothelial cells provide the machinery for receptor-mediated uptake of ferric iron-containing transferrin. Ferric iron can then undergo reduction to ferrous iron by ferrireductases inside endosomes followed by DMT1-mediated pumping into the cytosol and subsequently cellular export by ferroportin. The expression of soluble ceruloplasmin by brain endothelial cells, pericytes, and astrocytes that together form the neurovascular unit (NVU) provides the ferroxidase activity necessary to reoxidize ferrous iron once released inside the brain.

Estimates of micro-, nano-, and picoplankton contributions to particle export in the northeast Pacific

NASA Astrophysics Data System (ADS)

Mackinson, B. L.; Moran, S. B.; Lomas, M. W.; Stewart, G. M.; Kelly, R. P.

2015-06-01

The contributions of micro-, nano-, and picoplankton to particle export were estimated from measurements of size-fractionated particulate 234Th, organic carbon, and phytoplankton indicator pigments obtained during five cruises between 2010 and 2012 along Line P in the subarctic northeast Pacific Ocean. Sinking fluxes of particulate organic carbon (POC) and indicator pigments were calculated from 234Th-238U disequilibria and, during two cruises, measured by a sediment trap at Ocean Station Papa. POC fluxes at 100 m ranged from 0.65 to 7.95 mmol m-2 d-1, similar in magnitude to previous results at Line P. Microplankton pigments dominate indicator pigment fluxes (averaging 69 ± 19% of total pigment flux), while nanoplankton pigments comprised the majority of pigment standing stocks (averaging 64 ± 23% of total pigment standing stocks). Indicator pigment loss rates (the ratio of pigment export flux to pigment standing stocks) point to preferential export of larger microplankton relative to smaller nano- and picoplankton. However, indicator pigments do not quantitatively trace particle export resulting from zooplankton grazing, which may be an important pathway for the export of small phytoplankton. These results have important implications for understanding the magnitude and mechanisms controlling the biological pump at Line P in particular, and more generally in oligotrophic gyres and high-nutrient, low-chlorophyll (HNLC) regions where small phytoplankton represent a major component of the autotrophic community.

Estimates of micro-, nano-, and picoplankton contributions to particle export in the northeast Pacific

NASA Astrophysics Data System (ADS)

Mackinson, B. L.; Moran, S. B.; Lomas, M. W.; Stewart, G. M.; Kelly, R. P.

2014-08-01

The contributions of micro-, nano-, and picoplankton to particle export were estimated from measurements of size-fractionated particulate 234Th, organic carbon, and phytoplankton indicator pigments obtained during five cruises between 2010 and 2012 along Line P in the subarctic northeast Pacific Ocean. Sinking fluxes of particulate organic carbon (POC) and indicator pigments were calculated from 234Th-238U disequilibria and, during two cruises, measured by sediment trap at Ocean Station Papa. POC fluxes at 100 m ranged from 0.65-7.95 mmol m-2 d-1, similar in magnitude to previous results at Line P. Microplankton pigments dominate indicator pigment fluxes (averaging 69 ± 19% of total pigment flux), while nanoplankton pigments comprised the majority of pigment standing stocks (averaging 64 ± 23% of total pigment standing stock). Indicator pigment loss rates (the ratio of pigment export flux to pigment standing stock) point to preferential export of larger microplankton relative to smaller nano- and picoplankton. However, indicator pigments do not quantitatively trace particle export resulting from zooplankton grazing, which may be an important pathway for the export of small phytoplankton. These results have important implications for understanding the magnitude and mechanisms controlling the biological pump at Line P in particular, and more generally in oligotrophic gyres and high-nutrient, low-chlorophyll regions where small phytoplankton represent a major component of the autotrophic community.

Nuclear Export of Messenger RNA

PubMed Central

Katahira, Jun

2015-01-01

Transport of messenger RNA (mRNA) from the nucleus to the cytoplasm is an essential step of eukaryotic gene expression. In the cell nucleus, a precursor mRNA undergoes a series of processing steps, including capping at the 5' ends, splicing and cleavage/polyadenylation at the 3' ends. During this process, the mRNA associates with a wide variety of proteins, forming a messenger ribonucleoprotein (mRNP) particle. Association with factors involved in nuclear export also occurs during transcription and processing, and thus nuclear export is fully integrated into mRNA maturation. The coupling between mRNA maturation and nuclear export is an important mechanism for providing only fully functional and competent mRNA to the cytoplasmic translational machinery, thereby ensuring accuracy and swiftness of gene expression. This review describes the molecular mechanism of nuclear mRNA export mediated by the principal transport factors, including Tap-p15 and the TREX complex. PMID:25836925

High-frequency monitoring reveals nutrient sources and transport processes in an agriculture-dominated lowland water system

NASA Astrophysics Data System (ADS)

van der Grift, Bas; Broers, Hans Peter; Berendrecht, Wilbert; Rozemeijer, Joachim; Osté, Leonard; Griffioen, Jasper

2016-05-01

Many agriculture-dominated lowland water systems worldwide suffer from eutrophication caused by high nutrient loads. Insight in the hydrochemical functioning of embanked polder catchments is highly relevant for improving the water quality in such areas or for reducing export loads to downstream water bodies. This paper introduces new insights in nutrient sources and transport processes in a polder in the Netherlands situated below sea level using high-frequency monitoring technology at the outlet, where the water is pumped into a higher situated lake, combined with a low-frequency water quality monitoring programme at six locations within the drainage area. Seasonal trends and short-scale temporal dynamics in concentrations indicated that the NO3 concentration at the pumping station originated from N loss from agricultural lands. The NO3 loads appear as losses via tube drains after intensive rainfall events during the winter months due to preferential flow through the cracked clay soil. Transfer function-noise modelling of hourly NO3 concentrations reveals that a large part of the dynamics in NO3 concentrations during the winter months can be related to rainfall. The total phosphorus (TP) concentration and turbidity almost doubled during operation of the pumping station, which points to resuspension of particulate P from channel bed sediments induced by changes in water flow due to pumping. Rainfall events that caused peaks in NO3 concentrations did not results in TP concentration peaks. The rainfall induced and NO3 enriched quick interflow, may also be enriched in TP but retention of TP due to sedimentation of particulate P then results in the absence of rainfall induced TP concentration peaks. Increased TP concentrations associated with run-off events is only observed during a rainfall event at the end of a freeze-thaw cycle. All these observations suggest that the P retention potential of polder water systems is primarily due to the artificial pumping regime that buffers high flows. As the TP concentration is affected by operation of the pumping station, timing of sampling relative to the operating hours of the pumping station should be accounted for when calculating P export loads, determining trends in water quality, or when judging water quality status of polder water systems.

Structures and transport dynamics of a Campylobacter jejuni multidrug efflux pump

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

Su, Chih-Chia; Yin, Linxiang; Kumar, Nitin

2017-08-01

Resistance-nodulation-cell division efflux pumps are integral membrane proteins that catalyze the export of substrates across cell membranes. Within the hydrophobe-amphiphile efflux subfamily, these resistance-nodulation-cell division proteins largely form trimeric efflux pumps. The drug efflux process has been proposed to entail a synchronized motion between subunits of the trimer to advance the transport cycle, leading to the extrusion of drug molecules. Here we use X-ray crystallography and single-molecule fluorescence resonance energy transfer imaging to elucidate the structures and functional dynamics of the Campylobacter jejuni CmeB multidrug efflux pump. We find that the CmeB trimer displays a very unique conformation. A directmore » observation of transport dynamics in individual CmeB trimers embedded in membrane vesicles indicates that each CmeB subunit undergoes conformational transitions uncoordinated and independent of each other. On the basis of our findings and analyses, we propose a model for transport mechanism where CmeB protomers function independently within the trimer.« less

The impact on atmospheric CO2 of iron fertilization induced changes in the ocean's biological pump

NASA Astrophysics Data System (ADS)

Jin, X.; Gruber, N.; Frenzel, H.; Doney, S. C.; McWilliams, J. C.

2007-10-01

Using numerical simulations, we quantify the impact of changes in the ocean's biological pump on the air-sea balance of CO2 by fertilizing a small surface patch in the high-nutrient, low-chlorophyll region of the eastern tropical Pacific with iron. Decade-long fertilization experiments are conducted in a basin-scale, eddy-permitting coupled physical biogeochemical ecological model. In contrast to previous studies, we find that most of the dissolved inorganic carbon (DIC) removed from the euphotic zone by the enhanced biological export is replaced by uptake of CO2 from the atmosphere. Atmospheric uptake efficiencies, the ratio of the perturbation in air-sea CO2 flux to the perturbation in export flux across 100 m, are 0.75 to 0.93 in our patch size-scale experiments. The atmospheric uptake efficiency is insensitive to the duration of the experiment. The primary factor controlling the atmospheric uptake efficiency is the vertical distribution of the enhanced biological production. Iron fertilization at the surface tends to induce production anomalies primarily near the surface, leading to high efficiencies. In contrast, mechanisms that induce deep production anomalies (e.g. altered light availability) tend to have a low uptake efficiency, since most of the removed DIC is replaced by lateral and vertical transport and mixing. Despite high atmospheric uptake efficiencies, patch-scale iron fertilization of the ocean's biological pump tends to remove little CO2 from the atmosphere over the decadal timescale considered here.

Multidrug resistance: prospects for clinical management.

PubMed

Mansouri, A; Henle, K J; Nagle, W A

1992-01-01

Clinical success in the treatment of tumors with chemotherapy has significantly improved over the past several years. However, treatment failures due to drug resistance of cancer cells has remained a major problem. The classical form of multiple drug resistance is perhaps also the most common type of drug resistance, and represents the overexpression of a transmembrane glycoprotein pump (P-170) that mediates the efflux of a spectrum of structurally and functionally unrelated drugs. Here, we discuss recent evidence that support the concept that the total phenomenon of multiple drug resistance (MDR) involves several other mechanisms in addition to that underlying "classical" MDR. These include the action of other energy-dependent membrane efflux pumps, elevated levels of GSH for drug conjugation and detoxification to facilitate export, enhanced DNA repair facility, gene amplification and oncogene activation. The combination of mechanisms used by any particular cell line is variable and suggests that many of these mechanisms are independent. Successful reversal of drug resistance appears to require the identification of relevant operative resistance mechanisms. An example is the competitive inhibition of P-170 with verapamil, quinine and tamoxifen. A broadly successful strategy for killing drug-resistant cancer cells, however, could be based on either selective energy depletion of cancer cells or the permeabilization of tumor cells with an effective bypass of efflux pumps, since many mechanisms of drug resistance entail the energy-dependent export of toxins. The latter approach may be achieved via membrane lipid modifications or the introduction of membrane pores by biological or physical (electroporation) means.

Positional cloning of zebrafish ferroportin1 identifies a conserved vertebrate iron exporter.

PubMed

Donovan, A; Brownlie, A; Zhou, Y; Shepard, J; Pratt, S J; Moynihan, J; Paw, B H; Drejer, A; Barut, B; Zapata, A; Law, T C; Brugnara, C; Lux, S E; Pinkus, G S; Pinkus, J L; Kingsley, P D; Palis, J; Fleming, M D; Andrews, N C; Zon, L I

2000-02-17

Defects in iron absorption and utilization lead to iron deficiency and overload disorders. Adult mammals absorb iron through the duodenum, whereas embryos obtain iron through placental transport. Iron uptake from the intestinal lumen through the apical surface of polarized duodenal enterocytes is mediated by the divalent metal transporter, DMTi. A second transporter has been postulated to export iron across the basolateral surface to the circulation. Here we have used positional cloning to identify the gene responsible for the hypochromic anaemia of the zebrafish mutant weissherbst. The gene, ferroportin1, encodes a multiple-transmembrane domain protein, expressed in the yolk sac, that is a candidate for the elusive iron exporter. Zebrafish ferroportin1 is required for the transport of iron from maternally derived yolk stores to the circulation and functions as an iron exporter when expressed in Xenopus oocytes. Human Ferroportin1 is found at the basal surface of placental syncytiotrophoblasts, suggesting that it also transports iron from mother to embryo. Mammalian Ferroportin1 is expressed at the basolateral surface of duodenal enterocytes and could export cellular iron into the circulation. We propose that Ferroportin1 function may be perturbed in mammalian disorders of iron deficiency or overload.

Expression and role of the genes involved in the transport of bile acids in the liver and kidneys in mice.

PubMed

Attakpa, Eugène S; Djibril, Naguibou M; Baba-Moussa, Farid; Yessoufou, Ganiou; Sezan, Alphonse

2013-01-01

Bile acids are synthesized in the liver from cholesterol. This study investigated the impact and expression of different carriers of bile acid in the liver and kidneys. Eight-week-old male mice were used, which were fed for 15 days and divided into two groups: 15 mice fed with standard diet (control group) and another 15 mice fed with a rich diet of 5% cholesterol (second group). Bile acid dosage was based on their oxidation by 7α hydroxyl-steroid dehydrogenize. The mRNA expression was quantitatively analyzed by the real time of polymerase chain reaction (RT-PCR), and the expression of the renal carrier bile acid protein was analyzed by Western blot. The expression of bile salt export pump involved in the uptake of bile acids in the basolateral membrane of hepatocytes revealed no differences between the two groups of mice. However, the expression of multidrug resistance-associated protein 2 was reduced in mice of the second group. Moreover, the expressions of organic anion transporting polypeptide 4, organic anion transporting polypeptide 1, and sodium taurocholate co-transporting polypeptide (Ntcp) involved in the uptake of bile acids in the apical pole of hepatocytes are suppressed in mice of the second group. The expression of multidrug resistance-associated protein 3 involved in the secretion of bile acids in the apical membrane of hepatocytes revealed no significant differences between the two groups. In mice of the second group, blood concentration of bile acids on the last day was increased. In those mice, the expression of intestinal bile acid transporter was reduced in the kidneys compared with the control mice.

Fluorescent recovery after photobleaching (FRAP) analysis of nuclear export rates identifies intrinsic features of nucleocytoplasmic transport.

PubMed

Cardarelli, Francesco; Tosti, Luca; Serresi, Michela; Beltram, Fabio; Bizzarri, Ranieri

2012-02-17

A quantitative description of carrier-mediated nuclear export in live cells is presented. To this end, we fused a prototypical leucine-rich nuclear export signal (NES) to GFP as a cargo model and expressed the fluorescent chimera in live CHO-K1 cells. By modeling FRAP data, we calculate the NES affinity for the export machinery and the maximum rate of nuclear export achievable at saturation of endogenous carriers. The measured active-export time through the Nuclear Pore Complex (NPC) is 18 ms, remarkably similar to the previously determined active-import rate. Also, our results reveal that active export/import and active export/passive diffusion fluxes are uncoupled, thus complementing previous reports on active import/passive diffusion uncoupling. These findings suggest differential gating at the NPC level.

10 CFR Appendix K to Part 110 - Illustrative List of Equipment and Components Under NRC Export Licensing Authority for Use in a...

Code of Federal Regulations, 2014 CFR

2014-01-01

... from synthesis gas through contact with liquid ammonia in the presence of a catalyst. The systhesis gas... contactors which promote intimate gas/liquid contact. Stage pumps include especially designed submersible... down the towers while the hydrogen sulphide gas circulates from the bottom to the top of the towers. A...

10 CFR Appendix K to Part 110 - Illustrative List of Equipment and Components Under NRC Export Licensing Authority for Use in a...

Code of Federal Regulations, 2012 CFR

2012-01-01

... from synthesis gas through contact with liquid ammonia in the presence of a catalyst. The systhesis gas... contactors which promote intimate gas/liquid contact. Stage pumps include especially designed submersible... down the towers while the hydrogen sulphide gas circulates from the bottom to the top of the towers. A...

10 CFR Appendix K to Part 110 - Illustrative List of Equipment and Components Under NRC Export Licensing Authority for Use in a...

Code of Federal Regulations, 2013 CFR

2013-01-01

... from synthesis gas through contact with liquid ammonia in the presence of a catalyst. The systhesis gas... contactors which promote intimate gas/liquid contact. Stage pumps include especially designed submersible... down the towers while the hydrogen sulphide gas circulates from the bottom to the top of the towers. A...

10 CFR Appendix K to Part 110 - Illustrative List of Equipment and Components Under NRC Export Licensing Authority for Use in a...

Code of Federal Regulations, 2011 CFR

2011-01-01

... from synthesis gas through contact with liquid ammonia in the presence of a catalyst. The systhesis gas... contactors which promote intimate gas/liquid contact. Stage pumps include especially designed submersible... down the towers while the hydrogen sulphide gas circulates from the bottom to the top of the towers. A...

10 CFR Appendix K to Part 110 - Illustrative List of Equipment and Components Under NRC Export Licensing Authority for Use in a...

Code of Federal Regulations, 2010 CFR

2010-01-01

... from synthesis gas through contact with liquid ammonia in the presence of a catalyst. The systhesis gas... contactors which promote intimate gas/liquid contact. Stage pumps include especially designed submersible... down the towers while the hydrogen sulphide gas circulates from the bottom to the top of the towers. A...

The role of diatom resting spores in pelagic-benthic coupling in the Southern Ocean

NASA Astrophysics Data System (ADS)

Rembauville, Mathieu; Blain, Stéphane; Manno, Clara; Tarling, Geraint; Thompson, Anu; Wolff, George; Salter, Ian

2018-05-01

Natural iron fertilization downstream of Southern Ocean island plateaus supports large phytoplankton blooms and promotes carbon export from the mixed layer. In addition to sequestering atmospheric CO2, the biological carbon pump also supplies organic matter (OM) to deep-ocean ecosystems. Although the total flux of OM arriving at the seafloor sets the energy input to the system, the chemical nature of OM is also of significance. However, a quantitative framework linking ecological flux vectors to OM composition is currently lacking. In the present study we report the lipid composition of export fluxes collected by five moored sediment traps deployed in contrasting productivity regimes of Southern Ocean island systems (Kerguelen, Crozet and South Georgia) and compile them with quantitative data on diatom and faecal pellet fluxes. At the three naturally iron-fertilized sites, the relative contribution of labile lipids (mono- and polyunsaturated fatty acids, unsaturated fatty alcohols) is 2-4 times higher than at low productivity sites. There is a strong attenuation of labile components as a function of depth, irrespective of productivity. The three island systems also display regional characteristics in lipid export. An enrichment of zooplankton dietary sterols, such as C27Δ5, at South Georgia is consistent with high zooplankton and krill biomass in the region and the importance of faecal pellets to particulate organic carbon (POC) flux. There is a strong association of diatom resting spore fluxes that dominate productive flux regimes with energy-rich unsaturated fatty acids. At the Kerguelen Plateau we provide a statistical framework to link seasonal variation in ecological flux vectors and lipid composition over a complete annual cycle. Our analyses demonstrate that ecological processes in the upper ocean, e.g. resting spore formation and grazing, not only impact the magnitude and stoichiometry of the Southern Ocean biological pump, but also regulate the composition of exported OM and the nature of pelagic-benthic coupling.

Influence of top-down control in the plankton food web on vertical carbon flux: a mesocosm study in the Chesapeake Bay

NASA Astrophysics Data System (ADS)

Stone, J.; Steinberg, D. K.

2016-02-01

The effects of predation on carbon export in planktonic food webs are poorly known, but likely play a key role in the biological pump. Gelatinous zooplankton (GZ) dominate the zooplankton community in the Chesapeake Bay during summer months, exerting considerable top-down control on the planktonic food web. The medusa Chrysaora quinquecirrha preys upon the ctenophore Mnemiopsis leidyi, which in turn is a major predator of the omnivorous copepod Acartia tonsa. This trophic cascade is known to significantly affect copepod abundance in Chesapeake Bay, but the resulting changes to particulate organic carbon (POC) flux are unknown. We hypothesized that additions or exclusions of GZ predators would result in changes in both total POC flux and the composition of exported particles (e.g., phytoplankton aggregates, fecal pellets). We conducted mesocosm experiments in the York River tributary of Chesapeake Bay during summer and fall, 2015 to quantify the cascading effects of GZ blooms on POC flux. The mesocosms contained a natural assemblage of phytoplankton and microzooplankton, and A. tonsa copepods, and received one of four treatments of GZ: 1) a control with no GZ added, 2) addition of ctenophores, 3) addition of medusae, and 4) addition of both ctenophores and medusae. POC flux from each mesocosm was measured over multiple 2-day experimental runs and grazing rates of GZ on each other and on copepods were calculated. There were no significant differences in total POC flux between treatments, but the composition of both the final zooplankton assemblage and exported organic matter differed between treatments. As a result of grazing on copepods by ctenophores, treatments which included GZ had lower final copepod abundances and a corresponding decrease in flux of copepod fecal pellets. We discuss how this change in composition of exported material as a result of cascading trophic interactions may affect the efficiency of the biological pump and benthic processes.

Efficient extracellular production of type I secretion pathway-dependent Pseudomonas fluorescens lipase in recombinant Escherichia coli by heterologous ABC protein exporters.

PubMed

Eom, Gyeong Tae; Lee, Seung Hwan; Oh, Young Hoon; Choi, Ji Eun; Park, Si Jae; Song, Jae Kwang

2014-10-01

Heterologous ABC protein exporters, the apparatus of type I secretion pathway in Gram-negative bacteria, were used for extracellular production of Pseudomonas fluorescens lipase (TliA) in recombinant Escherichia coli. The effect of the expression of different ABC protein exporter gene clusters (P. fluorescens tliDEF, Pseudomonas aeruginosa aprDEF, Erwinia chrysanthemi prtDEF, and Serratia marcescens lipBCD genes) was examined on the secretion of TliA at growth temperatures of 20, 25, 30 and 35 °C. TliA secretion in recombinant E. coli XL10-Gold varied depending upon type of ABC protein exporter and culture temperature. E. coli expressing S. marcescens lipBCD genes showed the highest secretion level of TliA (122.8 U ml(-1)) when cultured at 25 °C. Thus, optimized culture conditions for efficient extracellular production of lipase in recombinant E. coli can be designed by changing the type of ABC protein exporter and the growth temperature.

Safe intravenous administration in pediatrics: A 5-year Pediatric Intensive Care Unit experience with smart pumps.

PubMed

Manrique-Rodríguez, S; Sánchez-Galindo, A C; Fernández-Llamazares, C M; Calvo-Calvo, M M; Carrillo-Álvarez, Á; Sanjurjo-Sáez, M

2016-10-01

To estimate the impact of smart pump implementation in a pediatric intensive care unit in terms of number and type of administration errors intercepted. Observational, prospective study carried out from January 2010 to March 2015 with syringe and great volumen infusion pumps available in the hospital. A tertiary level hospital pediatric intensive care unit. Infusions delivered with infusion pumps in all pediatric intensive care unit patients. Design of a drug library with safety limits for all intravenous drugs prescribed. Users' compliance with drug library as well as number and type of errors prevented were analyzed. Two hundred and eighty-three errors were intercepted during 62 months of study. A high risk drug was involved in 58% of prevented errors, such as adrenergic agonists and antagonists, sedatives, analgesics, neuromuscular blockers, opioids, potassium and insulin. Users' average compliance with the safety software was 84%. Smart pumps implementation has proven effective in intercepting high risk drugs programming errors. These results might be exportable to other critical care units, involving pediatric or adult patients. Interdisciplinary colaboration is key to succeed in this process. Copyright © 2016 Elsevier España, S.L.U. y SEMICYUC. All rights reserved.

Biological and physical controls in the Southern Ocean on past millennial-scale atmospheric CO2 changes.

PubMed

Gottschalk, Julia; Skinner, Luke C; Lippold, Jörg; Vogel, Hendrik; Frank, Norbert; Jaccard, Samuel L; Waelbroeck, Claire

2016-05-17

Millennial-scale climate changes during the last glacial period and deglaciation were accompanied by rapid changes in atmospheric CO2 that remain unexplained. While the role of the Southern Ocean as a 'control valve' on ocean-atmosphere CO2 exchange has been emphasized, the exact nature of this role, in particular the relative contributions of physical (for example, ocean dynamics and air-sea gas exchange) versus biological processes (for example, export productivity), remains poorly constrained. Here we combine reconstructions of bottom-water [O2], export production and (14)C ventilation ages in the sub-Antarctic Atlantic, and show that atmospheric CO2 pulses during the last glacial- and deglacial periods were consistently accompanied by decreases in the biological export of carbon and increases in deep-ocean ventilation via southern-sourced water masses. These findings demonstrate how the Southern Ocean's 'organic carbon pump' has exerted a tight control on atmospheric CO2, and thus global climate, specifically via a synergy of both physical and biological processes.

Biological and physical controls in the Southern Ocean on past millennial-scale atmospheric CO2 changes

PubMed Central

Gottschalk, Julia; Skinner, Luke C.; Lippold, Jörg; Vogel, Hendrik; Frank, Norbert; Jaccard, Samuel L.; Waelbroeck, Claire

2016-01-01

Millennial-scale climate changes during the last glacial period and deglaciation were accompanied by rapid changes in atmospheric CO2 that remain unexplained. While the role of the Southern Ocean as a 'control valve' on ocean–atmosphere CO2 exchange has been emphasized, the exact nature of this role, in particular the relative contributions of physical (for example, ocean dynamics and air–sea gas exchange) versus biological processes (for example, export productivity), remains poorly constrained. Here we combine reconstructions of bottom-water [O2], export production and 14C ventilation ages in the sub-Antarctic Atlantic, and show that atmospheric CO2 pulses during the last glacial- and deglacial periods were consistently accompanied by decreases in the biological export of carbon and increases in deep-ocean ventilation via southern-sourced water masses. These findings demonstrate how the Southern Ocean's 'organic carbon pump' has exerted a tight control on atmospheric CO2, and thus global climate, specifically via a synergy of both physical and biological processes. PMID:27187527

Assessing mRNA nuclear export in mammalian cells by microinjection.

PubMed

Lee, Eliza S; Palazzo, Alexander F

2017-08-15

The nuclear export of mRNAs is an important yet little understood part of eukaryotic gene expression. One of the easiest methods for monitoring mRNA export in mammalian tissue culture cells is through the microinjection of DNA plasmids into the nucleus and monitoring the distribution of the transcribed product over time. Here we describe how to setup a microscope equipped with a micromanipulator used in cell microinjections, and we explain how to perform a nuclear mRNA export assay and obtain the nuclear export rate for any given mRNA. Copyright © 2017 Elsevier Inc. All rights reserved.

Nuclear export of RNA: Different sizes, shapes and functions.

PubMed

Williams, Tobias; Ngo, Linh H; Wickramasinghe, Vihandha O

2018-03-01

Export of protein-coding and non-coding RNA molecules from the nucleus to the cytoplasm is critical for gene expression. This necessitates the continuous transport of RNA species of different size, shape and function through nuclear pore complexes via export receptors and adaptor proteins. Here, we provide an overview of the major RNA export pathways in humans, highlighting the similarities and differences between each. Its importance is underscored by the growing appreciation that deregulation of RNA export pathways is associated with human diseases like cancer. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

Fluorescent Recovery after Photobleaching (FRAP) Analysis of Nuclear Export Rates Identifies Intrinsic Features of Nucleocytoplasmic Transport*

PubMed Central

Cardarelli, Francesco; Tosti, Luca; Serresi, Michela; Beltram, Fabio; Bizzarri, Ranieri

2012-01-01

A quantitative description of carrier-mediated nuclear export in live cells is presented. To this end, we fused a prototypical leucine-rich nuclear export signal (NES) to GFP as a cargo model and expressed the fluorescent chimera in live CHO-K1 cells. By modeling FRAP data, we calculate the NES affinity for the export machinery and the maximum rate of nuclear export achievable at saturation of endogenous carriers. The measured active-export time through the Nuclear Pore Complex (NPC) is 18 ms, remarkably similar to the previously determined active-import rate. Also, our results reveal that active export/import and active export/passive diffusion fluxes are uncoupled, thus complementing previous reports on active import/passive diffusion uncoupling. These findings suggest differential gating at the NPC level. PMID:22190681

RRE-dependent HIV-1 Env RNA effects on Gag protein expression, assembly and release

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

López, Claudia S., E-mail: lopezcl@ohsu.edu; Sloan, Rachel; Cylinder, Isabel

The HIV-1 Gag proteins are translated from the full-length HIV-1 viral RNA (vRNA), whereas the envelope (Env) protein is translated from incompletely spliced Env mRNAs. Nuclear export of vRNAs and Env mRNAs is mediated by the Rev accessory protein which binds to the rev-responsive element (RRE) present on these RNAs. Evidence has shown there is a direct or indirect interaction between the Gag protein, and the cytoplasmic tail (CT) of the Env protein. Our current work shows that env gene expression impacts HIV-1 Gag expression and function in two ways. At the protein level, full-length Env expression altered Gag proteinmore » expression, while Env CT-deletion proteins did not. At the RNA level, RRE-containing Env mRNA expression reduced Gag expression, processing, and virus particle release from cells. Our results support models in which Gag is influenced by the Env CT, and Env mRNAs compete with vRNAs for nuclear export. - Highlights: • At the protein level, full-length HIV-1 Env alters Gag protein expression. • HIV-1 Env RNA expression reduces Gag levels and virus release. • Env RNA effects on Gag are dependent on the RRE. • RRE-containing Env RNAs compete with vRNAs for nuclear export.« less

Natural ocean carbon cycle sensitivity to parameterizations of the recycling in a climate model

NASA Astrophysics Data System (ADS)

Romanou, A.; Romanski, J.; Gregg, W. W.

2014-02-01

Sensitivities of the oceanic biological pump within the GISS (Goddard Institute for Space Studies ) climate modeling system are explored here. Results are presented from twin control simulations of the air-sea CO2 gas exchange using two different ocean models coupled to the same atmosphere. The two ocean models (Russell ocean model and Hybrid Coordinate Ocean Model, HYCOM) use different vertical coordinate systems, and therefore different representations of column physics. Both variants of the GISS climate model are coupled to the same ocean biogeochemistry module (the NASA Ocean Biogeochemistry Model, NOBM), which computes prognostic distributions for biotic and abiotic fields that influence the air-sea flux of CO2 and the deep ocean carbon transport and storage. In particular, the model differences due to remineralization rate changes are compared to differences attributed to physical processes modeled differently in the two ocean models such as ventilation, mixing, eddy stirring and vertical advection. GISSEH(GISSER) is found to underestimate mixed layer depth compared to observations by about 55% (10%) in the Southern Ocean and overestimate it by about 17% (underestimate by 2%) in the northern high latitudes. Everywhere else in the global ocean, the two models underestimate the surface mixing by about 12-34%, which prevents deep nutrients from reaching the surface and promoting primary production there. Consequently, carbon export is reduced because of reduced production at the surface. Furthermore, carbon export is particularly sensitive to remineralization rate changes in the frontal regions of the subtropical gyres and at the Equator and this sensitivity in the model is much higher than the sensitivity to physical processes such as vertical mixing, vertical advection and mesoscale eddy transport. At depth, GISSER, which has a significant warm bias, remineralizes nutrients and carbon faster thereby producing more nutrients and carbon at depth, which eventually resurfaces with the global thermohaline circulation especially in the Southern Ocean. Because of the reduced primary production and carbon export in GISSEH compared to GISSER, the biological pump efficiency, i.e., the ratio of primary production and carbon export at 75 m, is half in the GISSEH of that in GISSER, The Southern Ocean emerges as a key region where the CO2 flux is as sensitive to biological parameterizations as it is to physical parameterizations. The fidelity of ocean mixing in the Southern Ocean compared to observations is shown to be a good indicator of the magnitude of the biological pump efficiency regardless of physical model choice.

Natural Ocean Carbon Cycle Sensitivity to Parameterizations of the Recycling in a Climate Model

NASA Technical Reports Server (NTRS)

Romanou, A.; Romanski, J.; Gregg, W. W.

2014-01-01

Sensitivities of the oceanic biological pump within the GISS (Goddard Institute for Space Studies ) climate modeling system are explored here. Results are presented from twin control simulations of the air-sea CO2 gas exchange using two different ocean models coupled to the same atmosphere. The two ocean models (Russell ocean model and Hybrid Coordinate Ocean Model, HYCOM) use different vertical coordinate systems, and therefore different representations of column physics. Both variants of the GISS climate model are coupled to the same ocean biogeochemistry module (the NASA Ocean Biogeochemistry Model, NOBM), which computes prognostic distributions for biotic and abiotic fields that influence the air-sea flux of CO2 and the deep ocean carbon transport and storage. In particular, the model differences due to remineralization rate changes are compared to differences attributed to physical processes modeled differently in the two ocean models such as ventilation, mixing, eddy stirring and vertical advection. GISSEH(GISSER) is found to underestimate mixed layer depth compared to observations by about 55% (10 %) in the Southern Ocean and overestimate it by about 17% (underestimate by 2%) in the northern high latitudes. Everywhere else in the global ocean, the two models underestimate the surface mixing by about 12-34 %, which prevents deep nutrients from reaching the surface and promoting primary production there. Consequently, carbon export is reduced because of reduced production at the surface. Furthermore, carbon export is particularly sensitive to remineralization rate changes in the frontal regions of the subtropical gyres and at the Equator and this sensitivity in the model is much higher than the sensitivity to physical processes such as vertical mixing, vertical advection and mesoscale eddy transport. At depth, GISSER, which has a significant warm bias, remineralizes nutrients and carbon faster thereby producing more nutrients and carbon at depth, which eventually resurfaces with the global thermohaline circulation especially in the Southern Ocean. Because of the reduced primary production and carbon export in GISSEH compared to GISSER, the biological pump efficiency, i.e., the ratio of primary production and carbon export at 75 m, is half in the GISSEH of that in GISSER, The Southern Ocean emerges as a key region where the CO2 flux is as sensitive to biological parameterizations as it is to physical parameterizations. The fidelity of ocean mixing in the Southern Ocean compared to observations is shown to be a good indicator of the magnitude of the biological pump efficiency regardless of physical model choice.

Decreased calcium pump expression in human erythrocytes is connected to a minor haplotype in the ATP2B4 gene.

PubMed

Zámbó, Boglárka; Várady, György; Padányi, Rita; Szabó, Edit; Németh, Adrienn; Langó, Tamás; Enyedi, Ágnes; Sarkadi, Balázs

2017-07-01

Plasma membrane Ca 2+ -ATPases are key calcium exporter proteins in most tissues, and PMCA4b is the main calcium transporter in the human red blood cells (RBCs). In order to assess the expression level of PMCA4b, we have developed a flow cytometry and specific antibody binding method to quantitatively detect this protein in the erythrocyte membrane. Interestingly, we found several healthy volunteers showing significantly reduced expression of RBC-PMCA4b. Western blot analysis of isolated RBC membranes confirmed this observation, and indicated that there are no compensatory alterations in other PMCA isoforms. In addition, reduced PMCA4b levels correlated with a lower calcium extrusion capacity in these erythrocytes. When exploring the potential genetic background of the reduced PMCA4b levels, we found no missense mutations in the ATP2B4 coding regions, while a formerly unrecognized minor haplotype in the predicted second promoter region closely correlated with lower erythrocyte PMCA4b protein levels. In recent GWA studies, SNPs in this ATP2B4 haplotype have been linked to reduced mean corpuscular hemoglobin concentrations (MCHC), and to protection against malaria infection. Our data suggest that an altered regulation of gene expression is responsible for the reduced RBC-PMCA4b levels that is probably linked to the development of human disease-related phenotypes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Analytical thermal model for end-pumped solid-state lasers

NASA Astrophysics Data System (ADS)

Cini, L.; Mackenzie, J. I.

2017-12-01

Fundamentally power-limited by thermal effects, the design challenge for end-pumped "bulk" solid-state lasers depends upon knowledge of the temperature gradients within the gain medium. We have developed analytical expressions that can be used to model the temperature distribution and thermal-lens power in end-pumped solid-state lasers. Enabled by the inclusion of a temperature-dependent thermal conductivity, applicable from cryogenic to elevated temperatures, typical pumping distributions are explored and the results compared with accepted models. Key insights are gained through these analytical expressions, such as the dependence of the peak temperature rise in function of the boundary thermal conductance to the heat sink. Our generalized expressions provide simple and time-efficient tools for parametric optimization of the heat distribution in the gain medium based upon the material and pumping constraints.

Gene expression of H+-pumps in plasma and vacuolar membranes of corn root cells under the effect of sodium ions and bioactive preparations.

PubMed

Kovalenko, N O; Palladina, T A

2016-01-01

Four isoforms of H+-ATPase of plasma membrane: MHA1, MHA2, MHA3, MHA4 are expressed in the corn seedling roots with prevalence of genes MHA3 і MHA4. The exposure of seedlings in the presence of 0.1 M NaCl activated the expression of MHA4 gene isoform, that demonstrates its important role in the processes of adaptation to salinization conditions. In vacuolar membrane, where potential is created by two Н+-pumps, sodium ions activated gene expression of only Н+-АТРase of V-type, taking no effect on the expression of Н+-pyrophosphatase. The seeds pretreatment by synthetic preparations Methyure and Ivine did not affect gene expression of Н+-pumps. Thus we can suppose that the ability of the above preparations to activate functioning of Н+-pumps in the presence of sodium ions is realized at the post-tranlation level.

Iron Export through the Transporter Ferroportin 1 Is Modulated by the Iron Chaperone PCBP2*

PubMed Central

Yanatori, Izumi; Richardson, Des R.; Imada, Kiyoshi; Kishi, Fumio

2016-01-01

Ferroportin 1 (FPN1) is an iron export protein found in mammals. FPN1 is important for the export of iron across the basolateral membrane of absorptive enterocytes and across the plasma membrane of macrophages. The expression of FPN1 is regulated by hepcidin, which binds to FPN1 and then induces its degradation. Previously, we demonstrated that divalent metal transporter 1 (DMT1) interacts with the intracellular iron chaperone protein poly(rC)-binding protein 2 (PCBP2). Subsequently, PCBP2 receives iron from DMT1 and then disengages from the transporter. In this study, we investigated the function of PCBP2 in iron export. Mammalian genomes encode four PCBPs (i.e. PCBP1–4). Here, for the first time, we demonstrated using both yeast and mammalian cells that PCBP2, but not PCBP1, PCBP3, or PCBP4, binds with FPN1. Importantly, iron-loaded, but not iron-depleted, PCBP2 interacts with FPN1. The PCBP2-binding domain of FPN1 was identified in its C-terminal cytoplasmic region. The silencing of PCBP2 expression suppressed FPN1-dependent iron export from cells. These results suggest that FPN1 exports iron received from the iron chaperone PCBP2. Therefore, it was found that PCBP2 modulates cellular iron export, which is an important physiological process. PMID:27302059

Amino acid stable isotope applications to deep-sea corals: A molecular geochemistry approach to reconstructing past ocean conditions

NASA Astrophysics Data System (ADS)

McMahon, K.; McCarthy, M. D.; Guilderson, T. P.; Sherwood, O.; Williams, B.; Larsen, T.; Glynn, D. S.

2017-12-01

Future climate change is predicted to alter ocean productivity, food web dynamics, biogeochemical cycling, and the efficacy of the biological pump. Proteinaceous deep-sea corals act as "living sediment traps," providing long-term, high-resolution records of exported surface ocean production and a window into past changes in ocean condition as a historical context for potential future changes. Here, we present recent work developing the application of compound-specific stable isotope analysis of individual amino acids to proteinaceous deep-sea corals to reconstruct past changes in phytoplankton community composition and biogeochemical cycling. We present new calibrations for molecular isotope comparisons between metabolically active coral polyp tissue and bioarchival proteinaceous skeleton. We then applied these techniques to deep-sea corals from the North Pacific Subtropical Gyre (NPSG) to reconstruct centennial to millennial time scale changes in phytoplankton community composition and biogeochemical cycling as a function of regional climate change. This work suggests that the NPSG has undergone multiple major phytoplankton regime shifts over the last millennium between prokaryotic and eukaryotic phytoplankton communities and associated sources of nitrogen fueling production. The most recent regime, which started around the end of the Little Ice Age and the onset of the Industrial era, is unprecedented in the last 1000 years and resulted in a 30-50% increase in diazotrophic cyanobacteria contribution to export production and an associated 17-27% increase in N2-fixation in the NPSG over last century. By offering the first direct phylogenetic context for long-term shifts in isotopic records of exported particulate organic matter, our data represent a major step forward in understanding the evolution of marine plankton community dynamics, food web architecture, biogeochemical cycling, and the climate feedback loops through the biological pump.

The Southern Ocean biogeochemical divide.

PubMed

Marinov, I; Gnanadesikan, A; Toggweiler, J R; Sarmiento, J L

2006-06-22

Modelling studies have demonstrated that the nutrient and carbon cycles in the Southern Ocean play a central role in setting the air-sea balance of CO(2) and global biological production. Box model studies first pointed out that an increase in nutrient utilization in the high latitudes results in a strong decrease in the atmospheric carbon dioxide partial pressure (pCO2). This early research led to two important ideas: high latitude regions are more important in determining atmospheric pCO2 than low latitudes, despite their much smaller area, and nutrient utilization and atmospheric pCO2 are tightly linked. Subsequent general circulation model simulations show that the Southern Ocean is the most important high latitude region in controlling pre-industrial atmospheric CO(2) because it serves as a lid to a larger volume of the deep ocean. Other studies point out the crucial role of the Southern Ocean in the uptake and storage of anthropogenic carbon dioxide and in controlling global biological production. Here we probe the system to determine whether certain regions of the Southern Ocean are more critical than others for air-sea CO(2) balance and the biological export production, by increasing surface nutrient drawdown in an ocean general circulation model. We demonstrate that atmospheric CO(2) and global biological export production are controlled by different regions of the Southern Ocean. The air-sea balance of carbon dioxide is controlled mainly by the biological pump and circulation in the Antarctic deep-water formation region, whereas global export production is controlled mainly by the biological pump and circulation in the Subantarctic intermediate and mode water formation region. The existence of this biogeochemical divide separating the Antarctic from the Subantarctic suggests that it may be possible for climate change or human intervention to modify one of these without greatly altering the other.

Coarse-grained Simulations of Substrate Export through Multidrug Efflux Transporter AcrB

NASA Astrophysics Data System (ADS)

Jewel, Yead; Dutta, Prashanta; Liu, Jin

2017-11-01

The treatment of bacterial infectious diseases hampered by the overexpression of multidrug resistance (MDR) systems. The MDR system actively pumps the antibiotic drugs as well as other toxic compounds out of the cells. During the pumping, AcrB (one of the key MDR components) undergoes a series of large-scale proton/substrate dependent conformational changes. In this work, we implement a hybrid coarse-grained PACE force field that couples the united-atom protein model with the coarse-grained MARTINI water/lipid, to investigate the conformational changes of AcrB. We first develop the substrate force field which is compatible with PACE, then we implement the force field to explore large scale structural changes of AcrB in microsecond simulations. The effects of the substrate and the protonation states of two key residues: Asp407 and Asp408, are investigated. Our results show that the drug export through AcrB is proton as well as substrate dependent. Our simulations explain molecular mechanisms of substrate transport through AcrB complex, as well as provide valuable insights for designing proper antibiotic drugs. Research reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number R01GM122081.

A Pseudomonas putida efflux pump acts on short-chain alcohols.

PubMed

Basler, Georg; Thompson, Mitchell; Tullman-Ercek, Danielle; Keasling, Jay

2018-01-01

The microbial production of biofuels is complicated by a tradeoff between yield and toxicity of many fuels. Efflux pumps enable bacteria to tolerate toxic substances by their removal from the cells while bypassing the periplasm. Their use for the microbial production of biofuels can help to improve cell survival, product recovery, and productivity. However, no native efflux pump is known to act on the class of short-chain alcohols, important next-generation biofuels, and it was considered unlikely that such an efflux pump exists. We report that controlled expression of the RND-type efflux pump TtgABC from Pseudomonas putida DOT-T1E strongly improved cell survival in highly toxic levels of the next-generation biofuels n -butanol, isobutanol, isoprenol, and isopentanol. GC-FID measurements indicated active efflux of n -butanol when the pump is expressed. Conversely, pump expression did not lead to faster growth in media supplemented with low concentrations of n -butanol and isopentanol. TtgABC is the first native efflux pump shown to act on multiple short-chain alcohols. Its controlled expression can be used to improve cell survival and increase production of biofuels as an orthogonal approach to metabolic engineering. Together with the increased interest in P. putida for metabolic engineering due to its flexible metabolism, high native tolerance to toxic substances, and various applications of engineering its metabolism, our findings endorse the strain as an excellent biocatalyst for the high-yield production of next-generation biofuels.

Transporter Expression in Liver Tissue from Subjects with Alcoholic or Hepatitis C Cirrhosis Quantified by Targeted Quantitative Proteomics

PubMed Central

Wang, Li; Collins, Carol; Kelly, Edward J.; Chu, Xiaoyan; Ray, Adrian S.; Salphati, Laurent; Xiao, Guangqing; Lee, Caroline; Lai, Yurong; Liao, Mingxiang; Mathias, Anita; Evers, Raymond; Humphreys, William; Hop, Cornelis E. C. A.; Kumer, Sean C.

2016-01-01

Although data are available on the change of expression/activity of drug-metabolizing enzymes in liver cirrhosis patients, corresponding data on transporter protein expression are not available. Therefore, using quantitative targeted proteomics, we compared our previous data on noncirrhotic control livers (n = 36) with the protein expression of major hepatobiliary transporters, breast cancer resistance protein (BCRP), bile salt export pump (BSEP), multidrug and toxin extrusion protein 1 (MATE1), multidrug resistance–associated protein (MRP)2, MRP3, MRP4, sodium taurocholate–cotransporting polypeptide (NTCP), organic anion–transporting polypeptides (OATP)1B1, 1B3, 2B1, organic cation transporter 1 (OCT1), and P-glycoprotein (P-gp) in alcoholic (n = 27) and hepatitis C cirrhosis (n = 30) livers. Compared with control livers, the yield of membrane protein from alcoholic and hepatitis C cirrhosis livers was significantly reduced by 56 and 67%, respectively. The impact of liver cirrhosis on transporter protein expression was transporter-dependent. Generally, reduced protein expression (per gram of liver) was found in alcoholic cirrhosis livers versus control livers, with the exception that the expression of MRP3 was increased, whereas no change was observed for MATE1, MRP2, OATP2B1, and P-gp. In contrast, the impact of hepatitis C cirrhosis on protein expression of transporters (per gram of liver) was diverse, showing an increase (MATE1), decrease (BSEP, MRP2, NTCP, OATP1B3, OCT1, and P-gp), or no change (BCRP, MRP3, OATP1B1, and 2B1). The expression of hepatobiliary transporter protein differed in different diseases (alcoholic versus hepatitis C cirrhosis). Finally, incorporation of protein expression of OATP1B1 in alcoholic cirrhosis into the Simcyp physiologically based pharmacokinetics cirrhosis module improved prediction of the disposition of repaglinide in liver cirrhosis patients. These transporter expression data will be useful in the future to predict transporter-mediated drug disposition in liver cirrhosis patients. PMID:27543206

Cooperativity among Rev-Associated Nuclear Export Signals Regulates HIV-1 Gene Expression and Is a Determinant of Virus Species Tropism

PubMed Central

Aligeti, Mounavya; Behrens, Ryan T.; Pocock, Ginger M.; Schindelin, Johannes; Dietz, Christian; Eliceiri, Kevin W.; Swanson, Chad M.; Malim, Michael H.; Ahlquist, Paul

2014-01-01

ABSTRACT Murine cells exhibit a profound block to HIV-1 virion production that was recently mapped to a species-specific structural attribute of the murine version of the chromosomal region maintenance 1 (mCRM1) nuclear export receptor and rescued by the expression of human CRM1 (hCRM1). In human cells, the HIV-1 Rev protein recruits hCRM1 to intron-containing viral mRNAs encoding the Rev response element (RRE), thereby facilitating viral late gene expression. Here we exploited murine 3T3 fibroblasts as a gain-of-function system to study hCRM1's species-specific role in regulating Rev's effector functions. We show that Rev is rapidly exported from the nucleus by mCRM1 despite only weak contributions to HIV-1's posttranscriptional stages. Indeed, Rev preferentially accumulates in the cytoplasm of murine 3T3 cells with or without hCRM1 expression, in contrast to human HeLa cells, where Rev exhibits striking en masse transitions between the nuclear and cytoplasmic compartments. Efforts to bias Rev's trafficking either into or out of the nucleus revealed that Rev encoding a second CRM1 binding domain (Rev-2xNES) or Rev-dependent viral gag-pol mRNAs bearing tandem RREs (GP-2xRRE), rescue virus particle production in murine cells even in the absence of hCRM1. Combined, these results suggest a model wherein Rev-associated nuclear export signals cooperate to regulate the number or quality of CRM1's interactions with viral Rev/RRE ribonucleoprotein complexes in the nucleus. This mechanism regulates CRM1-dependent viral gene expression and is a determinant of HIV-1's capacity to produce virions in nonhuman cell types. IMPORTANCE Cells derived from mice and other nonhuman species exhibit profound blocks to HIV-1 replication. Here we elucidate a block to HIV-1 gene expression attributable to the murine version of the CRM1 (mCRM1) nuclear export receptor. In human cells, hCRM1 regulates the nuclear export of viral intron-containing mRNAs through the activity of the viral Rev adapter protein that forms a multimeric complex on these mRNAs prior to recruiting hCRM1. We demonstrate that Rev-dependent gene expression is poor in murine cells despite the finding that, surprisingly, the bulk of Rev interacts efficiently with mCRM1 and is rapidly exported from the nucleus. Instead, we map the mCRM1 defect to the apparent inability of this factor to engage Rev multimers in the context of large viral Rev/RNA ribonucleoprotein complexes. These findings shed new light on HIV-1 gene regulation and could inform the development of novel antiviral strategies that target viral gene expression. PMID:25275125

pH regulates genes for flagellar motility, catabolism, and oxidative stress in Escherichia coli K-12.

PubMed

Maurer, Lisa M; Yohannes, Elizabeth; Bondurant, Sandra S; Radmacher, Michael; Slonczewski, Joan L

2005-01-01

Gene expression profiles of Escherichia coli K-12 W3110 were compared as a function of steady-state external pH. Cultures were grown to an optical density at 600 nm of 0.3 in potassium-modified Luria-Bertani medium buffered at pH 5.0, 7.0, and 8.7. For each of the three pH conditions, cDNA from RNA of five independent cultures was hybridized to Affymetrix E. coli arrays. Analysis of variance with an alpha level of 0.001 resulted in 98% power to detect genes showing a twofold difference in expression. Normalized expression indices were calculated for each gene and intergenic region (IG). Differential expression among the three pH classes was observed for 763 genes and 353 IGs. Hierarchical clustering yielded six well-defined clusters of pH profiles, designated Acid High (highest expression at pH 5.0), Acid Low (lowest expression at pH 5.0), Base High (highest at pH 8.7), Base Low (lowest at pH 8.7), Neutral High (highest at pH 7.0, lower in acid or base), and Neutral Low (lowest at pH 7.0, higher at both pH extremes). Flagellar and chemotaxis genes were repressed at pH 8.7 (Base Low cluster), where the cell's transmembrane proton potential is diminished by the maintenance of an inverted pH gradient. High pH also repressed the proton pumps cytochrome o (cyo) and NADH dehydrogenases I and II. By contrast, the proton-importing ATP synthase F1Fo and the microaerophilic cytochrome d (cyd), which minimizes proton export, were induced at pH 8.7. These observations are consistent with a model in which high pH represses synthesis of flagella, which expend proton motive force, while stepping up electron transport and ATPase components that keep protons inside the cell. Acid-induced genes, on the other hand, were coinduced by conditions associated with increased metabolic rate, such as oxidative stress. All six pH-dependent clusters included envelope and periplasmic proteins, which directly experience external pH. Overall, this study showed that (i) low pH accelerates acid consumption and proton export, while coinducing oxidative stress and heat shock regulons; (ii) high pH accelerates proton import, while repressing the energy-expensive flagellar and chemotaxis regulons; and (iii) pH differentially regulates a large number of periplasmic and envelope proteins.

Electricity for groundwater use: constraints and opportunities for adaptive response to climate change

NASA Astrophysics Data System (ADS)

Scott, Christopher A.

2013-09-01

Globally, groundwater use is intensifying to meet demands for irrigation, urban supply, industrialization, and, in some instances, electrical power generation. In response to hydroclimatic variability, surface water is being substituted with groundwater, which must be viewed as a strategic resource for climate adaptation. In this sense, the supply of electricity for pumping is an adaptation policy tool. Additionally, planning for climate-change mitigation must consider CO2 emissions resulting from pumping. This paper examines the influence of electricity supply and pricing on groundwater irrigation and resulting emissions, with specific reference to Mexico—a climate-water-energy ‘perfect storm’. Night-time power supply at tariffs below the already-subsidized rates for agricultural groundwater use has caused Mexican farmers to increase pumping, reversing important water and electricity conservation gains achieved. Indiscriminate groundwater pumping, including for virtual water exports of agricultural produce, threatens the long-term sustainability of aquifers, non-agricultural water uses, and stream-aquifer interactions that sustain riparian ecosystems. Emissions resulting from agricultural groundwater pumping in Mexico are estimated to be 3.6% of total national emissions and are equivalent to emissions from transporting the same agricultural produce to market. The paper concludes with an assessment of energy, water, and climate trends coupled with policy futures to address these challenges.

Biological production in the Indian Ocean upwelling zones - Part 1: refined estimation via the use of a variable compensation depth in ocean carbon models

NASA Astrophysics Data System (ADS)

Geethalekshmi Sreeush, Mohanan; Valsala, Vinu; Pentakota, Sreenivas; Venkata Siva Rama Prasad, Koneru; Murtugudde, Raghu

2018-04-01

Biological modelling approach adopted by the Ocean Carbon-Cycle Model Intercomparison Project (OCMIP-II) provided amazingly simple but surprisingly accurate rendition of the annual mean carbon cycle for the global ocean. Nonetheless, OCMIP models are known to have seasonal biases which are typically attributed to their bulk parameterisation of compensation depth. Utilising the criteria of surface Chl a-based attenuation of solar radiation and the minimum solar radiation required for production, we have proposed a new parameterisation for a spatially and temporally varying compensation depth which captures the seasonality in the production zone reasonably well. This new parameterisation is shown to improve the seasonality of CO2 fluxes, surface ocean pCO2, biological export and new production in the major upwelling zones of the Indian Ocean. The seasonally varying compensation depth enriches the nutrient concentration in the upper ocean yielding more faithful biological exports which in turn leads to accurate seasonality in the carbon cycle. The export production strengthens by ˜ 70 % over the western Arabian Sea during the monsoon period and achieves a good balance between export and new production in the model. This underscores the importance of having a seasonal balance in the model export and new productions for a better representation of the seasonality of the carbon cycle over upwelling regions. The study also implies that both the biological and solubility pumps play an important role in the Indian Ocean upwelling zones.

Effects of L-arabinose efflux on λ Red recombination-mediated gene knockout in multiple-antimicrobial-resistant Salmonella enterica serovar Choleraesuis.

PubMed

Liao, Shi-Wei; Lee, Jen-Jie; Ptak, Christopher P; Wu, Ying-Chen; Hsuan, Shih-Ling; Kuo, Chih-Jung; Chen, Ter-Hsin

2018-03-01

In this study, six swine-derived multiple-antimicrobial-resistant (MAR) strains of Salmonella Choleraesuis (S. Choleraesuis) were demonstrated to possess higher efflux pump activity than the wild-type (WT). L-Arabinose, a common inducer for gene expression, modulated S. Choleraesuis efflux pump activity in a dose-dependent manner. At low L-arabinose concentrations, increasing L-arabinose led to a corresponding increase in fluorophore efflux, while at higher L-arabinose concentrations, increasing L-arabinose decreased fluorophore efflux activity. The WT S. Choleraesuis that lacks TolC (ΔtolC), an efflux protein associated with bacterial antibiotic resistance and virulence, was demonstrated to possess a significantly reduced ability to extrude L-arabinose. Further, due to the rapid export of L-arabinose, an efficient method for recombination-mediated gene knockout, the L-arabinose-inducible bacteriophage λ Red recombinase system, has a reduced recombination frequency (~ 12.5%) in clinically isolated MAR Salmonella strains. An increased recombination frequency (up to 60%) can be achieved using a higher concentration of L-arabinose (fivefold) for genetic manipulation and functional analysis for MAR Salmonella using the λ Red system. The study suggests that L-arabinose serves not only as an inducer of the TolC-dependent efflux system but also acts as a competitive substrate of the efflux system. In addition, understanding the TolC-dependent efflux of L-arabinose should facilitate the optimization of L-arabinose induction in strains with high efflux activity.

NMD3 regulates both mRNA and rRNA nuclear export in African trypanosomes via an XPOI-linked pathway

PubMed Central

Bühlmann, Melanie; Walrad, Pegine; Rico, Eva; Ivens, Alasdair; Capewell, Paul; Naguleswaran, Arunasalam; Roditi, Isabel; Matthews, Keith R.

2015-01-01

Trypanosomes mostly regulate gene expression through post-transcriptional mechanisms, particularly mRNA stability. However, much mRNA degradation is cytoplasmic such that mRNA nuclear export must represent an important level of regulation. Ribosomal RNAs must also be exported from the nucleus and the trypanosome orthologue of NMD3 has been confirmed to be involved in rRNA processing and export, matching its function in other organisms. Surprisingly, we found that TbNMD3 depletion also generates mRNA accumulation of procyclin-associated genes (PAGs), these being co-transcribed by RNA polymerase I with the procyclin surface antigen genes expressed on trypanosome insect forms. By whole transcriptome RNA-seq analysis of TbNMD3-depleted cells we confirm the regulation of the PAG transcripts by TbNMD3 and using reporter constructs reveal that PAG1 regulation is mediated by its 5′UTR. Dissection of the mechanism of regulation demonstrates that it is not dependent upon translational inhibition mediated by TbNMD3 depletion nor enhanced transcription. However, depletion of the nuclear export factors XPO1 or MEX67 recapitulates the effects of TbNMD3 depletion on PAG mRNAs and mRNAs accumulated in the nucleus of TbNMD3-depleted cells. These results invoke a novel RNA regulatory mechanism involving the NMD3-dependent nuclear export of mRNA cargos, suggesting a shared platform for mRNA and rRNA export. PMID:25873624

Tdp studies and tests for C. A. Energia Electrica de Venezuela (enelven) at planta ramon laguna, units RL-17 and RL-10. Volume 3. Unit RL-10 turbine generator condition assessment report and units RL-10 and RL-11 boiler feed pump conditon assessment report. Export trade information

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

Not Available

1991-03-28

The study, conducted by Babcock and Wilcox, was funded by the U.S. Trade and Development agency on behalf of Enelven. In order to maximize generated power output and minimize operating costs at Planta Ramon Laguna, tests were done to evaluate the condition of equipment at the plant. In order to identify any damage and determine the operating output of each unit, assessments were done of the furnaces, boilers, generators and boiler feed pumps being used in the plant. The report presents the results of these tests. This is the last of three volumes and it is divided into the followingmore » sections: (1) Condition Assessment of Unit RL-10 Turbine-Generator; (2) Condition Assessment of Unit RL-10 and RL-11 Boiler Feed Pumps.« less

Scanning electrochemical microscopy of menadione-glutathione conjugate export from yeast cells

PubMed Central

Mauzeroll, Janine; Bard, Allen J.

2004-01-01

The uptake of menadione (2-methyl-1,4-naphthoquinone), which is toxic to yeast cells, and its expulsion as a glutathione complex were studied by scanning electrochemical microscopy. The progression of the in vitro reaction between menadione and glutathione was monitored electrochemically by cyclic voltammetry and correlated with the spectroscopic (UV–visible) behavior. By observing the scanning electrochemical microscope tip current of yeast cells suspended in a menadione-containing solution, the export of the conjugate from the cells with time could be measured. Similar experiments were performed on immobilized yeast cell aggregates stressed by a menadione solution. From the export of the menadione-glutathione conjugate detected at a 1-μm-diameter electrode situated 10 μm from the cells, a flux of about 30,000 thiodione molecules per second per cell was extracted. Numerical simulations based on an explicit finite difference method further revealed that the observation of a constant efflux of thiodione from the cells suggested the rate was limited by the uptake of menadione and that the efflux through the glutathione-conjugate pump was at least an order of magnitude faster. PMID:15148374

Short communication: Massive erosion in monsoonal central India linked to late Holocene land cover degradation

NASA Astrophysics Data System (ADS)

Giosan, Liviu; Ponton, Camilo; Usman, Muhammed; Blusztajn, Jerzy; Fuller, Dorian Q.; Galy, Valier; Haghipour, Negar; Johnson, Joel E.; McIntyre, Cameron; Wacker, Lukas; Eglinton, Timothy I.

2017-12-01

Soil erosion plays a crucial role in transferring sediment and carbon from land to sea, yet little is known about the rhythm and rates of soil erosion prior to the most recent few centuries. Here we reconstruct a Holocene erosional history from central India, as integrated by the Godavari River in a sediment core from the Bay of Bengal. We quantify terrigenous fluxes, fingerprint sources for the lithogenic fraction and assess the age of the exported terrigenous carbon. Taken together, our data show that the monsoon decline in the late Holocene significantly increased soil erosion and the age of exported organic carbon. This acceleration of natural erosion was later exacerbated by the Neolithic adoption and Iron Age extensification of agriculture on the Deccan Plateau. Despite a constantly elevated sea level since the middle Holocene, this erosion acceleration led to a rapid growth of the continental margin. We conclude that in monsoon conditions aridity boosts rather than suppresses sediment and carbon export, acting as a monsoon erosional pump modulated by land cover conditions.

Nuclear Export and Expression of Human T-Cell Leukemia Virus Type 1 tax/rex mRNA Are RxRE/Rex Dependent

PubMed Central

Bai, X. T.; Sinha-Datta, U.; Ko, N. L.; Bellon, M.

2012-01-01

Human T-cell leukemia virus type 1 (HTLV-1) is a complex retrovirus associated with the lymphoproliferative disease adult T-cell leukemia/lymphoma (ATL) and the neurodegenerative disorder tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM). Replication of HTLV-1 is under the control of two major trans-acting proteins, Tax and Rex. Previous studies suggested that Tax activates transcription from the viral long terminal repeat (LTR) through recruitment of cellular CREB and transcriptional coactivators. Other studies reported that Rex acts posttranscriptionally and allows the cytoplasmic export of unspliced or incompletely spliced viral mRNAs carrying gag/pol and env only. As opposed to HIV's Rev-responsive element (RRE), the Rex-responsive element (RxRE) is present in all viral mRNAs in HTLV-1. However, based on indirect observations, it is believed that nuclear export and expression of the doubly spliced tax/rex RNA are Rex independent. In this study, we demonstrate that Rex does stimulate Tax expression, through nuclear-cytoplasmic export of the tax/rex RNA, even though a Rex-independent basal export mechanism exists. This effect was dependent upon the RxRE element and the RNA-binding activity of Rex. In addition, Rex-mediated export of tax/rex RNA was CRM1 dependent and inhibited by leptomycin B treatment. RNA immunoprecipitation (RNA-IP) experiments confirmed Rex binding to the tax/rex RNA in both transfected cells with HTLV-1 molecular clones and HTLV-1-infected T cells. Since both Rex and p30 interact with the tax/rex RNA and with one another, this may offer a temporal and dynamic regulation of HTLV-1 replication. Our results shed light on HTLV-1 replication and reveal a more complex regulatory network than previously anticipated. PMID:22318152

Comparison of maternal milk (breastmilk) expression methods in an African nursery.

PubMed

Slusher, Tina M; Slusher, Ida L; Keating, Elizabeth M; Curtis, Beverly A; Smith, Eleanor A; Orodriyo, Elizabeth; Awori, Sussane; Nakakeeto, Margaret K

2012-04-01

This study compares maternal milk volumes (MMVs) of Ugandan mothers whose infants were in a special care nursery and who used one of three maternal milk expression techniques: double electric breast pump, single non-electric manual breast pump, and hand breastmilk expression. A convenience sample of 161 Ugandan mothers of infants who were either too immature or ill to independently feed from the breast yet healthy enough to survive in an environment without ventilator support (birth weights, 0.84-3.8 kg) were assigned to one of three maternal milk expressions: Group 1, double electric breast pump (n=55); Group 2, single non-electric manual breast pump (n=59); and Group 3, hand breastmilk expression (n=47). Data were collected over a 7-day period (from day 1 postpartum to day 7 postpartum), and mean MMVs were measured and compared among the groups. The mean daily MMVs were as follows: Group 1, mean=647 mL (SD=310); Group 2, mean=520 mL (SD=298); and Group 3, mean=434 mL (SD=291). Results from one-way analysis of variance revealed significant differences in the mean MMV based on the method of maternal milk expression (p=0.0019). Further analysis using Tukey's HSD Test revealed significant differences in the MMV between Groups 1 and 3 (p < 0.01), but not between Groups 1 and 2 or between Groups 2 and 3. Electric breast pumps provided the highest mean MMV; however, many mothers obtained adequate feeding volumes for their infants' daily nutritional needs with the single non-electric manual breast pump and hand breastmilk expression.

mRNA quality control is bypassed for immediate export of stress-responsive transcripts.

PubMed

Zander, Gesa; Hackmann, Alexandra; Bender, Lysann; Becker, Daniel; Lingner, Thomas; Salinas, Gabriela; Krebber, Heike

2016-12-12

Cells grow well only in a narrow range of physiological conditions. Surviving extreme conditions requires the instantaneous expression of chaperones that help to overcome stressful situations. To ensure the preferential synthesis of these heat-shock proteins, cells inhibit transcription, pre-mRNA processing and nuclear export of non-heat-shock transcripts, while stress-specific mRNAs are exclusively exported and translated. How cells manage the selective retention of regular transcripts and the simultaneous rapid export of heat-shock mRNAs is largely unknown. In Saccharomyces cerevisiae, the shuttling RNA adaptor proteins Npl3, Gbp2, Hrb1 and Nab2 are loaded co-transcriptionally onto growing pre-mRNAs. For nuclear export, they recruit the export-receptor heterodimer Mex67-Mtr2 (TAP-p15 in humans). Here we show that cellular stress induces the dissociation of Mex67 and its adaptor proteins from regular mRNAs to prevent general mRNA export. At the same time, heat-shock mRNAs are rapidly exported in association with Mex67, without the need for adapters. The immediate co-transcriptional loading of Mex67 onto heat-shock mRNAs involves Hsf1, a heat-shock transcription factor that binds to heat-shock-promoter elements in stress-responsive genes. An important difference between the export modes is that adaptor-protein-bound mRNAs undergo quality control, whereas stress-specific transcripts do not. In fact, regular mRNAs are converted into uncontrolled stress-responsive transcripts if expressed under the control of a heat-shock promoter, suggesting that whether an mRNA undergoes quality control is encrypted therein. Under normal conditions, Mex67 adaptor proteins are recruited for RNA surveillance, with only quality-controlled mRNAs allowed to associate with Mex67 and leave the nucleus. Thus, at the cost of error-free mRNA formation, heat-shock mRNAs are exported and translated without delay, allowing cells to survive extreme situations.

SRSF3 represses the expression of PDCD4 protein by coordinated regulation of alternative splicing, export and translation

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

Park, Seung Kuk; Jeong, Sunjoo, E-mail: sjsj@dankook.ac.kr

2016-02-05

Gene expression is regulated at multiple steps, such as transcription, splicing, export, degradation and translation. Considering diverse roles of SR proteins, we determined whether the tumor-related splicing factor SRSF3 regulates the expression of the tumor-suppressor protein, PDCD4, at multiple steps. As we have reported previously, knockdown of SRSF3 increased the PDCD4 protein level in SW480 colon cancer cells. More interestingly, here we showed that the alternative splicing and the nuclear export of minor isoforms of pdcd4 mRNA were repressed by SRSF3, but the translation step was unaffected. In contrast, only the translation step of the major isoform of pdcd4 mRNAmore » was repressed by SRSF3. Therefore, overexpression of SRSF3 might be relevant to the repression of all isoforms of PDCD4 protein levels in most types of cancer cell. We propose that SRSF3 could act as a coordinator of the expression of PDCD4 protein via two mechanisms on two alternatively spliced mRNA isoforms.« less

IRAK2 directs stimulus-dependent nuclear export of inflammatory mRNAs

PubMed Central

Yu, Minjia; Qian, Wen; Wang, Han; Zhou, Gao; Chen, Xing; Yang, Hui; Hong, Lingzi; Zhao, Junjie; Qin, Luke; Fukuda, Koichi; Flotho, Annette; Gao, Ji; Dongre, Ashok; Carman, Julie A; Kang, Zizhen; Su, Bing; Kern, Timothy S; Smith, Jonathan D; Hamilton, Thomas A; Melchior, Frauke; Fox, Paul L

2017-01-01

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation. PMID:28990926

IRAK2 directs stimulus-dependent nuclear export of inflammatory mRNAs.

PubMed

Zhou, Hao; Bulek, Katarzyna; Li, Xiao; Herjan, Tomasz; Yu, Minjia; Qian, Wen; Wang, Han; Zhou, Gao; Chen, Xing; Yang, Hui; Hong, Lingzi; Zhao, Junjie; Qin, Luke; Fukuda, Koichi; Flotho, Annette; Gao, Ji; Dongre, Ashok; Carman, Julie A; Kang, Zizhen; Su, Bing; Kern, Timothy S; Smith, Jonathan D; Hamilton, Thomas A; Melchior, Frauke; Fox, Paul L; Li, Xiaoxia

2017-10-09

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation.

Nucleocytoplasmic Transport of RNAs and RNA-Protein Complexes.

PubMed

Sloan, Katherine E; Gleizes, Pierre-Emmanuel; Bohnsack, Markus T

2016-05-22

RNAs and ribonucleoprotein complexes (RNPs) play key roles in mediating and regulating gene expression. In eukaryotes, most RNAs are transcribed, processed and assembled with proteins in the nucleus and then either function in the cytoplasm or also undergo a cytoplasmic phase in their biogenesis. This compartmentalization ensures that sequential steps in gene expression and RNP production are performed in the correct order and it allows important quality control mechanisms that prevent the involvement of aberrant RNAs/RNPs in these cellular pathways. The selective exchange of RNAs/RNPs between the nucleus and cytoplasm is enabled by nuclear pore complexes, which function as gateways between these compartments. RNA/RNP transport is facilitated by a range of nuclear transport receptors and adaptors, which are specifically recruited to their cargos and mediate interactions with nucleoporins to allow directional translocation through nuclear pore complexes. While some transport factors are only responsible for the export/import of a certain class of RNA/RNP, others are multifunctional and, in the case of large RNPs, several export factors appear to work together to bring about export. Recent structural studies have revealed aspects of the mechanisms employed by transport receptors to enable specific cargo recognition, and genome-wide approaches have provided the first insights into the diverse composition of pre-mRNPs during export. Furthermore, the regulation of RNA/RNP export is emerging as an important means to modulate gene expression under stress conditions and in disease. Copyright © 2015. Published by Elsevier Ltd.

Variable C : N : P stoichiometry of dissolved organic matter cycling in the Community Earth System Model

DOE PAGES

Letscher, R. T.; Moore, J. K.; Teng, Y. -C.; ...

2014-06-16

Dissolved organic matter (DOM) plays an important role in the ocean's biological carbon pump by providing an advective/mixing pathway for ~ 20% of export production. DOM is known to have a stoichiometry depleted in nitrogen (N) and phosphorus (P) compared to the particulate organic matter pool, a~fact that is often omitted from biogeochemical-ocean general circulation models. However the variable C : N : P stoichiometry of DOM becomes important when quantifying carbon export from the upper ocean and linking the nutrient cycles of N and P with that of carbon. Here we utilize recent advances in DOM observational data coveragemore » and offline tracer-modeling techniques to objectively constrain the variable production and remineralization rates of the DOM C / N / P pools in a simple biogeochemical-ocean model of DOM cycling. The optimized DOM cycling parameters are then incorporated within the Biogeochemical Elemental Cycling (BEC) component of the Community Earth System Model and validated against the compilation of marine DOM observations. The optimized BEC simulation including variable DOM C : N : P cycling was found to better reproduce the observed DOM spatial gradients than simulations that used the canonical Redfield ratio. Global annual average export of dissolved organic C, N, and P below 100 m was found to be 2.28 Pg C yr -1 (143 Tmol C yr -1), 16.4 Tmol N yr -1, and 1 Tmol P yr -1, respectively with an average export C : N : P stoichiometry of 225 : 19 : 1 for the semilabile (degradable) DOM pool. DOC export contributed ~ 25% of the combined organic C export to depths greater than 100 m.« less

Variable C : N : P stoichiometry of dissolved organic matter cycling in the Community Earth System Model

DOE PAGES

Letscher, R. T.; Moore, J. K.; Teng, Y. -C.; ...

2015-01-12

Dissolved organic matter (DOM) plays an important role in the ocean's biological carbon pump by providing an advective/mixing pathway for ~ 20% of export production. DOM is known to have a stoichiometry depleted in nitrogen (N) and phosphorus (P) compared to the particulate organic matter pool, a fact that is often omitted from biogeochemical ocean general circulation models. However the variable C : N : P stoichiometry of DOM becomes important when quantifying carbon export from the upper ocean and linking the nutrient cycles of N and P with that of carbon. Here we utilize recent advances in DOM observationalmore » data coverage and offline tracer-modeling techniques to objectively constrain the variable production and remineralization rates of the DOM C : N : P pools in a simple biogeochemical-ocean model of DOM cycling. The optimized DOM cycling parameters are then incorporated within the Biogeochemical Elemental Cycling (BEC) component of the Community Earth System Model (CESM) and validated against the compilation of marine DOM observations. The optimized BEC simulation including variable DOM C : N : P cycling was found to better reproduce the observed DOM spatial gradients than simulations that used the canonical Redfield ratio. Global annual average export of dissolved organic C, N, and P below 100 m was found to be 2.28 Pg C yr -1 (143 Tmol C yr -1, 16.4 Tmol N yr -1, and 1 Tmol P yr -1, respectively, with an average export C : N : P stoichiometry of 225 : 19 : 1 for the semilabile (degradable) DOM pool. Dissolved organic carbon (DOC) export contributed ~ 25% of the combined organic C export to depths greater than 100 m.« less

mRNA export in the apicomplexan parasite Toxoplasma gondii: emerging divergent components of a crucial pathway.

PubMed

Ávila, Andréa Rodrigues; Cabezas-Cruz, Alexjandro; Gissot, Mathieu

2018-01-25

Control of gene expression is crucial for parasite survival and is the result of a series of processes that are regulated to permit fine-tuning of gene expression in response to biological changes during the life-cycle of apicomplexan parasites. Control of mRNA nuclear export is a key process in eukaryotic cells but is poorly understood in apicomplexan parasites. Here, we review recent knowledge regarding this process with an emphasis on T. gondii. We describe the presence of divergent orthologs and discuss structural and functional differences in export factors between apicomplexans and other eukaryotic lineages. Undoubtedly, the use of the CRISPR/Cas9 system in high throughput screenings associated with the discovery of mRNA nuclear export complexes by proteomic analysis will contribute to identify these divergent factors. Ligand-based or structure-based strategies may be applied to investigate the potential use of these proteins as targets for new antiprotozoal agents.

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

Zhang, Aijie, E-mail: zhangaijie1986@163.com

Alpha-naphthylisothiocyanate (ANIT) is a toxicant that is widely used in rodents to model human intrahepatic cholestasis. The aim of the study is to investigate whether effects of dioscin on ANIT-induced cholestasis are related to changes in expression of hepatic transporters in rats. Effects of dioscin on cholestasis were examined by histology and biochemical marker levels. The functional changes of hepatic transporters were determined by in vitro, in situ and in vivo. qRT-PCR and western blot were used to assess the expression of hepatic transporters in cholestatic rats. Dioscin administration could ameliorate cholestasis, as evidenced by reduced biochemical markers as wellmore » as improved liver pathology. The uptakes of organic anion transporting polypeptide (Oatp) substrates were altered in liver uptake index in vivo, perfused rat liver in situ and isolated rat hepatocytes in vitro in cholestasis rats. qRT-PCR and western blot analysis indicated co-treatment of ANIT with dioscin prevented the adaptive down-regulation of Oatp1a1, 1b2, and prompted the up-regulation of Oatp1a4, multidrug resistance-associated protein (Mrp) 2 and bile salt export pump (Bsep). In addition, concerted effects on Mrp2 and Bsep occurred through up-regulation of small heterodimer partner by activating farnesoid X receptor. Dioscin might prevent impairment of hepatic function by restoring hepatic transporter expression. - Highlights: • Cholestasis was improved by dioscin via up-regulating the expression of Oatps, Mrp2 and Bsep. • Dioscin regulated Mrp2 and Bsep via activating FXR. • Dioscin may be a candidate drug for the prevention of intrahepatic cholestasis.« less

Xanthohumol, a hop-derived prenylated flavonoid, promotes macrophage reverse cholesterol transport.

PubMed

Hirata, Hiroshi; Uto-Kondo, Harumi; Ogura, Masatsune; Ayaori, Makoto; Shiotani, Kazusa; Ota, Ami; Tsuchiya, Youichi; Ikewaki, Katsunori

2017-09-01

Xanthohumol, a prominent prenyl flavonoid from the hop plant (Humulus lupulus L.), is suggested to be antiatherogenic since it reportedly increases high-density lipoprotein (HDL) cholesterol levels. It is not clear whether xanthohumol promotes reverse cholesterol transport (RCT), the most important antiatherogenic property of HDL; therefore, we investigated the effects of xanthohumol on macrophage-to-feces RCT using a hamster model as a CETP-expressing species. In vivo RCT experiments showed that xanthohumol significantly increased fecal appearance of the tracer derived from intraperitoneally injected [ 3 H]-cholesterol-labeled macrophages. Ex vivo experiments were then employed to investigate the detailed mechanism by which xanthohumol enhanced RCT. Cholesterol efflux capacity from macrophages was 1.5-fold higher in xanthohumol-fed hamsters compared with the control group. In addition, protein expression and lecithin-cholesterol acyltransferase activity in the HDL fraction were significantly higher in xanthohumol-fed hamsters compared with the control, suggesting that xanthohumol promoted HDL maturation. Hepatic transcript analysis revealed that xanthohumol increased mRNA expression of abcg8 and cyp7a1. In addition, protein expressions of liver X receptor α and bile pump export protein were increased in the liver by xanthohumol administration when compared with the control, implying that it stimulated bile acid synthesis and cholesterol excretion to feces. In conclusion, our data demonstrate that xanthohumol improves RCT in vivo through cholesterol efflux from macrophages and excretion to feces, leading to antiatherosclerosis effects. It remains to be elucidated whether enhancement of RCT by xanthohumol could prove valuable in humans. Copyright © 2017 Elsevier Inc. All rights reserved.

A short conserved motif in ALYREF directs cap- and EJC-dependent assembly of export complexes on spliced mRNAs

PubMed Central

Gromadzka, Agnieszka M.; Steckelberg, Anna-Lena; Singh, Kusum K.; Hofmann, Kay; Gehring, Niels H.

2016-01-01

The export of messenger RNAs (mRNAs) is the final of several nuclear posttranscriptional steps of gene expression. The formation of export-competent mRNPs involves the recruitment of export factors that are assumed to facilitate transport of the mature mRNAs. Using in vitro splicing assays, we show that a core set of export factors, including ALYREF, UAP56 and DDX39, readily associate with the spliced RNAs in an EJC (exon junction complex)- and cap-dependent manner. In order to elucidate how ALYREF and other export adaptors mediate mRNA export, we conducted a computational analysis and discovered four short, conserved, linear motifs present in RNA-binding proteins. We show that mutation in one of the new motifs (WxHD) in an unstructured region of ALYREF reduced RNA binding and abolished the interaction with eIF4A3 and CBP80. Additionally, the mutation impaired proper localization to nuclear speckles and export of a spliced reporter mRNA. Our results reveal important details of the orchestrated recruitment of export factors during the formation of export competent mRNPs. PMID:26773052

Evaluation of expression of NorA efflux pump in ciprofloxacin resistant Staphylococcus aureus against hexahydroquinoline derivative by real-time PCR.

PubMed

Pourmand, Mohammad Reza; Yousefi, Masoud; Salami, Seyed Alireza; Amini, Mohsen

2014-01-01

Staphylococcus aureus causes a wide variety of infections worldwide. Methicillin-resistant S. aureus is one of most common causes of nosocomial and community acquired infections. The fluoroquinolones are an important class of antibiotics that used to treat infections caused by S. aureus. Today, a significant increase in the rate of ciprofloxacin resistance in methicillin-resistant S. aureus strains is concerning. The norA efflux pump is considered as contributors to antibiotic resistance. Here, we aimed to evaluate the expression of norA efflux pump in the presence of hexahydroquinoline derivative in methicillin and ciprofloxacin resistant S. aureus. We were determined minimum inhibitory concentration of ciprofloxacin and hexahydroquinoline derivative and their combination by broth microdilution method against ciprofloxacin resistant S. aureus. The expression of the norA efflux pump gene was evaluated by quantitative Real-time PCR. This study showed that minimum inhibitory concentrations of ciprofloxacin in the presence of hexahydroquinoline derivative in comparison to ciprofloxacin were decreased. Quantitative Real-time PCR identified the increased expression of norA efflux pump gene in methicillin and ciprofloxacin resistant S. aureus strain. The increased expression of norA efflux pump gene may have resulted in the effort of S. aureus to survive. The results showed that the hexahydroquinoline derivative enhanced the antibacterial effect of ciprofloxacin against methicillin and ciprofloxacin resistant S. aureus. Therefore, the derivatives may be used as inhibitors of antibiotic resistance for combination therapy.

NaK-ATPase pump sites in cultured bovine corneal endothelium of varying cell density at confluence.

PubMed

Crawford, K M; Ernst, S A; Meyer, R F; MacCallum, D K

1995-06-01

The driving force for ion and water flow necessary for efficient deturgesence of the corneal stroma resides in the ouabain-sensitive sodium (Na) pump of corneal endothelial cells. Using a cell culture model of corneal endothelial cell hypertrophy, the authors examined the expression of Na pumps at the cell surface to see how this central element of the endothelial pump changed as corneal endothelial cell density decreased to a level associated with corneal decompensation in vivo. 3H-ouabain binding to NaK-ATPase at saturating conditions was used to quantitate the number of Na pump sites on cultured bovine corneal endothelial cells as the confluent density decreased from approximately 2750 cells/mm2 to approximately 275 cells/mm2. The mean number of Na pump sites per cell at confluence (1.92 +/- 0.07 x 10(6)) did not change as the cell density decreased 2.7-fold from 2763 cells/mm2 to 1000 cells/mm2. However, pump site expression doubled to approximately 4 x 10(6) sites/cell as the cell density decreased from 1000 cells/mm2 to 275 cells/mm2. Despite the incremental increase in Na pump site expression that occurred as the cells hypertrophied below a density of 1000/mm2 to achieve confluence, this increase was insufficient to prevent a decrease in Na pump site density of the intact monolayer, expressed as pump sites/mm2. The confluent cell density of cultured bovine corneal endothelial cells can be varied from that found in the normal native cornea to that associated with corneal decompensation. In confluent cultures with cell densities ranging from 2750 cells/mm2 to 1000 cells/mm2, the number of pump sites per cell remains relatively unchanged. Below cell densities of 1000 cells/mm2, the number of pump sites per cell progressively increases. The increased Na pump site abundance in markedly hypertrophied endothelial cells cannot adequately compensate for the progressive reduction in the number of transporting cells per unit area within the intact monolayer. Even when considered with the decrease in the size of the paracellular ion conductive pathway that is a consequence of progressive endothelial hypertrophy, the overall pumping capacity of the intact endothelial monolayer declines.

The Apical Localization of Na+, K+-ATPase in Cultured Human Retinal Pigment Epithelial Cells Depends on Expression of the β2 Subunit

PubMed Central

Lobato-Álvarez, Jorge A.; Roldán, María L.; López-Murillo, Teresa del Carmen; González-Ramírez, Ricardo; Bonilla-Delgado, José; Shoshani, Liora

2016-01-01

Na+, K+-ATPase, or the Na+ pump, is a key component in the maintenance of the epithelial phenotype. In most epithelia, the pump is located in the basolateral domain. Studies from our laboratory have shown that the β1 subunit of Na+, K+-ATPase plays an important role in this mechanism because homotypic β1-β1 interactions between neighboring cells stabilize the pump in the lateral membrane. However, in the retinal pigment epithelium (RPE), the Na+ pump is located in the apical domain. The mechanism of polarization in this epithelium is unclear. We hypothesized that the apical polarization of the pump in RPE cells depends on the expression of its β2 subunit. ARPE-19 cells cultured for up to 8 weeks on inserts did not polarize, and Na+, K+-ATPase was expressed in the basolateral membrane. In the presence of insulin, transferrin and selenic acid (ITS), ARPE-19 cells cultured for 4 weeks acquired an RPE phenotype, and the Na+ pump was visible in the apical domain. Under these conditions, Western blot analysis was employed to detect the β2 isoform and immunofluorescence analysis revealed an apparent apical distribution of the β2 subunit. qPCR results showed a time-dependent increase in the level of β2 isoform mRNA, suggesting regulation at the transcriptional level. Moreover, silencing the expression of the β2 isoform in ARPE-19 cells resulted in a decrease in the apical localization of the pump, as assessed by the mislocalization of the α2 subunit in that domain. Our results demonstrate that the apical polarization of Na+, K+-ATPase in RPE cells depends on the expression of the β2 subunit. PMID:27774068

The Apical Localization of Na+, K+-ATPase in Cultured Human Retinal Pigment Epithelial Cells Depends on Expression of the β2 Subunit.

PubMed

Lobato-Álvarez, Jorge A; Roldán, María L; López-Murillo, Teresa Del Carmen; González-Ramírez, Ricardo; Bonilla-Delgado, José; Shoshani, Liora

2016-01-01

Na + , K + -ATPase, or the Na + pump, is a key component in the maintenance of the epithelial phenotype. In most epithelia, the pump is located in the basolateral domain. Studies from our laboratory have shown that the β 1 subunit of Na + , K + -ATPase plays an important role in this mechanism because homotypic β 1 -β 1 interactions between neighboring cells stabilize the pump in the lateral membrane. However, in the retinal pigment epithelium (RPE), the Na + pump is located in the apical domain. The mechanism of polarization in this epithelium is unclear. We hypothesized that the apical polarization of the pump in RPE cells depends on the expression of its β 2 subunit. ARPE-19 cells cultured for up to 8 weeks on inserts did not polarize, and Na + , K + -ATPase was expressed in the basolateral membrane. In the presence of insulin, transferrin and selenic acid (ITS), ARPE-19 cells cultured for 4 weeks acquired an RPE phenotype, and the Na + pump was visible in the apical domain. Under these conditions, Western blot analysis was employed to detect the β 2 isoform and immunofluorescence analysis revealed an apparent apical distribution of the β 2 subunit. qPCR results showed a time-dependent increase in the level of β 2 isoform mRNA, suggesting regulation at the transcriptional level. Moreover, silencing the expression of the β 2 isoform in ARPE-19 cells resulted in a decrease in the apical localization of the pump, as assessed by the mislocalization of the α 2 subunit in that domain. Our results demonstrate that the apical polarization of Na + , K + -ATPase in RPE cells depends on the expression of the β 2 subunit.

Effect of Yin-Zhi-Huang on up-regulation of Oatp2, Ntcp, and Mrp2 proteins in estrogen-induced rat cholestasis.

PubMed

Zhang, Guoqiang; Zhou, Yan; Rao, Zhi; Qin, Hongyan; Wei, Yuhui; Ren, Jiangxia; Zhou, Liting; Wu, Xin'an

2015-03-01

Yin-Zhi-Huang (YZH), a prescription of traditional Chinese medicine, is widely used to treat neonatal jaundice or cholestasis. This study investigates the regulatory effect of YZH on the localization and expression of organic anion transporting polypeptides 2 (Oatp2), Na(+)-taurocholate co-transporting polypeptide (Ntcp), multidrug-resistance-associated protein 2 (Mrp2), and bile salt export pump (Bsep) in estrogen-induced cholestasis rats. Cholestasis model rats were induced via subcutaneous injection of estradiol benzoate (EB, 5 mg/kg/d) for 5 d. Other EB-induced rats were treated with saline (2 ml) or YZH (1.5 g/kg, two times a day) for 7, 14, and 21 d. The biochemical and pathologic examinations were performed. Moreover, the localization and expression of Oatp2, Ntcp, Mrp2, and Bsep were determined by immunohistochemisty and Western blotting, respectively. YZH treatment could significantly decrease the serum total bile acids (TBA) (4.9 ± 0.6-2.8 ± 0.8) and direct bilirubin (DBIL) (2.6 ± 0.7-1.0 ± 0.1) levels, improve the histological disorganization, and, respectively, increase the expression of Oatp2 and Ntcp by 46% and 28% compared with saline-treated (p < 0.05) rats at 14 d. The expression of Mrp2 increased by 45% was observed in YZH treated compared with saline-treated (p < 0.05) rats at 7 d. However, there was a little change in the expression of Bsep (p > 0.05) after YZH treatment for 7, 14, and 21 d. In conclusion, the therapeutic effect of YZH to cholestasis could be attributed to the regulation of Oatp2, Ntcp, Mrp2, and Bsep.

Nuclear factor-E2-related factor 2 is a major determinant of bile acid homeostasis in the liver and intestine

PubMed Central

Weerachayaphorn, Jittima; Mennone, Albert; Soroka, Carol J.; Harry, Kathy; Hagey, Lee R.; Kensler, Thomas W.

2012-01-01

The transcription factor nuclear factor-E2-related factor 2 (Nrf2) is a key regulator for induction of hepatic detoxification and antioxidant mechanisms, as well as for certain hepatobiliary transporters. To examine the role of Nrf2 in bile acid homeostasis and cholestasis, we assessed the determinants of bile secretion and bile acid synthesis and transport before and after bile duct ligation (BDL) in Nrf2−/− mice. Our findings indicate reduced rates of biliary bile acid and GSH excretion, higher levels of intrahepatic bile acids, and decreased expression of regulators of bile acid synthesis, Cyp7a1 and Cyp8b1, in Nrf2−/− compared with wild-type control mice. The mRNA expression of the bile acid transporters bile salt export pump (Bsep) and organic solute transporter (Ostα) were increased in the face of impaired expression of the multidrug resistance-associated proteins Mrp3 and Mrp4. Deletion of Nrf2 also decreased ileal apical sodium-dependent bile acid transporter (Asbt) expression, leading to reduced bile acid reabsorption and increased loss of bile acid in feces. Finally, when cholestasis is induced by BDL, liver injury was not different from that in wild-type BDL mice. These Nrf2−/− mice also had increased pregnane X receptor (Pxr) and Cyp3a11 mRNA expression in association with enhanced hepatic bile acid hydroxylation. In conclusion, this study finds that Nrf2 plays a major role in the regulation of bile acid homeostasis in the liver and intestine. Deletion of Nrf2 results in a cholestatic phenotype but does not augment liver injury following BDL. PMID:22345550

Dietary fish oil regulates gene expression of cholesterol and bile acid transporters in mice.

PubMed

Kamisako, Toshinori; Tanaka, Yuji; Ikeda, Takanori; Yamamoto, Kazuo; Ogawa, Hiroshi

2012-03-01

  Fish oil rich in n-3 polyunsaturated fatty acids is known to affect hepatic lipid metabolism. Several studies have demonstrated that fish oil may affect the bile acid metabolism as well as lipid metabolism, whereas only scarce data are available. The aim of this study was to investigate the effect of fish oil on the gene expression of the transporters and enzymes related to bile acid as well as lipid metabolism in the liver and small intestine.   Seven-week old male C57BL/6 mice were fed diets enriched in 10% soybean oil or 10% fish oil for 4 weeks. After 4 weeks, blood, liver and small intestine were obtained.   Hepatic mRNA expression of lipids (Abcg5/8, multidrug resistance gene product 2) and bile acids transporters (bile salt export pump, multidrug resistance associated protein 2 and 3, organic solute transporter α) was induced in fish oil-fed mice. Hepatic Cyp8b1, Cyp27a1 and bile acid CoA : amino acid N-acyltransferase were increased in fish oil-fed mice compared with soybean-oil fed mice. Besides, intestinal cholesterol (Abcg5/8) and bile acid transporters (multidrug resistance associated protein 2 and organic solute transporter α) were induced in fish oil-fed mice.   Fish oil induced the expression of cholesterol and bile acid transporters not only in liver but in intestine. The upregulation of Abcg5/g8 by fish oil is caused by an increase in cellular 27-HOC through Cyp27a1 induction. The hepatic induction of bile acid synthesis through Cyp27a1 may upregulate expression of bile acid transporters in both organs. © 2012 The Japan Society of Hepatology.

Role of regulatory subunits and protein kinase inhibitor (PKI) in determining nuclear localization and activity of the catalytic subunit of protein kinase A.

PubMed

Wiley, J C; Wailes, L A; Idzerda, R L; McKnight, G S

1999-03-05

Regulation of protein kinase A by subcellular localization may be critical to target catalytic subunits to specific substrates. We employed epitope-tagged catalytic subunit to correlate subcellular localization and gene-inducing activity in the presence of regulatory subunit or protein kinase inhibitor (PKI). Transiently expressed catalytic subunit distributed throughout the cell and induced gene expression. Co-expression of regulatory subunit or PKI blocked gene induction and prevented nuclear accumulation. A mutant PKI lacking the nuclear export signal blocked gene induction but not nuclear accumulation, demonstrating that nuclear export is not essential to inhibit gene induction. When the catalytic subunit was targeted to the nucleus with a nuclear localization signal, it was not sequestered in the cytoplasm by regulatory subunit, although its activity was completely inhibited. PKI redistributed the nuclear catalytic subunit to the cytoplasm and blocked gene induction, demonstrating that the nuclear export signal of PKI can override a strong nuclear localization signal. With increasing PKI, the export process appeared to saturate, resulting in the return of catalytic subunit to the nucleus. These results demonstrate that both the regulatory subunit and PKI are able to completely inhibit the gene-inducing activity of the catalytic subunit even when the catalytic subunit is forced to concentrate in the nuclear compartment.

21 CFR 884.5160 - Powered breast pump.

Code of Federal Regulations, 2010 CFR

2010-04-01

... § 884.5160 Powered breast pump. (a) Identification. A powered breast pump in an electrically powered suction device used to express milk from the breast. (b) Classification. Class II (performance standards). ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Powered breast pump. 884.5160 Section 884.5160...

21 CFR 884.5160 - Powered breast pump.

Code of Federal Regulations, 2013 CFR

2013-04-01

... § 884.5160 Powered breast pump. (a) Identification. A powered breast pump in an electrically powered suction device used to express milk from the breast. (b) Classification. Class II (performance standards). ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Powered breast pump. 884.5160 Section 884.5160...

21 CFR 884.5160 - Powered breast pump.

Code of Federal Regulations, 2011 CFR

2011-04-01

... § 884.5160 Powered breast pump. (a) Identification. A powered breast pump in an electrically powered suction device used to express milk from the breast. (b) Classification. Class II (performance standards). ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Powered breast pump. 884.5160 Section 884.5160...

21 CFR 884.5160 - Powered breast pump.

Code of Federal Regulations, 2012 CFR

2012-04-01

... § 884.5160 Powered breast pump. (a) Identification. A powered breast pump in an electrically powered suction device used to express milk from the breast. (b) Classification. Class II (performance standards). ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Powered breast pump. 884.5160 Section 884.5160...

21 CFR 884.5160 - Powered breast pump.

Code of Federal Regulations, 2014 CFR

2014-04-01

... § 884.5160 Powered breast pump. (a) Identification. A powered breast pump in an electrically powered suction device used to express milk from the breast. (b) Classification. Class II (performance standards). ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Powered breast pump. 884.5160 Section 884.5160...

The sodium pump α1 sub-unit: a disease progression–related target for metastatic melanoma treatment

PubMed Central

Mathieu, Véronique; Pirker, Christine; Martin de Lassalle, Elisabeth; Vernier, Mathieu; Mijatovic, Tatjana; DeNeve, Nancy; Gaussin, Jean-François; Dehoux, Mischael; Lefranc, Florence; Berger, Walter; Kiss, Robert

2009-01-01

Melanomas remain associated with dismal prognosis because they are naturally resistant to apoptosis and they markedly metastasize. Up-regulated expression of sodium pump α sub-units has previously been demonstrated when comparing metastatic to non-metastatic melanomas. Our previous data revealed that impairing sodium pump α1 activity by means of selective ligands, that are cardiotonic steroids, markedly impairs cell migration and kills apoptosis-resistant cancer cells. The objective of this study was to determine the expression levels of sodium pump α sub-units in melanoma clinical samples and cell lines and also to characterize the role of α1 sub-units in melanoma cell biology. Quantitative RT-PCR, Western blotting and immunohistochemistry were used to determine the expression levels of sodium pump α sub-units. In vitro cytotoxicity of various cardenolides and of an anti-α1 siRNA was evaluated by means of MTT assay, quantitative videomicroscopy and through apoptosis assays. The in vivo activity of a novel cardenolide UNBS1450 was evaluated in a melanoma brain metastasis model. Our data show that all investigated human melanoma cell lines expressed high levels of the α1 sub-unit, and 33% of human melanomas displayed significant α1 sub-unit expression in correlation with the Breslow index. Furthermore, cardenolides (notably UNBS1450; currently in Phase I clinical trials) displayed marked anti-tumour effects against melanomas in vitro. This activity was closely paralleled by decreases in cMyc expression and by increases in apoptotic features. UNBS1450 also displayed marked anti-tumour activity in the aggressive human metastatic brain melanoma model in vivo. The α1 sodium pump sub-unit could represent a potential novel target for combating melanoma. PMID:19243476

Simulated ground-water flow for a pond-dominated aquifer system near Great Sandy Bottom Pond, Pembroke, Massachusetts

USGS Publications Warehouse

Carlson, Carl S.; Lyford, Forest P.

2005-01-01

A ground-water flow simulation for a 66.4-square-mile area around Great Sandy Bottom (GSB) Pond (105 acres) near Pembroke, Massachusetts, was developed for use by local and State water managers to assess the yields for public water supply of local ponds and wells for average climatic and drought conditions and the effects of water withdrawals on nearby water levels and streamflows. Wetlands and ponds cover about 30 percent of the study area and the aquifer system is dominated by interactions between ground water and the ponds. The three largest surface-water bodies in the study area are Silver Lake (640 acres), Monponsett Pond (590 acres), and Oldham Pond (236 acres). The study area is drained by tributaries of the Taunton River to the southwest, the South and North Rivers to the northeast, and the Jones River to the southeast. In 2002, 10.8 million gallons per day of water was exported from ponds and 3.5 million gallons per day from wells was used locally for public supply. A transient ground-water-flow model with 69 monthly stress periods spanning the period from January 1998 through September 2003 was calibrated to stage at GSB Pond and nearby Silver Lake and streamflow and water levels collected from September 2002 through September 2003. The calibrated model was used to assess hydrologic responses to a variety of water-use and climatic conditions. Simulation of predevelopment (no pumping or export) average monthly (1949-2002) water-level conditions caused the GSB Pond level to increase by 6.3 feet from the results of a simulation using average 2002 pumping for all wells, withdrawals, and exports. Most of this decline can be attributed to pumping, withdrawals, and exports of water from sites away from GSB Pond. The effects of increasing the export rate from GSB Pond by 1.25 and 1.5 times the 2002 rate were a lowering of pond levels by a maximum of 1.6 and 2.8 feet, respectively. Simulated results for two different drought conditions, one mild drought similar to that of 1979-82 and a more severe drought similar to that of 1963-66, but with current (2002) pumping, were compared to results for average monthly recharge conditions (1949-2002). Simulated mild drought conditions showed a reduction of GSB Pond level of about 1.3 feet and a lower streamflow of about 1.7 percent in the nearby stream. Simulated severe drought conditions reduced the pond level at GSB Pond by almost 7 feet and lowered streamflow by about 37 percent. Varying cranberry-irrigation practices had little effect on simulated GSB Pond water levels, but may be important in other ponds. The model was most sensitive to changes in areal recharge. An increase and decrease of 22 percent in recharge produced changes in the GSB Pond water level of +1.4 feet and -2.4 feet, respectively. The accuracy of simulation results was best in the central portion of the study area in the immediate location of GSB Pond. The model was developed with the study-area boundary far enough away from the GSB Pond area that the boundary would have minimal effect on the water levels in GSB Pond, nearby ponds, and the underlying aquifer system. The model is best suited for use by local and State water managers to assess the effects of different withdrawal scenarios for wells and ponds near GSB Pond and for general delineation of areas contributing recharge to wells and ponds in the vicinity of GSB Pond. The model in its current form may not be well suited to detailed analyses of water budgets and flow patterns for parts of the study area farther from GSB Pond without further investigation, calibration, and data collection.

Sustaining nutrient supply and carbon export in a seasonally-stratifying shelf sea through inconsistent production and remineralisation stoichiometry

NASA Astrophysics Data System (ADS)

Humphreys, Matthew; Moore, Mark; Achterberg, Eric; Chowdhury, Mohammed; Griffiths, Alex; Hartman, Susan; Hopkins, Joanne; Hull, Tom; Kivimäe, Caroline; Sivyer, Dave; Smilenova, Angelina; Wihsgott, Juliane; Woodward, Malcolm

2017-04-01

Continental shelf seas support 15-20% of global primary productivity despite covering only about 5% of the Earth's ocean surface area. As a result, they may have a significant role in oceanic uptake and storage of carbon dioxide (CO2) from the atmosphere, through the 'continental shelf pump' mechanism. The northwest European continental shelf, in particular the Celtic Sea (50°N 8°W), was the target of extensive biogeochemical sampling from March 2014 to September 2015, as part of the UK Shelf Sea Biogeochemistry research programme (UK-SSB). Here, we use the UK-SSB carbonate chemistry and macronutrient measurements to investigate the biogeochemical seasonality in the temperate, seasonally-stratifying Celtic Sea. During the spring-summer, near-surface biological activity removed dissolved inorganic carbon and nutrients, some of which were then exported into the deeper layer. We calculated vertical inventories of these variables throughout 1.5 seasonal cycles and attempted to correct these for air-sea CO2 exchange, advection and denitrification, thus isolating the combined effect of net community production and remineralisation on the inorganic macronutrient inventories, and revealing fluctuating deviations from Redfield stoichiometry. Here, we discuss the capacity of these stoichiometric inconsistencies to sustain the Celtic Sea nutrient supply, and thus examine whether an effective continental shelf pump for CO2 could operate in this region.

Substantial energy input to the mesopelagic ecosystem from the seasonal mixed-layer pump

PubMed Central

Dall’Olmo, Giorgio; Dingle, James; Polimene, Luca; Brewin, Robert J.W.; Claustre, Hervé

2016-01-01

The “mesopelagic” is the region of the ocean between about 100 and 1000 m that harbours one of the largest ecosystems and fish stocks on the planet1,2. This vastly unexplored ecosystem is believed to be mostly sustained by chemical energy, in the form of fast-sinking particulate organic carbon, supplied by the biological carbon pump3. Yet, this supply appears insufficient to match mesopelagic metabolic demands4–6. The mixed-layer pump is a physically-driven biogeochemical process7–11 that could further contribute to meet these energetic requirements. However, little is known about the magnitude and spatial distribution of this process at the global scale. Here we show that the mixed-layer pump supplies an important seasonal flux of organic carbon to the mesopelagic. By combining mixed-layer depths from Argo floats with satellite retrievals of particulate organic carbon, we estimate that this pump exports a global flux of about 0.3 Pg C yr−1 (range 0.1 – 0.5 Pg C yr−1). In high-latitude regions where mixed-layers are deep, this flux is on average 23%, but can be greater than 100% of the carbon supplied by fast sinking particles. Our results imply that a relatively large flux of organic carbon is missing from current energy budgets of the mesopelagic. PMID:27857779

High particulate organic carbon export during the decline of a vast diatom bloom in the Atlantic sector of the Southern Ocean

NASA Astrophysics Data System (ADS)

Roca-Martí, Montserrat; Puigcorbé, Viena; Iversen, Morten H.; van der Loeff, Michiel Rutgers; Klaas, Christine; Cheah, Wee; Bracher, Astrid; Masqué, Pere

2017-04-01

Carbon fixation by phytoplankton plays a key role in the uptake of atmospheric CO2 in the Southern Ocean. Yet, it still remains unclear how efficiently the particulate organic carbon (POC) is exported and transferred from ocean surface waters to depth during phytoplankton blooms. In addition, little is known about the processes that control the flux attenuation within the upper twilight zone. Here, we present results of downward POC and particulate organic nitrogen fluxes during the decline of a vast diatom bloom in the Atlantic sector of the Southern Ocean in summer 2012. We used thorium-234 (234Th) as a particle tracer in combination with drifting sediment traps (ST). Their simultaneous use evidenced a sustained high export rate of 234Th at 100 m depth in the weeks prior to and during the sampling period. The entire study area, of approximately 8000 km2, showed similar vertical export fluxes in spite of the heterogeneity in phytoplankton standing stocks and productivity, indicating a decoupling between production and export. The POC fluxes at 100 m were high, averaging 26±15 mmol C m-2 d-1, although the strength of the biological pump was generally low. Only <20% of the daily primary production reached 100 m, presumably due to an active recycling of carbon and nutrients. Pigment analyses indicated that direct sinking of diatoms likely caused the high POC transfer efficiencies ( 60%) observed between 100 and 300 m, although faecal pellets and transport of POC linked to zooplankton vertical migration might have also contributed to downward fluxes.

Do Continental Shelves Act as an Atmospheric CO2 Sink?

NASA Astrophysics Data System (ADS)

Cai, W.

2003-12-01

Recent air-to-sea CO2 flux measurements at several major continental shelves (European Atlantic Shelves, East China Sea and U.S. Middle Atlantic Bight) suggest that shelves may act as a one-way pump and absorb atmospheric CO2 into the ocean. These observations also favor the argument that continental shelves are autotrophic (i.e., net production of organic carbon, OC). The U.S. South Atlantic Bight (SAB) contrasts these findings in that it acts as a strong source of CO2 to the atmosphere while simultaneously exporting dissolved inorganic carbon (DIC) to the open ocean. We report pCO2, DIC, and alkalinity data from the SAB collected in 8 cruises along a transect from the shore to the shelf break in the central SAB. The shelf-wide net heterotrophy and carbon exports in the SAB are subsidized by the export of OC from the abundant intertidal marshes, which are a sink for atmospheric CO2. It is proposed here that the SAB represents a marsh-dominated heterotrophic ocean margin as opposed to river-dominated autotrophic margins. To further investigate why margins may behave differently in term of CO2 sink/source, the physical and biological conditions of several western boundary current margins are compared. Based on this and other studies, DIC export flux from margins to the open ocean must be significant in the overall global ocean carbon budget.

Iron defecation by sperm whales stimulates carbon export in the Southern Ocean

PubMed Central

Lavery, Trish J.; Roudnew, Ben; Gill, Peter; Seymour, Justin; Seuront, Laurent; Johnson, Genevieve; Mitchell, James G.; Smetacek, Victor

2010-01-01

The iron-limited Southern Ocean plays an important role in regulating atmospheric CO2 levels. Marine mammal respiration has been proposed to decrease the efficiency of the Southern Ocean biological pump by returning photosynthetically fixed carbon to the atmosphere. Here, we show that by consuming prey at depth and defecating iron-rich liquid faeces into the photic zone, sperm whales (Physeter macrocephalus) instead stimulate new primary production and carbon export to the deep ocean. We estimate that Southern Ocean sperm whales defecate 50 tonnes of iron into the photic zone each year. Molar ratios of Cexport ∶Feadded determined during natural ocean fertilization events are used to estimate the amount of carbon exported to the deep ocean in response to the iron defecated by sperm whales. We find that Southern Ocean sperm whales stimulate the export of 4 × 105 tonnes of carbon per year to the deep ocean and respire only 2 × 105 tonnes of carbon per year. By enhancing new primary production, the populations of 12 000 sperm whales in the Southern Ocean act as a carbon sink, removing 2 × 105 tonnes more carbon from the atmosphere than they add during respiration. The ability of the Southern Ocean to act as a carbon sink may have been diminished by large-scale removal of sperm whales during industrial whaling. PMID:20554546

Vitamin K3 Induces the Expression of the Stenotrophomonas maltophilia SmeVWX Multidrug Efflux Pump.

PubMed

Blanco, P; Corona, F; Sánchez, M B; Martínez, J L

2017-05-01

Stenotrophomonas maltophilia is an opportunistic pathogen with increasing prevalence, which is able to cause infections in immunocompromised patients or in those with a previous pathology. The treatment of the infections caused by this bacterium is often complicated due to the several intrinsic antibiotic resistance mechanisms that it presents. Multidrug efflux pumps are among the best-studied mechanisms of S. maltophilia antibiotic resistance. Some of these efflux pumps have a basal expression level but, in general, their expression is often low and only reaches high levels when the local regulator is mutated or bacteria are in the presence of an effector. In the current work, we have developed a yellow fluorescent protein (YFP)-based sensor with the aim to identify effectors able to trigger the expression of SmeVWX, an efflux pump that confers resistance to quinolones, chloramphenicol, and tetracycline when it is expressed at high levels. With this purpose in mind, we tested a variety of different compounds and analyzed the fluorescence signal given by the expression of YFP under the control of the smeVWX promoter. Among the tested compounds, vitamin K 3 , which is a compound belonging to the 2-methyl-1,4-naphthoquinone family, is produced by plants in defense against infection, and has increasing importance in human therapy, was able to induce the expression of the SmeVWX efflux pump. In addition, a decrease in the susceptibility of S. maltophilia to ofloxacin and chloramphenicol was observed in the presence of vitamin K 3 , in both wild-type and smeW -deficient strains. Copyright © 2017 American Society for Microbiology.

A short conserved motif in ALYREF directs cap- and EJC-dependent assembly of export complexes on spliced mRNAs.

PubMed

Gromadzka, Agnieszka M; Steckelberg, Anna-Lena; Singh, Kusum K; Hofmann, Kay; Gehring, Niels H

2016-03-18

The export of messenger RNAs (mRNAs) is the final of several nuclear posttranscriptional steps of gene expression. The formation of export-competent mRNPs involves the recruitment of export factors that are assumed to facilitate transport of the mature mRNAs. Using in vitro splicing assays, we show that a core set of export factors, including ALYREF, UAP56 and DDX39, readily associate with the spliced RNAs in an EJC (exon junction complex)- and cap-dependent manner. In order to elucidate how ALYREF and other export adaptors mediate mRNA export, we conducted a computational analysis and discovered four short, conserved, linear motifs present in RNA-binding proteins. We show that mutation in one of the new motifs (WxHD) in an unstructured region of ALYREF reduced RNA binding and abolished the interaction with eIF4A3 and CBP80. Additionally, the mutation impaired proper localization to nuclear speckles and export of a spliced reporter mRNA. Our results reveal important details of the orchestrated recruitment of export factors during the formation of export competent mRNPs. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Nuclear DDX3 expression predicts poor outcome in colorectal and breast cancer

PubMed Central

Heerma van Voss, Marise R; Vesuna, Farhad; Bol, Guus M; Meeldijk, Jan; Raman, Ana; Offerhaus, G Johan; Buerger, Horst; Patel, Arvind H; van der Wall, Elsken; van Diest, Paul J; Raman, Venu

2017-01-01

Purpose DEAD box protein 3 (DDX3) is an RNA helicase with oncogenic properties that shuttles between the cytoplasm and nucleus. The majority of DDX3 is found in the cytoplasm, but a subset of tumors has distinct nuclear DDX3 localization of yet unknown biological significance. This study aimed to evaluate the significance of and mechanisms behind nuclear DDX3 expression in colorectal and breast cancer. Methods Expression of nuclear DDX3 and the nuclear exporter chromosome region maintenance 1 (CRM1) was evaluated by immunohistochemistry in 304 colorectal and 292 breast cancer patient samples. Correlations between the subcellular localization of DDX3 and CRM1 and the difference in overall survival between patients with and without nuclear DDX3 were studied. In addition, DDX3 mutants were created for in vitro evaluation of the mechanism behind nuclear retention of DDX3. Results DDX3 was present in the nucleus of 35% of colorectal and 48% of breast cancer patient samples and was particularly strong in the nucleolus. Nuclear DDX3 correlated with worse overall survival in both colorectal (hazard ratio [HR] 2.34, P<0.001) and breast cancer (HR 2.39, P=0.004) patients. Colorectal cancers with nuclear DDX3 expression more often had cytoplasmic expression of the nuclear exporter CRM1 (relative risk 1.67, P=0.04). In vitro analysis of DDX3 deletion mutants demonstrated that CRM1-mediated export was most dependent on the N-terminal nuclear export signal. Conclusion Overall, we conclude that nuclear DDX3 is partially CRM1-mediated and predicts worse survival in colorectal and breast cancer patients, putting it forward as a target for therapeutic intervention with DDX3 inhibitors under development in these cancer types. PMID:28761359

Nuclear DDX3 expression predicts poor outcome in colorectal and breast cancer.

PubMed

Heerma van Voss, Marise R; Vesuna, Farhad; Bol, Guus M; Meeldijk, Jan; Raman, Ana; Offerhaus, G Johan; Buerger, Horst; Patel, Arvind H; van der Wall, Elsken; van Diest, Paul J; Raman, Venu

2017-01-01

DEAD box protein 3 (DDX3) is an RNA helicase with oncogenic properties that shuttles between the cytoplasm and nucleus. The majority of DDX3 is found in the cytoplasm, but a subset of tumors has distinct nuclear DDX3 localization of yet unknown biological significance. This study aimed to evaluate the significance of and mechanisms behind nuclear DDX3 expression in colorectal and breast cancer. Expression of nuclear DDX3 and the nuclear exporter chromosome region maintenance 1 (CRM1) was evaluated by immunohistochemistry in 304 colorectal and 292 breast cancer patient samples. Correlations between the subcellular localization of DDX3 and CRM1 and the difference in overall survival between patients with and without nuclear DDX3 were studied. In addition, DDX3 mutants were created for in vitro evaluation of the mechanism behind nuclear retention of DDX3. DDX3 was present in the nucleus of 35% of colorectal and 48% of breast cancer patient samples and was particularly strong in the nucleolus. Nuclear DDX3 correlated with worse overall survival in both colorectal (hazard ratio [HR] 2.34, P <0.001) and breast cancer (HR 2.39, P =0.004) patients. Colorectal cancers with nuclear DDX3 expression more often had cytoplasmic expression of the nuclear exporter CRM1 (relative risk 1.67, P =0.04). In vitro analysis of DDX3 deletion mutants demonstrated that CRM1-mediated export was most dependent on the N-terminal nuclear export signal. Overall, we conclude that nuclear DDX3 is partially CRM1-mediated and predicts worse survival in colorectal and breast cancer patients, putting it forward as a target for therapeutic intervention with DDX3 inhibitors under development in these cancer types.

Nuclear localization signal regulates porcine circovirus type 2 capsid protein nuclear export through phosphorylation.

PubMed

Hou, Qiang; Hou, Shaohua; Chen, Qing; Jia, Hong; Xin, Ting; Jiang, Yitong; Guo, Xiaoyu; Zhu, Hongfei

2018-02-15

The open reading frame 2 (ORF2) of Porcine circovirus type 2 (PCV2) encodes the major Capsid (Cap) protein, which self-assembles into virus-like particle (VLP) of similar morphology to the PCV2 virion and accumulates in the nucleus through the N-terminal arginine-rich nuclear localization signal (NLS). In this study, PCV2 Cap protein and its derivates were expressed via the baculovirus expression system, and the cellular localization of the recombinant proteins were investigated using anti-Cap mAb by imaging flow cytometry. Analysis of subcellular localization of Cap protein and its variants demonstrated that NLS mediated Cap protein nuclear export as well as nuclear import, and a phosphorylation site (S17) was identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in the NLS domain to regulate Cap protein nuclear export. Phosphorylation of NLS regulating the PCV2 Cap protein nuclear export was also demonstrated in PK15 cells by fluorescence microscopy. Moreover, the influence of Rep and Rep' protein on Cap protein subcellular localization was investigated in PK15 cells. Phosphorylation of NLS regulating Cap protein nuclear export provides more detailed knowledge of the PCV2 viral life cycle. Copyright © 2018 Elsevier B.V. All rights reserved.

Multiple Export Mechanisms for mRNAs

PubMed Central

Delaleau, Mildred; Borden, Katherine L. B.

2015-01-01

Nuclear mRNA export plays an important role in gene expression. We describe the mechanisms of mRNA export including the importance of mRNP assembly, docking with the nuclear basket of the nuclear pore complex (NPC), transit through the central channel of the NPC and cytoplasmic release. We describe multiple mechanisms of mRNA export including NXF1 and CRM1 mediated pathways. Selective groups of mRNAs can be preferentially transported in order to respond to cellular stimuli. RNAs can be selected based on the presence of specific cis-acting RNA elements and binding of specific adaptor proteins. The role that dysregulation of this process plays in human disease is also discussed. PMID:26343730

27 CFR 28.251 - Railway express receipts.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Railway express receipts... § 28.251 Railway express receipts. Where the exportation is to a contiguous foreign country and the shipment is by railway express, a receipt issued by the railway express agency may be accepted in lieu of...

Phase 0 and phase III transport in various organs: combined concept of phases in xenobiotic transport and metabolism.

PubMed

Döring, Barbara; Petzinger, Ernst

2014-08-01

The historical phasing concept of drug metabolism and elimination was introduced to comprise the two phases of metabolism: phase I metabolism for oxidations, reductions and hydrolyses, and phase II metabolism for synthesis. With this concept, biological membrane barriers obstructing the accessibility of metabolism sites in the cells for drugs were not considered. The concept of two phases was extended to a concept of four phases when drug transporters were detected that guided drugs and drug metabolites in and out of the cells. In particular, water soluble or charged drugs are virtually not able to overcome the phospholipid membrane barrier. Drug transporters belong to two main clusters of transporter families: the solute carrier (SLC) families and the ATP binding cassette (ABC) carriers. The ABC transporters comprise seven families with about 20 carriers involved in drug transport. All of them operate as pumps at the expense of ATP splitting. Embedded in the former phase concept, the term "phase III" was introduced by Ishikawa in 1992 for drug export by ABC efflux pumps. SLC comprise 52 families, from which many carriers are drug uptake transporters. Later on, this uptake process was referred to as the "phase 0 transport" of drugs. Transporters for xenobiotics in man and animal are most expressed in liver, but they are also present in extra-hepatic tissues such as in the kidney, the adrenal gland and lung. This review deals with the function of drug carriers in various organs and their impact on drug metabolism and elimination.

Expression and characterization of the Plasmodium translocon of the exported proteins component EXP2.

PubMed

Hakamada, Kazuaki; Watanabe, Hirokazu; Kawano, Ryuji; Noguchi, Keiichi; Yohda, Masafumi

2017-01-22

The malaria parasite Plasmodium falciparum requires the Plasmodium translocon of exported proteins (PTEX) to proliferate in human red blood cells. During the blood stages of malaria, several hundred parasite-encoded proteins are exported from the parasite into the cytosol of red blood cells. PTEX is the translocon for protein export and comprises 5 proteins: EXP2, PTEX150, PTEX88, Hsp101 and TRX2. Among them, EXP2 is thought to constitute the transmembrane pore, whereas the other components seem to play a role in unfolding the luggage proteins or providing a driving force. However, detailed functional and structural characterizations of PTEX proteins have not been performed. In this study, we expressed and characterized the membrane-associated component EXP2. Because expression of EXP2 is lethal to E. coli, EXP2 was expressed as a fusion protein with GST, and the recombinant EXP2 was obtained by protease digestion. The recombinant EXP2 formed pores in bilayer lipid membranes. The inner diameter of the pore was estimated to be approximately 3.5 nm based on electron microscopy images and channel currents. From this size and the molecular mass as determined by size exclusion chromatography and blue native polyacrylamide gel electrophoresis, we determined that the pore comprises approximately 10-12 EXP2 subunits. However, there is a possibility that the pore structure is different in the PTEX complex. These results provide important insights in the protein transport mechanism of PTEX, which will aid in developing new drugs targeting PTEX. Copyright © 2016 Elsevier Inc. All rights reserved.

Feasibility study for the construction of a new LNG receiving terminal, turkey. Volume 2. Appendix. Export trade information. [LNG (liquified natural gas)

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

Not Available

1993-06-01

The report was prepared by The M. W. Kellogg Co. for BOTAS Petroleum Pipeline Corporation of Ankara, Turkey. The study was undertaken to evaluate the cost and economics of constructing a second liquified natural gas (LNG) terminal in Turkey to meet future requirements for natural gas. Volume 2 contains the following appendices: LNG Storage Tanks; Vaporizers; Compressors; Pumps; Loading Arms; Marine Installations; Shipping; and Seismic Study.

Longtime variation of phytoplankton in the South China Sea from the perspective of carbon fixation

NASA Astrophysics Data System (ADS)

Li, Teng; Bai, Yan; Chen, Xiaoyan; Zhu, Qiankun; Gong, Fang; Wang, Difeng

2017-10-01

The ocean is a huge carbon pool in the earth, and about half of the anthropogenic emissions of carbon dioxide are absorbed by the ocean each year. By converting inorganic carbon into organic carbon, the photosynthesis process of phytoplankton affords an important way for carbon sequestration in the ocean. According to previous researches, primary production (NPP) and the structure of phytoplankton community are important in regulate the efficiency of biological carbon pump. This study examined the spatiotemporal variability of satellite remote sensing derived chlorophyll a concentration (Chla), phytoplankton carbon biomass (Carbon), composition ratio of micro-, nano- and pico- phytoplankton, NPP and integrated particulate organic carbon (IPOC) during 1998-2007 in the South China Sea (SCS). Micro-, nano-phytoplankton and NPP showed similar seasonal variation with highest values in winter (January) (especially in the western ocean of Luzon Strait) and lowest values in summer (July) in SCS. Chla, phytoplankton carbon biomass, and IPOC showed different seasonal trends with one peak values occurred in winter and lowest in spring. Two sampling areas (A, N:17-21°, E:117.5-120° and B, N:12.5-15°, E:112-119°) in SCS were selected based on spatial distribution of the standard deviation of research parameters mentioned above. Compared to Chla, phytoplankton carbon biomass, NPP and IPOC, the interannual changes of phytoplankton community structure were remarkable in the two areas. The fraction of micro- and nano- phytoplankton in SCS tend to rise when La Nina events occur. Our results contribute to an understanding of the response of phytoplankton to climate change in the marginal sea. To quantify the efficiency of biological carbon pump in this area, more attention should be paid to the development of remote sensing algorithms of export NPP (or POC export flux) as well as the regulate mechanism of export NPP.

Zooplankton diel vertical migration and contribution to deep active carbon flux in the NW Mediterranean

NASA Astrophysics Data System (ADS)

Isla, Alejandro; Scharek, Renate; Latasa, Mikel

2015-03-01

The diel vertical migration (DVM) of zooplankton contributes to the biological pump transporting material from surface to deep waters. We examined the DVM of the zooplankton community in different size fractions (53-200 μm, 200-500 μm, 500-1000 μm, 1000-2000 μm and > 2000 μm) during three cruises carried out in the open NW Mediterranean Sea. We assessed their metabolic rates from empirical published relationships and estimated the active fluxes of dissolved carbon to the mesopelagic zone driven by migrant zooplankton. Within the predominantly oligotrophic Mediterranean Sea, the NW region is one of the most productive ones, with a seasonal cycle characterized by a prominent spring bloom. The study area was visited at three different phases of the seasonal cycle: during the spring bloom, the post-bloom, and strongly stratified oligotrophic conditions. We found seasonal differences in DVM, less evident during the bloom. Changes in DVM intensity were related to the composition of the zooplanktonic assemblage, which also varied between cruises. Euphausiids appeared as the most active migrants in all seasons, and their life cycle conditioned the observed pattern. Immature stages, which are unable to perform large diel vertical movements, dominated during the bloom, in contrast to the higher relative importance of migrating adults in the other two sampling periods. The amount of dissolved carbon exported was determined by the migrant zooplankton biomass, being highest during the post-bloom (2.2 mmol C respired m- 2 d- 1, and up to 3.1 mmol C exported m- 2 d- 1 when DOC release estimations are added). The active transport by diel migrants represented a substantial contribution to total carbon export to deep waters, especially under stratified oligotrophic conditions, revealing the importance of zooplankton in the biological pump operating in the study area.

A quasi-lentiviral green fluorescent protein reporter exhibits nuclear export features of late human immunodeficiency virus type 1 transcripts

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

Graf, Marcus; Ludwig, Christine; Kehlenbeck, Sylvia

2006-09-01

We have previously shown that Rev-dependent expression of HIV-1 Gag from CMV immediate early promoter critically depends on the AU-rich codon bias of the gag gene. Here, we demonstrate that adaptation of the green fluorescent protein (GFP) reporter gene to HIV codon bias is sufficient to turn this hivGFP RNA into a quasi-lentiviral message following the rules of late lentiviral gene expression. Accordingly, GFP expression was significantly decreased in transfected cells strictly correlating with reduced RNA levels. In the presence of the HIV 5' major splice donor, the hivGFP RNAs were stabilized in the nucleus and efficiently exported to themore » cytoplasm following fusion of the 3' Rev-responsive element (RRE) and coexpression of HIV-1 Rev. This Rev-dependent translocation was specifically inhibited by leptomycin B suggesting export via the CRM1-dependent pathway used by late lentiviral transcripts. In conclusion, this quasi-lentiviral reporter system may provide a new platform for developing sensitive Rev screening assays.« less

Efflux Pump Gene Expression in Multidrug-Resistant Mycobacterium tuberculosis Clinical Isolates

PubMed Central

Jiang, Yi; Wei, Jianhao; Zhao, Li-li; Zhao, Xiuqin; Lu, Jianxin; Wan, Kanglin

2015-01-01

Isoniazid (INH) and rifampicin (RIF) are the two most effective drugs in tuberculosis therapy. Understanding the molecular mechanisms of resistance to these two drugs is essential to quickly diagnose multidrug-resistant (MDR) tuberculosis and extensive drug-resistant tuberculosis. Nine clinical Mycobacterium tuberculosis isolates resistant to only INH and RIF and 10 clinical pan-sensitive isolates were included to evaluate the expression of 20 putative drug efflux pump genes and sequence mutations in rpoB (RIF), katG (INH), the inhA promoter (INH), and oxyR-ahpC (INH). Nine and three MDR isolates were induced to overexpress efflux pump genes by INH and RIF, respectively. Eight and two efflux pump genes were induced to overexpress by INH and RIF in MDR isolates, respectively. drrA, drrB, efpA, jefA (Rv2459), mmr, Rv0849, Rv1634, and Rv1250 were overexpressed under INH or RIF stress. Most efflux pump genes were overexpressed under INH stress in a MDR isolates that carried the wild-type katG, inhA, and oxyR-ahpC associated with INH resistance than in those that carried mutations. The expression levels of 11 genes (efpA, Rv0849, Rv1250, P55 (Rv1410c), Rv1634, Rv2994, stp, Rv2459, pstB, drrA, and drrB) without drug inducement were significantly higher (P < 0.05) in nine MDR isolates than in 10 pan-sensitive isolates. In conclusion, efflux pumps may play an important role in INH acquired resistance in MDR M. tuberculosis, especially in those strains having no mutations in genes associated with INH resistance; basal expression levels of some efflux pump genes are higher in MDR isolates than in pan-sensitive isolates and the basal expressional differences may be helpful to diagnose and treat resistant tuberculosis. PMID:25695504

Integration of mRNP formation and export.

PubMed

Björk, Petra; Wieslander, Lars

2017-08-01

Expression of protein-coding genes in eukaryotes relies on the coordinated action of many sophisticated molecular machineries. Transcription produces precursor mRNAs (pre-mRNAs) and the active gene provides an environment in which the pre-mRNAs are processed, folded, and assembled into RNA-protein (RNP) complexes. The dynamic pre-mRNPs incorporate the growing transcript, proteins, and the processing machineries, as well as the specific protein marks left after processing that are essential for export and the cytoplasmic fate of the mRNPs. After release from the gene, the mRNPs move by diffusion within the interchromatin compartment, making up pools of mRNPs. Here, splicing and polyadenylation can be completed and the mRNPs recruit the major export receptor NXF1. Export competent mRNPs interact with the nuclear pore complex, leading to export, concomitant with compositional and conformational changes of the mRNPs. We summarize the integrated nuclear processes involved in the formation and export of mRNPs.

The simulated climate of the Last Glacial Maximum and insights into the global marine carbon cycle

NASA Astrophysics Data System (ADS)

Buchanan, Pearse J.; Matear, Richard J.; Lenton, Andrew; Phipps, Steven J.; Chase, Zanna; Etheridge, David M.

2016-12-01

The ocean's ability to store large quantities of carbon, combined with the millennial longevity over which this reservoir is overturned, has implicated the ocean as a key driver of glacial-interglacial climates. However, the combination of processes that cause an accumulation of carbon within the ocean during glacial periods is still under debate. Here we present simulations of the Last Glacial Maximum (LGM) using the CSIRO Mk3L-COAL (Carbon-Ocean-Atmosphere-Land) earth system model to test the contribution of physical and biogeochemical processes to ocean carbon storage. For the LGM simulation, we find a significant global cooling of the surface ocean (3.2 °C) and the expansion of both minimum and maximum sea ice cover broadly consistent with proxy reconstructions. The glacial ocean stores an additional 267 Pg C in the deep ocean relative to the pre-industrial (PI) simulation due to stronger Antarctic Bottom Water formation. However, 889 Pg C is lost from the upper ocean via equilibration with a lower atmospheric CO2 concentration and a global decrease in export production, causing a net loss of carbon relative to the PI ocean. The LGM deep ocean also experiences an oxygenation ( > 100 mmol O2 m-3) and deepening of the calcite saturation horizon (exceeds the ocean bottom) at odds with proxy reconstructions. With modifications to key biogeochemical processes, which include an increased export of organic matter due to a simulated release from iron limitation, a deepening of remineralisation and decreased inorganic carbon export driven by cooler temperatures, we find that the carbon content of the glacial ocean can be sufficiently increased (317 Pg C) to explain the reduction in atmospheric and terrestrial carbon at the LGM (194 ± 2 and 330 ± 400 Pg C, respectively). Assuming an LGM-PI difference of 95 ppm pCO2, we find that 55 ppm can be attributed to the biological pump, 28 ppm to circulation changes and the remaining 12 ppm to solubility. The biogeochemical modifications also improve model-proxy agreement in export production, carbonate chemistry and dissolved oxygen fields. Thus, we find strong evidence that variations in the oceanic biological pump exert a primary control on the climate.

Inactivation of a putative efflux pump (LmrB) in Streptococcus mutans results in altered biofilm structure and increased exopolysaccharide synthesis: implications for biofilm resistance.

PubMed

Liu, Jia; Zhang, Jianying; Guo, Lihong; Zhao, Wei; Hu, Xiaoli; Wei, Xi

2017-07-01

Efflux pumps are a mechanism associated with biofilm formation and resistance. There is limited information regarding efflux pumps in Streptococcus mutans, a major pathogen in dental caries. The aim of this study was to investigate potential roles of a putative efflux pump (LmrB) in S. mutans biofilm formation and susceptibility. Upon lmrB inactivation and antimicrobial exposure, the biofilm structure and expression of other efflux pumps were examined using confocal laser scanning microscopy (CLSM) and qRT-PCR. lmrB inactivation resulted in biofilm structural changes, increased EPS formation and EPS-related gene transcription (p < 0.05), but no improvement in susceptibility was observed. The expression of most efflux pump genes increased upon lmrB inactivation when exposed to antimicrobials (p < 0.05), suggesting a feedback mechanism that activated the transcription of other efflux pumps to compensate for the loss of lmrB. These observations imply that sole inactivation of lmrB is not an effective solution to control biofilms.

Amazon River carbon dioxide outgassing fuelled by wetlands.

PubMed

Abril, Gwenaël; Martinez, Jean-Michel; Artigas, L Felipe; Moreira-Turcq, Patricia; Benedetti, Marc F; Vidal, Luciana; Meziane, Tarik; Kim, Jung-Hyun; Bernardes, Marcelo C; Savoye, Nicolas; Deborde, Jonathan; Souza, Edivaldo Lima; Albéric, Patrick; Landim de Souza, Marcelo F; Roland, Fabio

2014-01-16

River systems connect the terrestrial biosphere, the atmosphere and the ocean in the global carbon cycle. A recent estimate suggests that up to 3 petagrams of carbon per year could be emitted as carbon dioxide (CO2) from global inland waters, offsetting the carbon uptake by terrestrial ecosystems. It is generally assumed that inland waters emit carbon that has been previously fixed upstream by land plant photosynthesis, then transferred to soils, and subsequently transported downstream in run-off. But at the scale of entire drainage basins, the lateral carbon fluxes carried by small rivers upstream do not account for all of the CO2 emitted from inundated areas downstream. Three-quarters of the world's flooded land consists of temporary wetlands, but the contribution of these productive ecosystems to the inland water carbon budget has been largely overlooked. Here we show that wetlands pump large amounts of atmospheric CO2 into river waters in the floodplains of the central Amazon. Flooded forests and floating vegetation export large amounts of carbon to river waters and the dissolved CO2 can be transported dozens to hundreds of kilometres downstream before being emitted. We estimate that Amazonian wetlands export half of their gross primary production to river waters as dissolved CO2 and organic carbon, compared with only a few per cent of gross primary production exported in upland (not flooded) ecosystems. Moreover, we suggest that wetland carbon export is potentially large enough to account for at least the 0.21 petagrams of carbon emitted per year as CO2 from the central Amazon River and its floodplains. Global carbon budgets should explicitly address temporary or vegetated flooded areas, because these ecosystems combine high aerial primary production with large, fast carbon export, potentially supporting a substantial fraction of CO2 evasion from inland waters.

Estimates of Gelatinous Zooplankton Carbon Flux in the Global Oceans

NASA Astrophysics Data System (ADS)

Luo, J. Y.; Condon, R.; Cowen, R. K.

2016-02-01

Gelatinous zooplankton (GZ), which include the cnidarians, ctenophores, and pelagic tunicates, are a common feature of marine ecosystems worldwide, but their contribution to global biogeochemical fluxes has never been assessed. We constructed a carbon-cycle model with a single, annual time-step and resolved to a 5° spatial grid for the three major GZ groups in order to evaluate the GZ-mediated carbon fluxes and export to depth. Biomass inputs (totaling 0.149 Pg C) were based off of Lucas et al. (2014) and updated using the JeDI database (Condon et al. 2015). From the upper ocean, biomass export flux from cnidarians, ctenophores, and tunicates totaled 2.96 ± 2.82 Pg C y-1, though only 0.199 ± 0.023 Pg C y-1 of GZ carbon were transferred to upper trophic levels, roughly amounting to one-quarter of all mesozooplankton production flux. In contrast, GZ fluxes to DOC only comprised ca. 2% of labile DOC flux. Egestion flux from the upper ocean totaled 2.56 ± 3.35 Pg C y-1, with over 80% being fast-sinking tunicate fecal pellets. Due to fast sinking rates of carcasses and fecal pellets, 26% of all C export from the upper ocean reached the seafloor, such that GZ fecal matter is estimated to comprise between 20-30% of global POC surface export and 11-30% of POC seafloor deposition. Finally, results from sensitivity analyses showed no increase in cnidarian and ctenophore export fluxes with increased temperature and jelly biomass, though tunicate export fluxes showed some increase with both temperature and biomass. These results suggest that current estimates of global POC flux from the surface oceans, which range between 8.6 - 12.9 Pg C y-1, may be underestimated by as much as 20 - 25%, implying a definite need to incorporate GZ mediated flux in estimating the biological pump transfer efficiency. Our study represents the first effort to quantify the role of gelatinous zooplankton in the global marine carbon cycle.

Dual function of the nuclear export signal of the Borna disease virus nucleoprotein in nuclear export activity and binding to viral phosphoprotein.

PubMed

Yanai, Mako; Sakai, Madoka; Makino, Akiko; Tomonaga, Keizo

2017-07-11

Borna disease virus (BoDV), which has a negative-sense, single-stranded RNA genome, causes persistent infection in the cell nucleus. The nuclear export signal (NES) of the viral nucleoprotein (N) consisting of leucine at positions 128 and 131 and isoleucine at positions 133 and 136 overlaps with one of two predicted binding sites for the viral phosphoprotein (P). A previous study demonstrated that higher expression of BoDV-P inhibits nuclear export of N; however, the function of N NES in the interaction with P remains unclear. We examined the subcellular localization, viral polymerase activity, and P-binding ability of BoDV-N NES mutants. We also characterized a recombinant BoDV (rBoDV) harboring an NES mutation of N. BoDV-N with four alanine-substitutions in the leucine and isoleucine residues of the NES impaired its cytoplasmic localization and abolished polymerase activity and P-binding ability. Although an alanine-substitution at position 131 markedly enhanced viral polymerase activity as determined by a minigenome assay, rBoDV harboring this mutation showed expression of viral RNAs and proteins relative to that of wild-type rBoDV. Our results demonstrate that BoDV-N NES has a dual function in BoDV replication, i.e., nuclear export of N and an interaction with P, affecting viral polymerase activity in the nucleus.

[Development of new therapeutic strategy for transporter-related diseases].

PubMed

Hayashi, Hisamitsu

2014-01-01

Significant technological advances in gene sequence analysis and construction of genetically-modified animals during the last two decades made it possible to reveal that many transporters are associated with diseases. The bile salt export pump (BSEP/ABCB11), a member of the family of ATP-binding cassette transporters, is localized on the canalicular membrane of hepatocytes and predominantly mediates the biliary excretion of bile salts. A hereditary defect of BSEP results in severe cholestasis called progressive familial intrahepatic cholestasis type 2 (PFIC2). Without liver transplantation, this disease progresses to liver failure and death before adulthood; therefore the development of new, less invasive medical therapy for PFIC2 is of the highest priority. We have previously shown that in many cases of PFIC2 patients, the dysfunction of BSEP is attributable to decreased BSEP expression on the hepatocanalicular membrane and that 4-phenylbutyrate (4PB), an approved drug for urea cycle disorder, may be a compound with potential to restore BSEP expression. This drug inhibits ubiquitination of cell surface-resident BSEP and thereby its clathrin-mediated endocytosis through the AP2 adaptor complex, leading to increase in BSEP expression on the canalicular membrane. Clinical studies to investigate the efficacy of 4PB in the treatment of PFIC2 revealed that 4PB therapy biochemically and histologically improved liver function without any side effect. Therefore, 4PB therapy may become the preferred choice, instead of liver transplantation, for PFIC2 patients. The strategy employed and findings in this study would be valuable for the drug development of transporter-related disorders.

The inhibition of hepatic bile acids transporters Ntcp and Bsep is involved in the pathogenesis of isoniazid/rifampicin-induced hepatotoxicity.

PubMed

Guo, Yao Xue; Xu, Xue Fei; Zhang, Qi Zhi; Li, Chun; Deng, Ye; Jiang, Pei; He, Lei Yan; Peng, Wen Xing

2015-01-01

Co-treatment of isoniazid (INH) and rifampicin (RFP) is well known for clinically apparent liver injury. However, the mechanism of INH/RFP-induced liver injury is controversial. Emerging evidence shows links between inhibition of bile acids transporters and drug-induced liver injury (DILI). The present study investigates whether sodium taurocholate cotransporting polypeptide (NTCP/Ntcp; SLC10A1) and bile salt export pump (BSEP/Bsep; ABCB11) are involved in the anti-tuberculosis medicines induced liver injury. ICR female mice were intragastrically treated with INH (50 or 100 mg/kg), RFP (100 or 200 mg/kg), or the combination of INH/RFP (50 + 100 mg/kg or 100 + 200 mg/kg) for 14 consecutive days. Liver histopathological examination, serum biochemical and liver malondialdehyde tests were evaluated. Apparent histopathological alterations and hepatic oxidative stress showed in INH (100 mg/kg), RFP (200 mg/kg) and their combination group. The hepatoxic effect was also indicated by increased serum biomarkers, such as aspartate transaminase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), direct bilirubin (DBil), total bilirubin (TBil) and total bile acids (TBA). Both doses of INH/RFP administration significantly down-regulated the expression of Ntcp and Bsep in liver. Furthermore, the combination of INH and RFP displayed stronger effect on the expression of Ntcp compared with the corresponding dose of INH or RFP alone. In conclusion, down-regulated expression of hepatic Ntcp and Bsep might play an important role in the development of INH and RFP induced liver injury.

Protein Kinase D-dependent Phosphorylation and Nuclear Export of Histone Deacetylase 5 Mediates Vascular Endothelial Growth Factor-induced Gene Expression and Angiogenesis*S⃞

PubMed Central

Ha, Chang Hoon; Wang, Weiye; Jhun, Bong Sook; Wong, Chelsea; Hausser, Angelika; Pfizenmaier, Klaus; McKinsey, Timothy A.; Olson, Eric N.; Jin, Zheng-Gen

2008-01-01

Vascular endothelial growth factor (VEGF) is essential for normal and pathological angiogenesis. However, the signaling pathways linked to gene regulation in VEGF-induced angiogenesis are not fully understood. Here we demonstrate a critical role of protein kinase D (PKD) and histone deacetylase 5 (HDAC5) in VEGF-induced gene expression and angiogenesis. We found that VEGF stimulated HDAC5 phosphorylation and nuclear export in endothelial cells through a VEGF receptor 2-phospholipase Cγ-protein kinase C-PKD-dependent pathway. We further showed that the PKD-HDAC5 pathway mediated myocyte enhancer factor-2 transcriptional activation and a specific subset of gene expression in response to VEGF, including NR4A1, an orphan nuclear receptor involved in angiogenesis. Specifically, inhibition of PKD by overexpression of the PKD kinase-negative mutant prevents VEGF-induced HDAC5 phosphorylation and nuclear export as well as NR4A1 induction. Moreover, a mutant of HDAC5 specifically deficient in PKD-dependent phosphorylation inhibited VEGF-mediated NR4A1 expression, endothelial cell migration, and in vitro angiogenesis. These findings suggest that the PKD-HDAC5 pathway plays an important role in VEGF regulation of gene transcription and angiogenesis. PMID:18332134

Effects of flow diversions on water and habitat quality: Examples from California's highly manipulated Sacramento–San Joaquin Delta

USGS Publications Warehouse

Monsen, Nancy E.; Cloern, James E.; Burau, Jon R.

2007-01-01

We use selected monitoring data to illustrate how localized water diversions from seasonal barriers, gate operations, and export pumps alter water quality across the Sacramento-San Joaquin Delta (California). Dynamics of water-quality variability are complex because the Delta is a mixing zone of water from the Sacramento and San Joaquin Rivers, agricultural return water, and the San Francisco Estuary. Each source has distinct water-quality characteristics, and the contribution of each source varies in response to natural hydrologic variability and water diversions. We use simulations with a tidal hydrodynamic model to reveal how three diversion events, as case studies, influence water quality through their alteration of Delta-wide water circulation patterns and flushing time. Reduction of export pumping decreases the proportion of Sacramento- to San Joaquin-derived fresh water in the central Delta, leading to rapid increases in salinity. Delta Cross Channel gate operations control salinity in the western Delta and alter the freshwater source distribution in the central Delta. Removal of the head of Old River barrier, in autumn, increases the flushing time of the Stockton Ship Channel from days to weeks, contributing to a depletion of dissolved oxygen. Each shift in water quality has implications either for habitat quality or municipal drinking water, illustrating the importance of a systems view to anticipate the suite of changes induced by flow manipulations, and to minimize the conflicts inherent in allocations of scarce resources to meet multiple objectives.

Breast pump use amongst mothers of healthy term infants in Melbourne, Australia: A prospective cohort study.

PubMed

Johns, Helene M; Amir, Lisa H; McLachlan, Helen L; Forster, Della A

2016-02-01

this paper describes the use of breast pumps amongst a group of mothers of healthy term infants in Melbourne, Australia, between birth and six months post partum. a prospective cohort study; data were collected using structured questionnaires. 1003 postpartum women who had given birth to healthy infants at term were recruited from three Melbourne maternity hospitals between July 2009 and April 2011. Data were collected by face-to-face interview at recruitment and by telephone interview three and six months later. at recruitment (24 to 48 hours post partum), 60% (605/1003) of women already had a breast pump. At two weeks post partum expressing was common; 62% (290/466) of women had expressed by this time, with 40% (186/466) doing so several times a day. By six months post partum 83% (754/911) of the women had a breast pump and 40% (288/715) were expressing, although most just occasionally. The most common reasons for any expressing in the first six months were 'to be able to go out and leave the baby' (35%; 268/772); milk supply 'not enough'(27%; 207/772); and having 'too much' milk (19%; 147/772). The increasing popularity of expressing breast milk to feed infants is not driven by women returning to the workforce, as only 10% of women (80/772) expressed because they had returned to paid employment. health professionals should be aware that in some settings breast pump use is common in the first six months, and this is not always related to maternal workforce participation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Gene expression analysis of the SdeAB multidrug efflux pump in antibiotic-resistant clinical isolates of Serratia marcescens.

PubMed

Dalvi, S D; Worobec, E A

2012-01-01

Many isolates of Serratia marcescens, a well-known opportunistic pathogen, can be multidrug resistant. Fluoroquinolones are among the most important groups of antibiotics used for treatment of these organisms. However, fluoroquinolone resistance among S. marcescens isolates is fast increasing. Drug extrusion through efflux pumps like SdeAB/ HasF is one of the major mechanisms of resistance to fluoroquinolones. This study was carried out to analyze, through gene expression analysis of sdeB, the relative contribution of this mechanism toward fluoroquinolone resistance in clinical isolates of Serratia. Total RNA from 45 clinical isolates of S. marcescens was isolated. Quantitative real-time RT PCR was performed on the extracted RNA to study the gene expression of sdeB and was normalized to the sdeB expression in the standard strain of S. marcescens. Of the 45 isolates analyzed, sdeB expression was found to be elevated in 20 isolates (44%). Of these 20 isolates, eight (40%) were fully resistant to at least one of the fluoroquinolones studied. Conversely, of the 20 isolates that over-expressed sdeB, 12 (60%) were fully sensitive to all fluoroquinolones tested. Drug efflux pumps are an important means of fluoroquinolone resistance among clinically important species of Serratia. The expression of these pumps can be up-regulated in the presence of antibiotics and have the potential for changing the phenotype from sensitive to resistant, thus contributing to therapeutic failures.

UAP56 is a conserved crucial component of a divergent mRNA export pathway in Toxoplasma gondii.

PubMed

Serpeloni, Mariana; Jiménez-Ruiz, Elena; Vidal, Newton Medeiros; Kroeber, Constanze; Andenmatten, Nicole; Lemgruber, Leandro; Mörking, Patricia; Pall, Gurman S; Meissner, Markus; Ávila, Andréa R

2016-11-01

Nucleo-cytoplasmic RNA export is an essential post-transcriptional step to control gene expression in eukaryotic cells and is poorly understood in apicomplexan parasites. With the exception of UAP56, a component of TREX (Transcription Export) complex, other components of mRNA export machinery are not well conserved in divergent supergroups. Here, we use Toxoplasma gondii as a model system to functionally characterize TgUAP56 and its potential interaction factors. We demonstrate that TgUAP56 is crucial for mRNA export and that functional interference leads to significant accumulation of mRNA in the nucleus. It was necessary to employ bioinformatics and phylogenetic analysis to identify orthologs related to mRNA export, which show a remarkable low level of conservation in T. gondii. We adapted a conditional Cas9/CRISPR system to carry out a genetic screen to verify if these factors were involved in mRNA export in T. gondii. Only the disruption of TgRRM_1330 caused accumulation of mRNA in the nucleus as found with TgUAP56. This protein is potentially a divergent partner of TgUAP56, and provides insight into a divergent mRNA export pathway in apicomplexans. © 2016 The Authors. Molecular Microbiology Published by John Wiley & Sons Ltd.

UAP56 is a conserved crucial component of a divergent mRNA export pathway in Toxoplasma gondii

PubMed Central

Serpeloni, Mariana; Jiménez‐Ruiz, Elena; Vidal, Newton Medeiros; Kroeber, Constanze; Andenmatten, Nicole; Lemgruber, Leandro; Mörking, Patricia; Pall, Gurman S.

2016-01-01

Summary Nucleo‐cytoplasmic RNA export is an essential post‐transcriptional step to control gene expression in eukaryotic cells and is poorly understood in apicomplexan parasites. With the exception of UAP56, a component of TREX (Transcription Export) complex, other components of mRNA export machinery are not well conserved in divergent supergroups. Here, we use Toxoplasma gondii as a model system to functionally characterize TgUAP56 and its potential interaction factors. We demonstrate that TgUAP56 is crucial for mRNA export and that functional interference leads to significant accumulation of mRNA in the nucleus. It was necessary to employ bioinformatics and phylogenetic analysis to identify orthologs related to mRNA export, which show a remarkable low level of conservation in T. gondii. We adapted a conditional Cas9/CRISPR system to carry out a genetic screen to verify if these factors were involved in mRNA export in T. gondii. Only the disruption of TgRRM_1330 caused accumulation of mRNA in the nucleus as found with TgUAP56. This protein is potentially a divergent partner of TgUAP56, and provides insight into a divergent mRNA export pathway in apicomplexans. PMID:27542978

Xenobiotic, Bile Acid, and Cholesterol Transporters: Function and Regulation

PubMed Central

Aleksunes, Lauren M.

2010-01-01

Transporters influence the disposition of chemicals within the body by participating in absorption, distribution, and elimination. Transporters of the solute carrier family (SLC) comprise a variety of proteins, including organic cation transporters (OCT) 1 to 3, organic cation/carnitine transporters (OCTN) 1 to 3, organic anion transporters (OAT) 1 to 7, various organic anion transporting polypeptide isoforms, sodium taurocholate cotransporting polypeptide, apical sodium-dependent bile acid transporter, peptide transporters (PEPT) 1 and 2, concentrative nucleoside transporters (CNT) 1 to 3, equilibrative nucleoside transporter (ENT) 1 to 3, and multidrug and toxin extrusion transporters (MATE) 1 and 2, which mediate the uptake (except MATEs) of organic anions and cations as well as peptides and nucleosides. Efflux transporters of the ATP-binding cassette superfamily, such as ATP-binding cassette transporter A1 (ABCA1), multidrug resistance proteins (MDR) 1 and 2, bile salt export pump, multidrug resistance-associated proteins (MRP) 1 to 9, breast cancer resistance protein, and ATP-binding cassette subfamily G members 5 and 8, are responsible for the unidirectional export of endogenous and exogenous substances. Other efflux transporters [ATPase copper-transporting β polypeptide (ATP7B) and ATPase class I type 8B member 1 (ATP8B1) as well as organic solute transporters (OST) α and β] also play major roles in the transport of some endogenous chemicals across biological membranes. This review article provides a comprehensive overview of these transporters (both rodent and human) with regard to tissue distribution, subcellular localization, and substrate preferences. Because uptake and efflux transporters are expressed in multiple cell types, the roles of transporters in a variety of tissues, including the liver, kidneys, intestine, brain, heart, placenta, mammary glands, immune cells, and testes are discussed. Attention is also placed upon a variety of regulatory factors that influence transporter expression and function, including transcriptional activation and post-translational modifications as well as subcellular trafficking. Sex differences, ontogeny, and pharmacological and toxicological regulation of transporters are also addressed. Transporters are important transmembrane proteins that mediate the cellular entry and exit of a wide range of substrates throughout the body and thereby play important roles in human physiology, pharmacology, pathology, and toxicology. PMID:20103563

Plasmid-Encoded Tetracycline Efflux Pump Protein Alters Bacterial Stress Responses and Ecological Fitness of Acinetobacter oleivorans

PubMed Central

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment. PMID:25229538

Plasmid-encoded tetracycline efflux pump protein alters bacterial stress responses and ecological fitness of Acinetobacter oleivorans.

PubMed

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment.

The Novel Mouse Mutation Oblivion Inactivates the PMCA2 Pump and Causes Progressive Hearing Loss

PubMed Central

de Angelis, Martin Hrabé; Fuchs, Helmut; Lim, Dmitry; Ortolano, Saida; Ingham, Neil J.; Brini, Marisa; Carafoli, Ernesto; Mammano, Fabio; Steel, Karen P.

2008-01-01

Progressive hearing loss is common in the human population, but we have few clues to the molecular basis. Mouse mutants with progressive hearing loss offer valuable insights, and ENU (N-ethyl-N-nitrosourea) mutagenesis is a useful way of generating models. We have characterised a new ENU-induced mouse mutant, Oblivion (allele symbol Obl), showing semi-dominant inheritance of hearing impairment. Obl/+ mutants showed increasing hearing impairment from post-natal day (P)20 to P90, and loss of auditory function was followed by a corresponding base to apex progression of hair cell degeneration. Obl/Obl mutants were small, showed severe vestibular dysfunction by 2 weeks of age, and were completely deaf from birth; sensory hair cells were completely degenerate in the basal turn of the cochlea, although hair cells appeared normal in the apex. We mapped the mutation to Chromosome 6. Mutation analysis of Atp2b2 showed a missense mutation (2630C→T) in exon 15, causing a serine to phenylalanine substitution (S877F) in transmembrane domain 6 of the PMCA2 pump, the resident Ca2+ pump of hair cell stereocilia. Transmembrane domain mutations in these pumps generally are believed to be incompatible with normal targeting of the protein to the plasma membrane. However, analyses of hair cells in cultured utricular maculae of Obl/Obl mice and of the mutant Obl pump in model cells showed that the protein was correctly targeted to the plasma membrane. Biochemical and biophysical characterisation showed that the pump had lost a significant portion of its non-stimulated Ca2+ exporting ability. These findings can explain the progressive loss of auditory function, and indicate the limits in our ability to predict mechanism from sequence alone. PMID:18974863

The role of TREX in gene expression and disease

PubMed Central

Heath, Catherine G.; Viphakone, Nicolas; Wilson, Stuart A.

2016-01-01

TRanscription and EXport (TREX) is a conserved multisubunit complex essential for embryogenesis, organogenesis and cellular differentiation throughout life. By linking transcription, mRNA processing and export together, it exerts a physiologically vital role in the gene expression pathway. In addition, this complex prevents DNA damage and regulates the cell cycle by ensuring optimal gene expression. As the extent of TREX activity in viral infections, amyotrophic lateral sclerosis and cancer emerges, the need for a greater understanding of TREX function becomes evident. A complete elucidation of the composition, function and interactions of the complex will provide the framework for understanding the molecular basis for a variety of diseases. This review details the known composition of TREX, how it is regulated and its cellular functions with an emphasis on mammalian systems. PMID:27679854

Internal hydraulic loss in a seal-less centrifugal Gyro pump.

PubMed

Makinouchi, K; Ohara, Y; Sakuma, I; Damm, G; Mizuguchi, K; Jikuya, T; Takatani, S; Noon, G P; Nosé, Y

1994-01-01

A new index "loss factor Z" defined by Eq. 1 was introduced as the absolute expression of the mock loop resistance for testing a nonpulsatile pump. [formula: see text] where gamma is specific gravity of the fluid, g is the acceleration of gravity, delta P is total pressure head, and Q is flow. Z is expected to be constant, regardless of the pumping parameters. Z values obtained in the same mock loop but with different rotary blood pumps were almost identical and were defined as Z0. New methods of analysis of the flow-restrictive conditions of various rotary blood pumps are proposed in this paper: namely, differential loss factor delta Z, and loss factor sensitivity delta Z/delta A. The proposed Z-Q curves demonstrated better performance mapping than the conventional delta P-Q curves. Delta Z is the difference between the Z-Q curves of two different pumps. A is a design parameter of the pump; therefore delta Z/delta A is a quantitative expression of the effect of the design change on the hydraulic performance. These various indices were used to analyze the internal hydraulic loss of a centrifugal pump (Gyro pump). The relationship between its gap size (rotor casing) and hydraulic performance was assessed quantitatively by these indices. In this paper, the derivation processes and above-mentioned indices are described.

Regulation of mRNA Trafficking by Nuclear Pore Complexes

PubMed Central

Bonnet, Amandine; Palancade, Benoit

2014-01-01

Over the last two decades, multiple studies have explored the mechanisms governing mRNA export out of the nucleus, a crucial step in eukaryotic gene expression. During transcription and processing, mRNAs are assembled into messenger ribonucleoparticles (mRNPs). mRNPs are then exported through nuclear pore complexes (NPCs), which are large multiprotein assemblies made of several copies of a limited number of nucleoporins. A considerable effort has been put into the dissection of mRNA export through NPCs at both cellular and molecular levels, revealing the conserved contributions of a subset of nucleoporins in this process, from yeast to vertebrates. Several reports have also demonstrated the ability of NPCs to sort out properly-processed mRNPs for entry into the nuclear export pathway. Importantly, changes in mRNA export have been associated with post-translational modifications of nucleoporins or changes in NPC composition, depending on cell cycle progression, development or exposure to stress. How NPC modifications also impact on cellular mRNA export in disease situations, notably upon viral infection, is discussed. PMID:25184662

Protein kinase D1 phosphorylates HDAC7 and induces its nuclear export after T-cell receptor activation.

PubMed

Parra, Maribel; Kasler, Herbert; McKinsey, Timothy A; Olson, Eric N; Verdin, Eric

2005-04-08

HDAC7, a class II histone deacetylase that is highly expressed in thymocytes, inhibits both transcription of the orphan steroid nuclear receptor Nur77 and induction of apoptosis in response to activation of the T-cell receptor (TCR). Here, we report that HDAC7 is exported to the cytoplasm by a calcium-independent signaling pathway after TCR activation. Protein kinase D1 (PKD1) was activated after TCR engagement, interacted with HDAC7, and phosphorylated three serines (Ser155, Ser318, and Ser448) at its N terminus, leading to its export from the nucleus. Mutation of Ser155, Ser318, and Ser448 blocked the nucleocytoplasmic shuttling of HDAC7 in response to TCR activation, as did overexpression of a kinase-inactive form of PKD1. Consistent with the regulatory role of HDAC7 in Nur77 expression, PKD1 activation led to the transcriptional activation of Nur77 via myocyte enhancer factor 2-binding sites in its promoter. In a mouse model of negative selection, PKD1 was activated during thymocyte activation. These observations indicate that PKD1 regulates the expression of Nur77 during thymocyte activation at least in part by phosphorylating HDAC7.

Petroleum Prices: Past, Present, and Prospective,

DTIC Science & Technology

1987-01-01

other oil-exporting developing nations such as Mexico and Malaysia ) enjoyed relatively high export revenues and quickly adjusted their spending habits to... challenge Soviet oil production for first place in fuel-equivalent terms. Natural gas currently provides about 95 percent ansmuch energy as oil pro- duction...been expressed without challenge in many recent forecasts. By now, OPEC nations in general, and Persian Gulf producers in particular, appear to have

Functional reconstitution of Arabidopsis thaliana plant uncoupling mitochondrial protein (AtPUMP1) expressed in Escherichia coli.

PubMed

Borecký, J; Maia, I G; Costa, A D; Jezek, P; Chaimovich, H; de Andrade, P B; Vercesi, A E; Arruda, P

2001-09-14

The Arabidopsis thaliana uncoupling protein (UCP) gene was expressed in Escherichia coli and isolated protein reconstituted into liposomes. Linoleic acid-induced H+ fluxes were sensitive to purine nucleotide inhibition with an apparent K(i) (in mM) of 0.8 (GDP), 0.85 (ATP), 0.98 (GTP), and 1.41 (ADP); the inhibition was pH-dependent. Kinetics of AtPUMP1-mediated H+ fluxes were determined for lauric, myristic, palmitic, oleic, linoleic, and linolenic acids. Properties of recombinant AtPUMP1 indicate that it represents a plant counterpart of animal UCP2 or UCP3. This work brings the functional and genetic approaches together for the first time, providing strong support that AtPUMP1 is truly an UCP.

“It’s my blood”: ethical complexities in the use, storage and export of biological samples: perspectives from South African research participants

PubMed Central

2014-01-01

Background The use of biological samples in research raises a number of ethical issues in relation to consent, storage, export, benefit sharing and re-use of samples. Participant perspectives have been explored in North America and Europe, with only a few studies reported in Africa. The amount of research being conducted in Africa is growing exponentially with volumes of biological samples being exported from the African continent. In order to investigate the perspectives of African research participants, we conducted a study at research sites in the Western Cape and Gauteng, South Africa. Methods Data were collected using a semi-structured questionnaire that captured both quantitative and qualitative information at 6 research sites in South Africa. Interviews were conducted in English and Afrikaans. Data were analysed both quantitatively and qualitatively. Results Our study indicates that while the majority of participants were supportive of providing samples for research, serious concerns were voiced about future use, benefit sharing and export of samples. While researchers view the provision of biosamples as a donation, participants believe that they still have ownership rights and are therefore in favour of benefit sharing. Almost half of the participants expressed a desire to be re-contacted for consent for future use of their samples. Interesting opinions were expressed with respect to export of samples. Conclusions Eliciting participant perspectives is an important part of community engagement in research involving biological sample collection, export, storage and future use. A tiered consent process appears to be more acceptable to participants in this study. Eliciting opinions of researchers and research ethics committee (REC) members would contribute multiple perspectives. Further research is required to interrogate the concept of ownership and the consent process in research involving biological samples. PMID:24447822

Identification of a Novel System for Boron Transport: Atr1 Is a Main Boron Exporter in Yeast▿ †

PubMed Central

Kaya, Alaattin; Karakaya, Huseyin C.; Fomenko, Dmitri E.; Gladyshev, Vadim N.; Koc, Ahmet

2009-01-01

Boron is a micronutrient in plants and animals, but its specific roles in cellular processes are not known. To understand boron transport and functions, we screened a yeast genomic DNA library for genes that confer resistance to the element in Saccharomyces cerevisiae. Thirty boron-resistant transformants were isolated, and they all contained the ATR1 (YML116w) gene. Atr1 is a multidrug resistance transport protein belonging to the major facilitator superfamily. C-terminal green fluorescent protein-tagged Atr1 localized to the cell membrane and vacuole, and ATR1 gene expression was upregulated by boron and several stress conditions. We found that atr1Δ mutants were highly sensitive to boron treatment, whereas cells overexpressing ATR1 were boron resistant. In addition, atr1Δ cells accumulated boron, whereas ATR1-overexpressing cells had low intracellular levels of the element. Furthermore, atr1Δ cells showed stronger boron-dependent phenotypes than mutants deficient in genes previously reported to be implicated in boron metabolism. ATR1 is widely distributed in bacteria, archaea, and lower eukaryotes. Our data suggest that Atr1 functions as a boron efflux pump and is required for boron tolerance. PMID:19414602

The cytochrome bd oxidase of Escherichia coli prevents respiratory inhibition by endogenous and exogenous hydrogen sulfide

PubMed Central

Korshunov, Sergey; Imlay, Karin R. C.; Imlay, James A.

2016-01-01

Summary When sulfur compounds are scarce or difficult to process, E. coli adapts by inducing the high-level expression of sulfur-compound importers. If cystine then becomes available, the cystine is rapidly overimported and reduced, leading to a burgeoning pool of intracellular cysteine. Most of the excess cysteine is exported, but some is adventitiously degraded, with the consequent release of sulfide. Sulfide is a potent ligand of copper and heme moieties, raising the prospect that it interferes with enzymes. We observed that when cystine was provided and sulfide levels rose, E. coli became strictly dependent upon cytochrome bd oxidase for continued respiration. Inspection revealed that low-micromolar levels of sulfide inhibited the proton-pumping cytochrome bo oxidase that is regarded as the primary respiratory oxidase. In the absence of the back-up cytochrome bd oxidase, growth failed. Exogenous sulfide elicited the same effect. The potency of sulfide was enhanced when oxygen concentrations were low. Natural oxic-anoxic interfaces are often sulfidic, including the intestinal environment where E. coli dwells. We propose that the sulfide resistance of the cytochrome bd oxidase is a key trait that permits respiration in such habitats. PMID:26991114

Impact of charged amino acid substitution in the transmembrane domain of L-alanine exporter, AlaE, of Escherichia coli on the L-alanine export.

PubMed

Kim, Seryoung; Ihara, Kohei; Katsube, Satoshi; Ando, Tasuke; Isogai, Emiko; Yoneyama, Hiroshi

2017-01-01

The Escherichia coli alaE gene encodes the L-alanine exporter, AlaE, that catalyzes active export of L-alanine using proton electrochemical potential. The transporter comprises only 149 amino acid residues and four predicted transmembrane domains (TMs), which contain three charged amino acid residues. The AlaE-deficient L-alanine non-metabolizing cells (ΔalaE cells) appeared hypersusceptible to L-alanyl-L-alanine showing a minimum inhibitory concentration (MIC) of 2.5 µg/ml for the dipeptide due to a toxic accumulation of L-alanine. To elucidate the mechanism by which AlaE exports L-alanine, we replaced charged amino acid residues in the TMs, glutamic acid-30 (TM-I), arginine-45 (TM-II), and aspartic acid-84 (TM-III) with their respective charge-conserved amino acid or a net neutral cysteine. The ΔalaE cells producing R45K or R45C appeared hypersusceptible to the dipeptide, indicating that arginine-45 is essential for AlaE activity. MIC of the dipeptide in the ΔalaE cells expressing E30D and E30C was 156 µg/ml and >10,000 µg/ml, respectively, thereby suggesting that a negative charge at this position is not essential. The ΔalaE cells expressing D84E or D84C showed an MIC >10,000 and 78 µg/ml, respectively, implying that a negative charge is required at this position. These results were generally consistent with that of the L-alanine accumulation experiments in intact cells. We therefore concluded that charged amino acid residues (R45 and D84) in the AlaE transmembrane domain play a pivotal role in L-alanine export. Replacement of three cysteine residues at C22, C28 (both in TM-I), and C135 (C-terminal region) with alanine showed only a marginal effect on L-alanine export.

Cytoplasmic utilization of human immunodeficiency virus type 1 genomic RNA is not dependent on a nuclear interaction with gag.

PubMed

Grewe, Bastian; Hoffmann, Bianca; Ohs, Inga; Blissenbach, Maik; Brandt, Sabine; Tippler, Bettina; Grunwald, Thomas; Uberla, Klaus

2012-03-01

In some retroviruses, such as Rous sarcoma virus and prototype foamy virus, Gag proteins are known to shuttle between the nucleus and the cytoplasm and are implicated in nuclear export of the viral genomic unspliced RNA (gRNA) for subsequent encapsidation. A similar function has been proposed for human immunodeficiency virus type 1 (HIV-1) Gag based on the identification of nuclear localization and export signals. However, the ability of HIV-1 Gag to transit through the nucleus has never been confirmed. In addition, the lentiviral Rev protein promotes efficient nuclear gRNA export, and previous reports indicate a cytoplasmic interaction between Gag and gRNA. Therefore, functional effects of HIV-1 Gag on gRNA and its usage were explored. Expression of gag in the absence of Rev was not able to increase cytoplasmic gRNA levels of subgenomic, proviral, or lentiviral vector constructs, and gene expression from genomic reporter plasmids could not be induced by Gag provided in trans. Furthermore, Gag lacking the reported nuclear localization and export signals was still able to mediate an efficient packaging process. Although small amounts of Gag were detectable in the nuclei of transfected cells, a Crm1-dependent nuclear export signal in Gag could not be confirmed. Thus, our study does not provide any evidence for a nuclear function of HIV-1 Gag. The encapsidation process of HIV-1 therefore clearly differs from that of Rous sarcoma virus and prototype foamy virus.

Effect of ursodeoxycholic acid on bile acid profiles and intestinal detoxification machinery in primary biliary cirrhosis and health.

PubMed

Dilger, Karin; Hohenester, Simon; Winkler-Budenhofer, Ursula; Bastiaansen, Barbara A J; Schaap, Frank G; Rust, Christian; Beuers, Ulrich

2012-07-01

Ursodeoxycholic acid (UDCA) exerts anticholestatic, antifibrotic and antiproliferative effects in primary biliary cirrhosis (PBC) via mechanisms not yet fully understood. Its adequate biliary enrichment is considered mandatory for therapeutic efficacy. However, precise determination of biliary enrichment of UDCA is not possible in clinical practice. Therefore, we investigated (i) the relationship between biliary enrichment and plasma pharmacokinetics of UDCA, (ii) the effect of UDCA on plasma and biliary bile acid composition and conjugation patterns, and (iii) on the intestinal detoxification machinery in patients with PBC and healthy controls. In 11 PBC patients and 11 matched healthy subjects, cystic bile and duodenal tissue were collected before and after 3 weeks of administration of UDCA (15 mg/kg/day). Extensive pharmacokinetic profiling of bile acids was performed. The effect of UDCA on the intestinal detoxification machinery was studied by quantitative PCR and Western blotting. The relative fraction of UDCA and its conjugates in plasma at trough level[x] correlated with their biliary enrichment[y] (r=0.73, p=0.0001, y=3.65+0.49x). Taurine conjugates of the major hydrophobic bile acid, chenodeoxycholic acid, were more prominent in bile of PBC patients than in that of healthy controls. Biliary bile acid conjugation patterns normalized after treatment with UDCA. UDCA induced duodenal expression of key export pumps, BCRP and P-glycoprotein. Biliary and trough plasma enrichment of UDCA are closely correlated in PBC and health. Taurine conjugation may represent an adaptive mechanism in PBC against chenodeoxycholic acid-mediated bile duct damage. UDCA may stabilize small intestinal detoxification by upregulation of efflux pumps. Copyright © 2012. Published by Elsevier B.V.

Interplay between three RND efflux pumps in doxycycline-selected strains of Burkholderia thailandensis.

PubMed

Biot, Fabrice Vincent; Lopez, Mélanie Monique; Poyot, Thomas; Neulat-Ripoll, Fabienne; Lignon, Sabrina; Caclard, Arnaud; Thibault, François Michel; Peinnequin, Andre; Pagès, Jean-Marie; Valade, Eric

2013-01-01

Efflux systems are involved in multidrug resistance in most Gram-negative non-fermentative bacteria. We have chosen Burkholderia thailandensis to dissect the development of multidrug resistance phenotypes under antibiotic pressure. We used doxycycline selection to obtain several resistant B. thailandensis variants. The minimal inhibitory concentrations of a large panel of structurally unrelated antibiotics were determined ± the efflux pump inhibitor phenylalanine-arginine ß-naphthylamide (PAßN). Membrane proteins were identified by proteomic method and the expressions of major efflux pumps in the doxycycline selected variants were compared to those of the parental strains by a quantitative RT-PCR analysis. Doxycycline selected variants showed a multidrug resistance in two major levels corresponding to the overproduction of two efflux pumps depending on its concentration: AmrAB-OprA and BpeEF-OprC. The study of two mutants, each lacking one of these pumps, indicated that a third pump, BpeAB-OprB, could substitute for the defective pump. Surprisingly, we observed antagonistic effects between PAßN and aminoglycosides or some ß-lactams. PAßN induced the overexpression of AmrAB-OprA and BpeAB-OprB pump genes, generating this unexpected effect. These results may account for the weak activity of PAßN in some Gram-negative species. We clearly demonstrated two antagonistic effects of this molecule on bacterial cells: the blocking of antibiotic efflux and an increase in efflux pump gene expression. Thus, doxycycline is a very efficient RND efflux pump inducer and PAßN may promote the production of some efflux pumps. These results should be taken into account when considering antibiotic treatments and in future studies on efflux pump inhibitors.

Nuclear Export Signal Masking Regulates HIV-1 Rev Trafficking and Viral RNA Nuclear Export.

PubMed

Behrens, Ryan T; Aligeti, Mounavya; Pocock, Ginger M; Higgins, Christina A; Sherer, Nathan M

2017-02-01

HIV-1's Rev protein forms a homo-oligomeric adaptor complex linking viral RNAs to the cellular CRM1/Ran-GTP nuclear export machinery through the activity of Rev's prototypical leucine-rich nuclear export signal (NES). In this study, we used a functional fluorescently tagged Rev fusion protein as a platform to study the effects of modulating Rev NES identity, number, position, or strength on Rev subcellular trafficking, viral RNA nuclear export, and infectious virion production. We found that Rev activity was remarkably tolerant of diverse NES sequences, including supraphysiological NES (SNES) peptides that otherwise arrest CRM1 transport complexes at nuclear pores. Rev's ability to tolerate a SNES was both position and multimerization dependent, an observation consistent with a model wherein Rev self-association acts to transiently mask the NES peptide(s), thereby biasing Rev's trafficking into the nucleus. Combined imaging and functional assays also indicated that NES masking underpins Rev's well-known tendency to accumulate at the nucleolus, as well as Rev's capacity to activate optimal levels of late viral gene expression. We propose that Rev multimerization and NES masking regulates Rev's trafficking to and retention within the nucleus even prior to RNA binding. HIV-1 infects more than 34 million people worldwide causing >1 million deaths per year. Infectious virion production is activated by the essential viral Rev protein that mediates nuclear export of intron-bearing late-stage viral mRNAs. Rev's shuttling into and out of the nucleus is regulated by the antagonistic activities of both a peptide-encoded N-terminal nuclear localization signal and C-terminal nuclear export signal (NES). How Rev and related viral proteins balance strong import and export activities in order to achieve optimal levels of viral gene expression is incompletely understood. We provide evidence that multimerization provides a mechanism by which Rev transiently masks its NES peptide, thereby biasing its trafficking to and retention within the nucleus. Targeted pharmacological disruption of Rev-Rev interactions should perturb multiple Rev activities, both Rev-RNA binding and Rev's trafficking to the nucleus in the first place. Copyright © 2017 American Society for Microbiology.

Nuclear Export Signal Masking Regulates HIV-1 Rev Trafficking and Viral RNA Nuclear Export

PubMed Central

Behrens, Ryan T.; Aligeti, Mounavya; Pocock, Ginger M.; Higgins, Christina A.

2016-01-01

ABSTRACT HIV-1's Rev protein forms a homo-oligomeric adaptor complex linking viral RNAs to the cellular CRM1/Ran-GTP nuclear export machinery through the activity of Rev's prototypical leucine-rich nuclear export signal (NES). In this study, we used a functional fluorescently tagged Rev fusion protein as a platform to study the effects of modulating Rev NES identity, number, position, or strength on Rev subcellular trafficking, viral RNA nuclear export, and infectious virion production. We found that Rev activity was remarkably tolerant of diverse NES sequences, including supraphysiological NES (SNES) peptides that otherwise arrest CRM1 transport complexes at nuclear pores. Rev's ability to tolerate a SNES was both position and multimerization dependent, an observation consistent with a model wherein Rev self-association acts to transiently mask the NES peptide(s), thereby biasing Rev's trafficking into the nucleus. Combined imaging and functional assays also indicated that NES masking underpins Rev's well-known tendency to accumulate at the nucleolus, as well as Rev's capacity to activate optimal levels of late viral gene expression. We propose that Rev multimerization and NES masking regulates Rev's trafficking to and retention within the nucleus even prior to RNA binding. IMPORTANCE HIV-1 infects more than 34 million people worldwide causing >1 million deaths per year. Infectious virion production is activated by the essential viral Rev protein that mediates nuclear export of intron-bearing late-stage viral mRNAs. Rev's shuttling into and out of the nucleus is regulated by the antagonistic activities of both a peptide-encoded N-terminal nuclear localization signal and C-terminal nuclear export signal (NES). How Rev and related viral proteins balance strong import and export activities in order to achieve optimal levels of viral gene expression is incompletely understood. We provide evidence that multimerization provides a mechanism by which Rev transiently masks its NES peptide, thereby biasing its trafficking to and retention within the nucleus. Targeted pharmacological disruption of Rev-Rev interactions should perturb multiple Rev activities, both Rev-RNA binding and Rev's trafficking to the nucleus in the first place. PMID:27852860

The Gpn3 Q279* cancer-associated mutant inhibits Gpn1 nuclear export and is deficient in RNA polymerase II nuclear targeting.

PubMed

Barbosa-Camacho, Angel A; Méndez-Hernández, Lucía E; Lara-Chacón, Bárbara; Peña-Gómez, Sonia G; Romero, Violeta; González-González, Rogelio; Guerra-Moreno, José A; Robledo-Rivera, Angélica Y; Sánchez-Olea, Roberto; Calera, Mónica R

2017-11-01

Gpn3 is required for RNA polymerase II (RNAPII) nuclear targeting. Here, we investigated the effect of a cancer-associated Q279* nonsense mutation in Gpn3 cellular function. Employing RNAi, we replaced endogenous Gpn3 by wt or Q279* RNAi-resistant Gpn3R in epithelial model cells. RNAPII nuclear accumulation and transcriptional activity were markedly decreased in cells expressing only Gpn3R Q279*. Wild-type Gpn3R localized to the cytoplasm but a fraction of Gpn3R Q279* entered the cell nucleus and inhibited Gpn1-EYFP nuclear export. This property and the transcriptional deficit in Gpn3R Q279*-expressing cells required a PDZ-binding motif generated by the Q279* mutation. We conclude that an acquired PDZ-binding motif in Gpn3 Q279* caused Gpn3 nuclear entry, and inhibited Gpn1 nuclear export and Gpn3-mediated RNAPII nuclear targeting. © 2017 Federation of European Biochemical Societies.

Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network

PubMed Central

Widhalm, Joshua R.; Gutensohn, Michael; Yoo, Heejin; Adebesin, Funmilayo; Qian, Yichun; Guo, Longyun; Jaini, Rohit; Lynch, Joseph H.; McCoy, Rachel M.; Shreve, Jacob T.; Thimmapuram, Jyothi; Rhodes, David; Morgan, John A.; Dudareva, Natalia

2015-01-01

In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles, as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network. PMID:26356302

Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network

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

Widhalm, Joshua R.; Gutensohn, Michael; Yoo, Heejin

In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles,more » as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network.« less

Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network

DOE PAGES

Widhalm, Joshua R.; Gutensohn, Michael; Yoo, Heejin; ...

2015-09-10

In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles,more » as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network.« less

Mechanistic determinants of the directionality and energetics of active export by a heterodimeric ABC transporter

DOE PAGES

Grossmann, Nina; Vakkasoglu, Ahmet S.; Hulpke, Sabine; ...

2014-11-07

The ATP-binding cassette (ABC) transporter associated with antigen processing (TAP) participates in immune surveillance by moving proteasomal products into the endoplasmic reticulum (ER) lumen for major histocompatibility complex class I loading and cell surface presentation to cytotoxic T cells. Here we delineate the mechanistic basis for antigen translocation. Notably, TAP works as a molecular diode, translocating peptide substrates against the gradient in a strict unidirectional way. We reveal the importance of the D-loop at the dimer interface of the two nucleotide-binding domains (NBDs) in coupling substrate translocation with ATP hydrolysis and defining transport vectoriality. Substitution of the converved aspartate, whichmore » coordinates the ATP-binding site, decreases NBD dimerization affinity and turns the unidirectional primary active pump into a passive bidirectional nucleotide-gated facilitator. Thus, ATP hydrolysis is not required for translocation per se, but is essential for both active and unidirectional transport. As a result, our data provide detailed mechanistic insight into how heterodimeric ABC exporters operate.« less

Shallow Carbon Export from an Iron fertilised Plankton Bloom in the Southern Ocean

NASA Astrophysics Data System (ADS)

Sanders, R.; Pollard, R.; Morris, P.; Statham, P.; Moore, C. M. M.; Lucas, M.

2009-04-01

Some regions of the global ocean, notably the Southern Ocean, have high levels of macronutrients yet low levels of chlorophyll (the high nutrient, low chlorophyll or HNLC condition). Numerous artificial iron fertilization experiments conducted in the Southern Ocean have resulted in enhanced phytoplankton biomass and macronutrient drawdown. However the subsequent long-term biogeochemical consequences of such iron fertilization are unclear due in part to the limited size and duration of such experiments. An alternative way to assess the affect of iron over the Southern Ocean biological carbon pump is to observe the evolution of plankton production in regions of the Southern Ocean where shallow topography and Ocean currents interact to promote to release terrestrial iron into HNLC waters. During 2004-5 RRS Discovery conduced a complex programme of observations in such a region around the Crozet Islands in the SW Indian Ocean. The results of this programme, focussing on a quantitative estimate of carbon export per unit iron addition, will be presented.

The function of the inner nuclear envelope protein SUN1 in mRNA export is regulated by phosphorylation.

PubMed

Li, Ping; Stumpf, Maria; Müller, Rolf; Eichinger, Ludwig; Glöckner, Gernot; Noegel, Angelika A

2017-08-22

SUN1, a component of the LINC (Linker of Nucleoskeleton and Cytoskeleton) complex, functions in mammalian mRNA export through the NXF1-dependent pathway. It associates with mRNP complexes by direct interaction with NXF1. It also binds to the NPC through association with the nuclear pore component Nup153, which is involved in mRNA export. The SUN1-NXF1 association is at least partly regulated by a protein kinase C (PKC) which phosphorylates serine 113 (S113) in the N-terminal domain leading to reduced interaction. The phosphorylation appears to be important for the SUN1 function in nuclear mRNA export since GFP-SUN1 carrying a S113A mutation was less efficient in restoring mRNA export after SUN1 knockdown as compared to the wild type protein. By contrast, GFP-SUN1-S113D resembling the phosphorylated state allowed very efficient export of poly(A)+RNA. Furthermore, probing a possible role of the LINC complex component Nesprin-2 in this process we observed impaired mRNA export in Nesprin-2 knockdown cells. This effect might be independent of SUN1 as expression of a GFP tagged SUN-domain deficient SUN1, which no longer can interact with Nesprin-2, did not affect mRNA export.

Millennial-scale variability in dust deposition, marine export production, and nutrient consumption in the glacial subantarctic ocean (Invited)

NASA Astrophysics Data System (ADS)

Martinez-Garcia, A.; Sigman, D. M.; Anderson, R. F.; Ren, H. A.; Hodell, D. A.; Straub, M.; Jaccard, S.; Eglinton, T. I.; Haug, G. H.

2013-12-01

Based on the limitation of modern Southern Ocean phytoplankton by iron and the evidence of higher iron-bearing dust fluxes to the ocean during ice ages, it has been proposed that iron fertilization of Southern Ocean phytoplankton contributed to the reduction in atmospheric CO2 during ice ages. In the Subantarctic zone of the Atlantic Southern Ocean, glacial increases in dust flux and export production have been documented, supporting the iron fertilization hypothesis. However, these observations could be interpreted alternatively as resulting from the equatorward migration of Southern Ocean fronts during ice ages if the observed productivity rise was not accompanied by an increase in major nutrient consumption. Here, new 230Th-normalized lithogenic and opal fluxes are combined with high-resolution biomarker measurements to reconstruct millennial-scale changes in dust deposition and marine export production in the subantarctic Atlantic over the last glacial cycle. In the same record foraminifera-bound nitrogen isotopes are used to reconstruct ice age changes in surface nitrate utilization, providing a comprehensive test of the iron fertilization hypothesis. Elevation in foraminifera-bound δ15N, indicating more complete nitrate consumption, coincides with times of surface cooling and greater dust flux and export production. These observations indicate that the ice age Subantarctic was characterized by iron fertilized phytoplankton growth. The resulting strengthening of the Southern Ocean's biological pump can explain the ~40 ppm lowering of CO2 that characterizes the transitions from mid-climate states to full ice age conditions as well as the millennial-scale atmospheric CO2 fluctuations observed within the last ice age

Polyamines cause plasma membrane depolarization, activate Ca2+-, and modulate H+-ATPase pump activity in pea roots.

PubMed

Pottosin, Igor; Velarde-Buendía, Ana María; Bose, Jayakumar; Fuglsang, Anja T; Shabala, Sergey

2014-06-01

Polyamines regulate a variety of cation and K(+) channels, but their potential effects on cation-transporting ATPases are underexplored. In this work, noninvasive microelectrode ion flux estimation and conventional microelectrode techniques were applied to study the effects of polyamines on Ca(2+) and H(+) transport and membrane potential in pea roots. Externally applied spermine or putrescine (1mM) equally activated eosin yellow (EY)-sensitive Ca(2+) pumping across the root epidermis and caused net H(+) influx or efflux. Proton influx induced by spermine was suppressed by EY, supporting the mechanism in which Ca(2+) pump imports 2 H(+) per each exported Ca(2+). Suppression of the Ca(2+) pump by EY diminished putrescine-induced net H(+) efflux instead of increasing it. Thus, activities of Ca(2+) and H(+) pumps were coupled, likely due to the H(+)-pump inhibition by intracellular Ca(2+). Additionally, spermine but not putrescine caused a direct inhibition of H(+) pumping in isolated plasma membrane vesicles. Spermine, spermidine, and putrescine (1mM) induced membrane depolarization by 70, 50, and 35 mV, respectively. Spermine-induced depolarization was abolished by cation transport blocker Gd(3+), was insensitive to anion channels' blocker niflumate, and was dependent on external Ca(2+). Further analysis showed that uptake of polyamines but not polyamine-induced cationic (K(+)+Ca(2+)+H(+)) fluxes were a main cause of membrane depolarization. Polyamine increase is a common component of plant stress responses. Activation of Ca(2+) efflux by polyamines and contrasting effects of polyamines on net H(+) fluxes and membrane potential can contribute to Ca(2+) signalling and modulate a variety of transport processes across the plasma membrane under stress. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Diversity and regulation of plant Ca2+ pumps: insights from expression in yeast

NASA Technical Reports Server (NTRS)

Sze, H.; Liang, F.; Hwang, I.; Curran, A. C.; Harper, J. F.; Evans, M. L. (Principal Investigator)

2000-01-01

The spatial and temporal regulation of calcium concentration in plant cells depends on the coordinate activities of channels and active transporters located on different organelles and membranes. Several Ca2+ pumps have been identified and characterized by functional expression of plant genes in a yeast mutant (K616). This expression system has opened the way to a genetic and biochemical characterization of the regulatory and catalytic features of diverse Ca2+ pumps. Plant Ca(2+)-ATPases fall into two major types: AtECA1 represents one of four or more members of the type IIA (ER-type) Ca(2+)-ATPases in Arabidopsis, and AtACA2 is one of seven or more members of the type IIB (PM-type) Ca(2+)-ATPases that are regulated by a novel amino terminal domain. Type IIB pumps are widely distributed on membranes, including the PM (plasma membrane), vacuole, and ER (endoplasmic reticulum). The regulatory domain serves multiple functions, including autoinhibition, calmodulin binding, and sites for modification by phosphorylation. This domain, however, is considerably diverse among several type IIB ATPases, suggesting that the pumps are differentially regulated. Understanding of Ca2+ transporters at the molecular level is providing insights into their roles in signaling networks and in regulating fundamental processes of cell biology.

Heme controls ferroportin1 (FPN1) transcription involving Bach1, Nrf2 and a MARE/ARE sequence motif at position -7007 of the FPN1 promoter.

PubMed

Marro, Samuele; Chiabrando, Deborah; Messana, Erika; Stolte, Jens; Turco, Emilia; Tolosano, Emanuela; Muckenthaler, Martina U

2010-08-01

Macrophages of the reticuloendothelial system play a key role in recycling iron from hemoglobin of senescent or damaged erythrocytes. Heme oxygenase 1 degrades the heme moiety and releases inorganic iron that is stored in ferritin or exported to the plasma via the iron export protein ferroportin. In the plasma, iron binds to transferrin and is made available for de novo red cell synthesis. The aim of this study was to gain insight into the regulatory mechanisms that control the transcriptional response of iron export protein ferroportin to hemoglobin in macrophages. Iron export protein ferroportin mRNA expression was analyzed in RAW264.7 mouse macrophages in response to hemoglobin, heme, ferric ammonium citrate or protoporphyrin treatment or to siRNA mediated knockdown or overexpression of Btb And Cnc Homology 1 or nuclear accumulation of Nuclear Factor Erythroid 2-like. Iron export protein ferroportin promoter activity was analyzed using reporter constructs that contain specific truncations of the iron export protein ferroportin promoter or mutations in a newly identified MARE/ARE element. We show that iron export protein ferroportin is transcriptionally co-regulated with heme oxygenase 1 by heme, a degradation product of hemoglobin. The protoporphyrin ring of heme is sufficient to increase iron export protein ferroportin transcriptional activity while the iron released from the heme moiety controls iron export protein ferroportin translation involving the IRE in the 5'untranslated region. Transcription of iron export protein ferroportin is inhibited by Btb and Cnc Homology 1 and activated by Nuclear Factor Erythroid 2-like involving a MARE/ARE element located at position -7007/-7016 of the iron export protein ferroportin promoter. This finding suggests that heme controls a macrophage iron recycling regulon involving Btb and Cnc Homology 1 and Nuclear Factor Erythroid 2-like to assure the coordinated degradation of heme by heme oxygenase 1, iron storage and detoxification by ferritin, and iron export by iron export protein ferroportin.

A subset of FG-nucleoporins is necessary for efficient Msn5-mediated nuclear protein export

PubMed Central

Finn, Erin M.; DeRoo, Elise P.; Clement, George W.; Rao, Sheila; Kruse, Sarah E.; Kokanovich, Kate M.; Belanger, Kenneth D.

2013-01-01

The transport of proteins between the cytoplasm and nucleus requires interactions between soluble transport receptors (karyopherins) and phenylalanine-glycine (FG) repeat domains on nuclear pore complex proteins (nucleoporins). However, the role of specific FG repeat-containing nucleoporins in nuclear protein export has not been carefully investigated. We have developed a novel kinetic assay to investigate the relative export kinetics mediated by the karyopherin Msn5/Kap142 in yeast containing specific FG-Nup mutations. Using the Msn5 substrate Crz1 as a marker for Msn5-mediated protein export, we observe that deletions of NUP100 or NUP2 result in decreased rates of Crz1 export, while nup60Δ and nup42Δ mutants do not vary significantly from wild type. The decreased Msn5 export rate in nup100Δ was confirmed using Mig1-GFP as a transport substrate. A nup100ΔGLFG mutant shows defects in nuclear export kinetics similar to a nup100Δ deletion. Removal of FG-repeats from Nsp1 also decreases export kinetics, while a loss of Nup1 FXFGs does not. To confirm that our export data reflected functional differences in protein localization, we performed Crz1 transcription activation assays using a CDRE::LacZ reporter gene that is upregulated upon increased transcription activation by Crz1 in vivo. We observe that expression from this reporter increases in nup100ΔGLFG and nsp1ΔFGΔFXFG strains that exhibit decreased Crz1 export kinetics but resembles wild-type levels in nup1ΔFXFG strains that do not exhibit export defects. These data provide evidence that the export of Msn5 is likely mediated by a specific subset of FG-Nups and that the GLFG repeat domain of Nup100 is important for Msn5-mediated nuclear protein export. PMID:23295456

The B7-1 Cytoplasmic Tail Enhances Intracellular Transport and Mammalian Cell Surface Display of Chimeric Proteins in the Absence of a Linear ER Export Motif

PubMed Central

Lin, Yi-Chieh; Chen, Bing-Mae; Lu, Wei-Cheng; Su, Chien-I; Prijovich, Zeljko M.; Chung, Wen-Chuan; Wu, Pei-Yu; Chen, Kai-Chuan; Lee, I-Chiao; Juan, Ting-Yi; Roffler, Steve R.

2013-01-01

Membrane-tethered proteins (mammalian surface display) are increasingly being used for novel therapeutic and biotechnology applications. Maximizing surface expression of chimeric proteins on mammalian cells is important for these applications. We show that the cytoplasmic domain from the B7-1 antigen, a commonly used element for mammalian surface display, can enhance the intracellular transport and surface display of chimeric proteins in a Sar1 and Rab1 dependent fashion. However, mutational, alanine scanning and deletion analysis demonstrate the absence of linear ER export motifs in the B7 cytoplasmic domain. Rather, efficient intracellular transport correlated with the presence of predicted secondary structure in the cytoplasmic tail. Examination of the cytoplasmic domains of 984 human and 782 mouse type I transmembrane proteins revealed that many previously identified ER export motifs are rarely found in the cytoplasmic tail of type I transmembrane proteins. Our results suggest that efficient intracellular transport of B7 chimeric proteins is associated with the structure rather than to the presence of a linear ER export motif in the cytoplasmic tail, and indicate that short (less than ~ 10-20 amino acids) and unstructured cytoplasmic tails should be avoided to express high levels of chimeric proteins on mammalian cells. PMID:24073236

Pancreatic bicarbonate secretion involves two proton pumps.

PubMed

Novak, Ivana; Wang, Jing; Henriksen, Katrine L; Haanes, Kristian A; Krabbe, Simon; Nitschke, Roland; Hede, Susanne E

2011-01-07

Pancreas secretes fluid rich in digestive enzymes and bicarbonate. The alkaline secretion is important in buffering of acid chyme entering duodenum and for activation of enzymes. This secretion is formed in pancreatic ducts, and studies to date show that plasma membranes of duct epithelium express H(+)/HCO(3)(-) transporters, which depend on gradients created by the Na(+)/K(+)-ATPase. However, the model cannot fully account for high-bicarbonate concentrations, and other active transporters, i.e. pumps, have not been explored. Here we show that pancreatic ducts express functional gastric and non-gastric H(+)-K(+)-ATPases. We measured intracellular pH and secretion in small ducts isolated from rat pancreas and showed their sensitivity to H(+)-K(+) pump inhibitors and ion substitutions. Gastric and non-gastric H(+)-K(+) pumps were demonstrated on RNA and protein levels, and pumps were localized to the plasma membranes of pancreatic ducts. Quantitative analysis of H(+)/HCO(3)(-) and fluid transport shows that the H(+)-K(+) pumps can contribute to pancreatic secretion in several species. Our results call for revision of the bicarbonate transport physiology in pancreas, and most likely other epithelia. Furthermore, because pancreatic ducts play a central role in several pancreatic diseases, it is of high relevance to understand the role of H(+)-K(+) pumps in pathophysiology.

The Importance of Subsurface Production for Carbon Export - Evidence from Past Oceans

NASA Astrophysics Data System (ADS)

Kemp, A. E. S.

2016-02-01

The maxim of the geological concept of uniformitarianism is "the present is the key to the past", but in the context of our temporally and spatially minimal observational record of modern ocean biogeochemical processes, ancient ocean sediments may provide critical evidence of the key species involved in carbon flux. Specifically, laminated marine sediments that preserve the seasonal flux cycle represent "palaeo-sediment traps" that vastly expand our knowledge of the operations of the marine biological carbon pump. Several key subsurface-dwelling diatom taxa, hitherto thought to be biogeochemically insignificant, are dominant components of ancient marine sediments. For example, the sapropels and equivalent horizons that have accumulated in the Mediterranean over the past 5 million years, contain abundant rhizosolenid and hemiaulid diatoms. These deposits contain the highest concentrations of organic carbon and there is extensive evidence that this was produced by subsurface production in a deep chlorophyll maximum. The highly stratified conditions that led to this subsurface production and carbon flux are in contrast to prevailing views that have held upwelling systems as those with the highest potential for export in the global ocean. Similarly, studies of ancient "greenhouse" periods such as the Cretaceous, with highly stratified oceans and which are potential analogues for future climate change, show evidence for extensive subsurface production. Together with emerging evidence from stratified regions of the modern ocean, such as the subtropical gyres, insights from these ancient oceans suggest that a reappraisal is required of current views on key phytoplankton producers and their role the operation of the marine biological carbon pump.

Adaptation of H+-pumping and plasma membrane H+ ATPase activity in proteoid roots of white lupin under phosphate deficiency.

PubMed

Yan, Feng; Zhu, Yiyong; Müller, Caroline; Zörb, Christian; Schubert, Sven

2002-05-01

White lupin (Lupinus albus) is able to adapt to phosphorus deficiency by producing proteoid roots that release a huge amount of organic acids, resulting in mobilization of sparingly soluble soil phosphate in rhizosphere. The mechanisms responsible for the release of organic acids by proteoid root cells, especially the trans-membrane transport processes, have not been elucidated. Because of high cytosolic pH, the release of undissociated organic acids is not probable. In the present study, we focused on H+ export by plasma membrane H+ ATPase in active proteoid roots. In vivo, rhizosphere acidification of active proteoid roots was vanadate sensitive. Plasma membranes were isolated from proteoid roots and lateral roots from P-deficient and -sufficient plants. In vitro, in comparison with two types of lateral roots and proteoid roots of P-sufficient plants, the following increase of the various parameters was induced in active proteoid roots of P-deficient plants: (a) hydrolytic ATPase activity, (b) Vmax and Km, (c) H+ ATPase enzyme concentration of plasma membrane, (d) H+-pumping activity, (e) pH gradient across the membrane of plasmalemma vesicles, and (f) passive H+ permeability of plasma membrane. In addition, lower vanadate sensitivity and more acidic pH optimum were determined for plasma membrane ATPase of active proteoid roots. Our data support the hypothesis that in active proteoid root cells, H+ and organic anions are exported separately, and that modification of plasma membrane H+ ATPase is essential for enhanced rhizosphere acidification by active proteoid roots.

Modeling the effect of heatsink performance in high-peak-power laser-diode-bar pump sources for solid-state lasers 011 011

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

Honea, E.C., LLNL

We derive approximate expressions for transient output power and wavelength chirp of high- peak-power laser-diode bars assuming one-dimensional heat flow and linear temperature dependences for chirp and efficiency. The model is derived for pulse durations, 10 < {tau} < 1000 ps, typically used for diode-pumped solid-state lasers and is in good agreement with experimental data for Si heatsink mounted 940 nm laser-diode bars operating at 100 W/cm. The analytic expressions are more flexible and easily used than the results of operating point dependent numerical modeling. In addition, the analytic expressions used here can be integrated to describe the energy permore » unit wavelength for a given pulse duration, initial emission bandwidth and heatsink material. We find that the figure-of-merit for a heatsink material in this application is ({rho}C{sub p}K) where {rho}C{sub p} is the volumetric heat capacity and K is the thermal conductivity. As an example of the utility of the derived expressions, we determine an effective absorption coefficient as a function of pump pulse duration for a diode-pumped solid-state laser utilizing Yb:Sr{sub 5}(PO{sub 4}){sub 3}F (Yb:S-FAP) as the gain medium.« less

Evaluation of the hydrologic system and selected water-management alternatives in the Owens Valley, California

USGS Publications Warehouse

Danskin, Wesley R.

1998-01-01

The Owens Valley, a long, narrow valley along the east side of the Sierra Nevada in eastcentral California, is the main source of water for the city of Los Angeles. The city diverts most of the surface water in the valley into the Owens River?Los Angeles Aqueduct system, which transports the water more than 200 miles south to areas of distribution and use. Additionally, ground water is pumped or flows from wells to supplement the surface-water diversions to the river? aqueduct system. Pumpage from wells needed to supplement water export has increased since 1970, when a second aqueduct was put into service, and local residents have expressed concerns that the increased pumping may have a detrimental effect on the environment and the native vegetation (indigenous alkaline scrub and meadow plant communities) in the valley. Native vegetation on the valley floor depends on soil moisture derived from precipitation and from the unconfined part of a multilayered ground-water system. This report, which describes the evaluation of the hydrologic system and selected water-management alternatives, is one in a series designed to identify the effects that ground-water pumping has on native vegetation and evaluate alternative strategies to mitigate any adverse effects caused by pumping. The hydrologic system of the Owens Valley can be conceptualized as having three parts: (1) an unsaturated zone affected by precipitation and evapotranspiration; (2) a surface-water system composed of the Owens River, the Los Angeles Aqueduct, tributary streams, canals, ditches, and ponds; and (3) a saturated ground-water system contained in the valley fill. Analysis of the hydrologic system was aided by development of a ground-water flow model of the ?aquifer system,? which is defined as the most active part of the ground-water system and which includes nearly all of the Owens Valley except for the area surrounding the Owens Lake. The model was calibrated and verified for water years 1963?88 and used to evaluate general concepts of the hydrologic system and the effects of past water-management practices. The model also was used to evaluate the likely effects of selected water-management alternatives designed to lessen the adverse effects of ground-water pumping on native vegetation. Results of the model simulations confirm that a major change in the hydrologic system was caused by the additional export of water from the valley beginning in 1970. Average ground-water pumpage increased by a factor of five, discharge from springs decreased almost to zero, reaches of the Owens River that previously had gained water from the aquifer system began losing water, and total evapotranspiration by native plants decreased by about 35 percent. Water-management practices as of 1988 were defined and evaluted using the model. Simulation results indicate that increased ground-water pumpage since 1985 for enhancement and mitigation projects within the Owens Valley has further stressed the aquifer system and resulted in declines of the water table and reduced evapotranspiration. Most of the water-table declines are beneath the western alluvial fans and in the immediate vicinity of production wells. The water-table altitude beneath the valley floor has remained relatively constant over time because of hydrologic buffers, such as evapotranspiration, springs, and permanent surface-water features. These buffers adjust the quantity of water exchanged with the aquifer system and effectively minimize variations in water-table altitude. The widespread presence of hydrologic buffers is the primary reason the water-table altitude beneath the valley floor has remained relatively constant since 1970 despite major changes in the type and location of ground-water discharge. Evaluation of selected water-management alternatives indicates that long-term variations in average runoff to the Owens Valley of as much as

How UK internet websites portray breast milk expression and breast pumps: a qualitative study of content.

PubMed

McInnes, Rhona J; Arbuckle, Alix; Hoddinott, Pat

2015-04-02

Exclusive breastfeeding for six months is recommended but few parents achieve this; particularly younger and less well-educated mothers. Many parents introduce infant formula milk to manage feeding but describe a desire to express breastmilk alongside a lack of support or information. The Internet is highlighted as a key resource. This study aimed to examine UK websites on expressing breastmilk to identify key messages and how information is provided. We used search terms in Google to identify websites with information rich content on expressing breastmilk and breast pumps. Ten sites were purposively selected at two time points in 2013 and 2014 to represent 3 categories: commercial, NHS or 3(rd) sector (voluntary or not for profit). Each site was reviewed by two researchers, data and reflective analytical notes were uploaded into NVivo and thematic data analysis undertaken. Sites varied considerably in their design, use of images, videos, audio files, product placement and marketing opportunities. Three key themes emerged: depiction of expressing; reasons to express; and recommendations about expressing. Inconsistent and conflicting information was common within and between sites. Expressing was portrayed as similar to, but easier than, breastfeeding although at the same time difficult and requiring to be learned. Expressed breastmilk is promoted by mainly commercial sites as immediately available, although pumps were also presented as needing to be concealed, not heard or seen. Health benefits were the overarching reason for expressing. Although predicated on separation from the baby, commercial sites identified this as a positive choice while other sites focused on separation due to circumstance. Commercial sites emphasised restrictions related to breastfeeding, lack of sleep and bonding with the father and wider family. Non-commercial sites emphasised hand expression, with some not mentioning breast pumps. Practical information about starting expressing in relation to infant age or duration of breastfeeding was conflicting. Internet information about expressing breastmilk is inconsistent, incomplete and not evidence informed. The lack of research evidence on the relationship between expressing and feeding outcomes has provided opportunities for commercial companies, which have the potential to further exacerbate observed health inequalities. Access to good quality information based on robust evidence is urgently required.

Paeoniflorin ameliorates cholestasis via regulating hepatic transporters and suppressing inflammation in ANIT-fed rats.

PubMed

Zhao, Yanling; He, Xuan; Ma, Xiao; Wen, Jianxia; Li, Pengyan; Wang, Jiabo; Li, Ruisheng; Zhu, Yun; Wei, Shizhang; Li, Haotian; Zhou, Xuelin; Li, Kun; Liu, Honghong; Xiao, Xiaohe

2017-05-01

Paeoniflorin has shown the obvious effect on cholestasis according to our previous research. However, its mechanism has not been absolutely explored yet. This study aims at evaluating the potential effect of paeoniflorin on alpha-naphthylisothiocyanate (ANIT) -induced cholestasis by inhibiting nuclear factor kappa-B (NF-κB) and simultaneously regulating hepatocyte transporters. Cholestasis was induced by administration of ANIT. The effect of paeoniflorin on serum indices such as total bilirubin (TBIL), direct bilirubin (DBIL), aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), γ-glutamyltranspeptidase (γ-GT), total bile acid (TBA) and histopathology of liver were determined. Liver protein levels of NF-κB, interleukin 1β (IL-1β) and the hepatocyte transporters such as Na + /taurocholate cotransporting polypeptide (NTCP), bile salt export pump (BSEP), multidrug resistance-associated protein 2 (MRP2) and cholesterol 7α-hydroxylase (Cyp7a1) were investigated by western blotting. The results demonstrated that paeoniflorin could decrease serum ALT, AST, ALP, γ-GT, TBIL, DBIL and TBA in ANIT-treated rats. Histological examination revealed that rats treated with paeoniflorin represented fewer neutrophils infiltration, edema and necrosis in liver tissue compared with ANIT rats. Moreover, paeoniflorin significantly reduced the over expressions of NF-κB and IL-1β induced by ANIT in liver tissue. In addition, the relative protein expressions of NTCP, BSEP, MRP2 but not Cyp7a1 were also restored by paeoniflorin. The potential mechanism of paeoniflorin in alleviating ANIT-induced cholestasis seems to be related to reduce the over expressions of NF-κB and hepatocyte transporters such as NTCP, BSEP as well as MRP2. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Phage insertion in mlrA and variations in rpoS limit curli expression and biofilm formation in Escherichia coli serotype O157:H7

USDA-ARS?s Scientific Manuscript database

Biofilm formation in Escherichia coli is a tightly controlled process requiring the expression of adhesive curli fibers and certain polysaccharides such as cellulose. The transcriptional regulator CsgD is central to biofilm formation, controlling the expression of the curli structural and export pro...

Marine phytoplankton stoichiometry mediates nonlinear interactions between nutrient supply, temperature, and atmospheric CO2

NASA Astrophysics Data System (ADS)

Moreno, Allison R.; Hagstrom, George I.; Primeau, Francois W.; Levin, Simon A.; Martiny, Adam C.

2018-05-01

Marine phytoplankton stoichiometry links nutrient supply to marine carbon export. Deviations of phytoplankton stoichiometry from Redfield proportions (106C : 1P) could therefore have a significant impact on carbon cycling, and understanding which environmental factors drive these deviations may reveal new mechanisms regulating the carbon cycle. To explore the links between environmental conditions, stoichiometry, and carbon cycling, we compared four different models of phytoplankton C : P: a fixed Redfield model, a model with C : P given as a function of surface phosphorus concentration (P), a model with C P given as a function of temperature, and a new multi-environmental model that predicts C : P as a function of light, temperature, and P. These stoichiometric models were embedded into a five-box ocean circulation model, which resolves the three major ocean biomes (high-latitude, subtropical gyres, and tropical upwelling regions). Contrary to the expectation of a monotonic relationship between surface nutrient drawdown and carbon export, we found that lateral nutrient transport from lower C : P tropical waters to high C : P subtropical waters could cause carbon export to decrease with increased tropical nutrient utilization. It has been hypothesized that a positive feedback between temperature and pCO2, atm will play an important role in anthropogenic climate change, with changes in the biological pump playing at most a secondary role. Here we show that environmentally driven shifts in stoichiometry make the biological pump more influential, and may reverse the expected positive relationship between temperature and pCO2, atm. In the temperature-only model, changes in tropical temperature have more impact on the Δ pCO2, atm (˜ 41 ppm) compared to subtropical temperature changes (˜ 4.5 ppm). Our multi-environmental model predicted a decline in pCO2, atm of ˜ 46 ppm when temperature spanned a change of 10 °C. Thus, we find that variation in marine phytoplankton stoichiometry and its environmental controlling factors can lead to nonlinear controls on pCO2, atm, suggesting the need for further studies of ocean C : P and the impact on ocean carbon cycling.

Circadian processes in the RNA life cycle.

PubMed

Torres, Manon; Becquet, Denis; Franc, Jean-Louis; François-Bellan, Anne-Marie

2018-05-01

The circadian clock drives daily rhythms of multiple physiological processes, allowing organisms to anticipate and adjust to periodic changes in environmental conditions. These physiological rhythms are associated with robust oscillations in the expression of at least 30% of expressed genes. While the ability for the endogenous timekeeping system to generate a 24-hr cycle is a cell-autonomous mechanism based on negative autoregulatory feedback loops of transcription and translation involving core-clock genes and their protein products, it is now increasingly evident that additional mechanisms also govern the circadian oscillations of clock-controlled genes. Such mechanisms can take place post-transcriptionally during the course of the RNA life cycle. It has been shown that many steps during RNA processing are regulated in a circadian manner, thus contributing to circadian gene expression. These steps include mRNA capping, alternative splicing, changes in splicing efficiency, and changes in RNA stability controlled by the tail length of polyadenylation or the use of alternative polyadenylation sites. RNA transport can also follow a circadian pattern, with a circadian nuclear retention driven by rhythmic expression within the nucleus of particular bodies (the paraspeckles) and circadian export to the cytoplasm driven by rhythmic proteins acting like cargo. Finally, RNA degradation may also follow a circadian pattern through the rhythmic involvement of miRNAs. In this review, we summarize the current knowledge of the post-transcriptional circadian mechanisms known to play a prominent role in shaping circadian gene expression in mammals. This article is categorized under: RNA Processing > Splicing Regulation/Alternative Splicing RNA Processing > RNA Editing and Modification RNA Export and Localization > Nuclear Export/Import. © 2018 Wiley Periodicals, Inc.

Arterial α2-Na+ pump expression influences blood pressure: lessons from novel, genetically engineered smooth muscle-specific α2 mice.

PubMed

Chen, Ling; Song, Hong; Wang, Youhua; Lee, Jane C; Kotlikoff, Michael I; Pritchard, Tracy J; Paul, Richard J; Zhang, Jin; Blaustein, Mordecai P

2015-09-01

Arterial myocytes express α1-catalytic subunit isoform Na(+) pumps (75-80% of total), which are ouabain resistant in rodents, and high ouabain affinity α2-Na(+) pumps. Mice with globally reduced α2-pumps (but not α1-pumps), mice with mutant ouabain-resistant α2-pumps, and mice with a smooth muscle (SM)-specific α2-transgene (α2 (SM-Tg)) that induces overexpression all have altered blood pressure (BP) phenotypes. We generated α2 (SM-DN) mice with SM-specific α2 (not α1) reduction (>50%) using nonfunctional dominant negative (DN) α2. We compared α2 (SM-DN) and α2 (SM-Tg) mice to controls to determine how arterial SM α2-pumps affect vasoconstriction and BP. α2 (SM-DN) mice had elevated basal mean BP (mean BP by telemetry: 117 ± 4 vs. 106 ± 1 mmHg, n = 7/7, P < 0.01) and enhanced BP responses to chronic ANG II infusion (240 ng·kg(-1)·min(-1)) and high (6%) NaCl. Several arterial Ca(2+) transporters, including Na(+)/Ca(2+) exchanger 1 (NCX1) and sarcoplasmic reticulum and plasma membrane Ca(2+) pumps [sarco(endo)plasmic reticulum Ca(2+)-ATPase 2 (SERCA2) and plasma membrane Ca(2+)-ATPase 1 (PMCA1)], were also reduced (>50%). α2 (SM-DN) mouse isolated small arteries had reduced myogenic reactivity, perhaps because of reduced Ca(2+) transporter expression. In contrast, α2 (SM-Tg) mouse aortas overexpressed α2 (>2-fold), NCX1, SERCA2, and PMCA1 (43). α2 (SM-Tg) mice had reduced basal mean BP (104 ± 1 vs. 109 ± 2 mmHg, n = 15/9, P < 0.02) and attenuated BP responses to chronic ANG II (300-400 ng·kg(-1)·min(-1)) with or without 2% NaCl but normal myogenic reactivity. NCX1 expression was inversely related to basal BP in SM-α2 engineered mice but was directly related in SM-NCX1 engineered mice. NCX1, which usually mediates arterial Ca(2+) entry, and α2-Na(+) pumps colocalize at plasma membrane-sarcoplasmic reticulum junctions and functionally couple via the local Na(+) gradient to help regulate cell Ca(2+). Altered Ca(2+) transporter expression in SM-α2 engineered mice apparently compensates to minimize Ca(2+) overload (α2 (SM-DN)) or depletion (α2 (SM-Tg)) and attenuate BP changes. In contrast, Ca(2+) transporter upregulation, observed in many rodent hypertension models, should enhance Ca(2+) entry and signaling and contribute significantly to BP elevation. Copyright © 2015 the American Physiological Society.

The plant energy-dissipating mitochondrial systems: depicting the genomic structure and the expression profiles of the gene families of uncoupling protein and alternative oxidase in monocots and dicots.

PubMed

Borecky, Jirí; Nogueira, Fábio T S; de Oliveira, Kívia A P; Maia, Ivan G; Vercesi, Aníbal E; Arruda, Paulo

2006-01-01

The simultaneous existence of alternative oxidases and uncoupling proteins in plants has raised the question as to why plants need two energy-dissipating systems with apparently similar physiological functions. A probably complete plant uncoupling protein gene family is described and the expression profiles of this family compared with the multigene family of alternative oxidases in Arabidopsis thaliana and sugarcane (Saccharum sp.) employed as dicot and monocot models, respectively. In total, six uncoupling protein genes, AtPUMP1-6, were recognized within the Arabidopsis genome and five (SsPUMP1-5) in a sugarcane EST database. The recombinant AtPUMP5 protein displayed similar biochemical properties as AtPUMP1. Sugarcane possessed four Arabidopsis AOx1-type orthologues (SsAOx1a-1d); no sugarcane orthologue corresponding to Arabidopsis AOx2-type genes was identified. Phylogenetic and expression analyses suggested that AtAOx1d does not belong to the AOx1-type family but forms a new (AOx3-type) family. Tissue-enriched expression profiling revealed that uncoupling protein genes were expressed more ubiquitously than the alternative oxidase genes. Distinct expression patterns among gene family members were observed between monocots and dicots and during chilling stress. These findings suggest that the members of each energy-dissipating system are subject to different cell or tissue/organ transcriptional regulation. As a result, plants may respond more flexibly to adverse biotic and abiotic conditions, in which oxidative stress is involved.

The E3 ubiquitin ligase, HECTD1, is involved in ABCA1-mediated cholesterol export from macrophages.

PubMed

Aleidi, Shereen M; Yang, Alryel; Sharpe, Laura J; Rao, Geetha; Cochran, Blake J; Rye, Kerry-Anne; Kockx, Maaike; Brown, Andrew J; Gelissen, Ingrid C

2018-04-01

The ABC lipid transporters, ABCA1 and ABCG1, are essential for maintaining lipid homeostasis in cells such as macrophages by exporting excess cholesterol to extracellular acceptors. These transporters are highly regulated at the post-translational level, including protein ubiquitination. Our aim was to investigate the role of the E3 ubiquitin ligase HECTD1, recently identified as associated with ABCG1, on ABCG1 and ABCA1 protein levels and cholesterol export function. Here, we show that HECTD1 protein is widely expressed in a range of human and murine primary cells and cell lines, including macrophages, neuronal cells and insulin secreting β-cells. siRNA knockdown of HECTD1 unexpectedly decreased overexpressed ABCG1 protein levels and cell growth, but increased native ABCA1 protein in CHO-K1 cells. Knockdown of HECTD1 in unloaded THP-1 macrophages did not affect ABCG1 but significantly increased ABCA1 protein levels, in wild-type as well as THP-1 cells that do not express ABCG1. Cholesterol export from macrophages to apoA-I over time was increased after knockdown of HECTD1, however these effects were not sustained in cholesterol-loaded cells. In conclusion, we have identified a new candidate, the E3 ubiquitin ligase HECTD1, that may be involved in the regulation of ABCA1-mediated cholesterol export from unloaded macrophages to apoA-I. The exact mechanism by which this ligase affects this pathway remains to be elucidated. Copyright © 2018 Elsevier B.V. All rights reserved.

Sodium Pumps Mediate Activity-Dependent Changes in Mammalian Motor Networks

PubMed Central

Picton, Laurence D.; Nascimento, Filipe; Broadhead, Matthew J.; Sillar, Keith T.

2017-01-01

Ubiquitously expressed sodium pumps are best known for maintaining the ionic gradients and resting membrane potential required for generating action potentials. However, activity- and state-dependent changes in pump activity can also influence neuronal firing and regulate rhythmic network output. Here we demonstrate that changes in sodium pump activity regulate locomotor networks in the spinal cord of neonatal mice. The sodium pump inhibitor, ouabain, increased the frequency and decreased the amplitude of drug-induced locomotor bursting, effects that were dependent on the presence of the neuromodulator dopamine. Conversely, activating the pump with the sodium ionophore monensin decreased burst frequency. When more “natural” locomotor output was evoked using dorsal-root stimulation, ouabain increased burst frequency and extended locomotor episode duration, whereas monensin slowed and shortened episodes. Decreasing the time between dorsal-root stimulation, and therefore interepisode interval, also shortened and slowed activity, suggesting that pump activity encodes information about past network output and contributes to feedforward control of subsequent locomotor bouts. Using whole-cell patch-clamp recordings from spinal motoneurons and interneurons, we describe a long-duration (∼60 s), activity-dependent, TTX- and ouabain-sensitive, hyperpolarization (∼5 mV), which is mediated by spike-dependent increases in pump activity. The duration of this dynamic pump potential is enhanced by dopamine. Our results therefore reveal sodium pumps as dynamic regulators of mammalian spinal motor networks that can also be affected by neuromodulatory systems. Given the involvement of sodium pumps in movement disorders, such as amyotrophic lateral sclerosis and rapid-onset dystonia parkinsonism, knowledge of their contribution to motor network regulation also has considerable clinical importance. SIGNIFICANCE STATEMENT The sodium pump is ubiquitously expressed and responsible for at least half of total brain energy consumption. The pumps maintain ionic gradients and the resting membrane potential of neurons, but increasing evidence suggests that activity- and state-dependent changes in pump activity also influence neuronal firing. Here we demonstrate that changes in sodium pump activity regulate locomotor output in the spinal cord of neonatal mice. We describe a sodium pump-mediated afterhyperpolarization in spinal neurons, mediated by spike-dependent increases in pump activity, which is affected by dopamine. Understanding how sodium pumps contribute to network regulation and are targeted by neuromodulators, including dopamine, has clinical relevance due to the role of the sodium pump in diseases, including amyotrophic lateral sclerosis, parkinsonism, epilepsy, and hemiplegic migraine. PMID:28123025

The RNA Export Factor, Nxt1, Is Required for Tissue Specific Transcriptional Regulation

PubMed Central

Jiang, Jianqiao; White-Cooper, Helen

2013-01-01

The highly conserved, Nxf/Nxt (TAP/p15) RNA nuclear export pathway is important for export of most mRNAs from the nucleus, by interacting with mRNAs and promoting their passage through nuclear pores. Nxt1 is essential for viability; using a partial loss of function allele, we reveal a role for this gene in tissue specific transcription. We show that many Drosophila melanogaster testis-specific mRNAs require Nxt1 for their accumulation. The transcripts that require Nxt1 also depend on a testis-specific transcription complex, tMAC. We show that loss of Nxt1 leads to reduced transcription of tMAC targets. A reporter transcript from a tMAC-dependent promoter is under-expressed in Nxt1 mutants, however the same transcript accumulates in mutants if driven by a tMAC-independent promoter. Thus, in Drosophila primary spermatocytes, the transcription factor used to activate expression of a transcript, rather than the RNA sequence itself or the core transcription machinery, determines whether this expression requires Nxt1. We additionally find that transcripts from intron-less genes are more sensitive to loss of Nxt1 function than those from intron-containing genes and propose a mechanism in which transcript processing feeds back to increase activity of a tissue specific transcription complex. PMID:23754955

FliH and FliI ensure efficient energy coupling of flagellar type III protein export in Salmonella.

PubMed

Minamino, Tohru; Kinoshita, Miki; Inoue, Yumi; Morimoto, Yusuke V; Ihara, Kunio; Koya, Satomi; Hara, Noritaka; Nishioka, Noriko; Kojima, Seiji; Homma, Michio; Namba, Keiichi

2016-06-01

For construction of the bacterial flagellum, flagellar proteins are exported via its specific export apparatus from the cytoplasm to the distal end of the growing flagellar structure. The flagellar export apparatus consists of a transmembrane (TM) export gate complex and a cytoplasmic ATPase complex consisting of FliH, FliI, and FliJ. FlhA is a TM export gate protein and plays important roles in energy coupling of protein translocation. However, the energy coupling mechanism remains unknown. Here, we performed a cross-complementation assay to measure robustness of the energy transduction system of the export apparatus against genetic perturbations. Vibrio FlhA restored motility of a Salmonella ΔflhA mutant but not that of a ΔfliH-fliI flhB(P28T) ΔflhA mutant. The flgM mutations significantly increased flagellar gene expression levels, allowing Vibrio FlhA to exert its export activity in the ΔfliH-fliI flhB(P28T) ΔflhA mutant. Pull-down assays revealed that the binding affinities of Vibrio FlhA for FliJ and the FlgN-FlgK chaperone-substrate complex were much lower than those of Salmonella FlhA. These suggest that Vibrio FlhA requires the support of FliH and FliI to efficiently and properly interact with FliJ and the FlgN-FlgK complex. We propose that FliH and FliI ensure robust and efficient energy coupling of protein export during flagellar assembly. © 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Influence of particulates on phosphorus loading exported from farm drainage during a storm event in the Everglades Agricultural Area

NASA Astrophysics Data System (ADS)

Bhadha, J. H.; Lang, T. A.; Daroub, S. H.

2015-12-01

The purpose of this study was to evaluate the influence of particulates on P loading captured during a single storm event. The Everglades Agricultural Area of Florida comprises 280,000 hectares of organic soil farmland artificially drained by ditches, canals and pumps. Phosphorus (P)-enriched suspended particulates in canals are susceptible to transport and can contribute significantly to the overall P loads in drainage water. A settling tank experiment was conducted to capture particulates during tropical storm Isaac in 2012 from three farms approximately 2.4 to 3.6 km2 in size. Farm canal discharge water was collected in a series of two 200 liter settling tanks over a seven-day drainage period, during tropical storm Isaac. Water from the settling tanks was siphoned through Imhoff settling cones, where the particulates were allowed to settle and collected for P-fractionation analyses, and compared to intact sediment cores collected from the bottom of the canals. The discharged particulates contained higher organic matter content (OM), total P, and labile P fractions compared to the canal bottom sediments. Based on the equilibrium P concentrations, surface sediments behave as a source of P to the water column. A seven-day continuous drainage event exported 4.7 to 11.1 metric tons of suspended solids per farm, corresponding to 32 to 63 kg of particulate P being lost to downstream ecosystems. Drainage associated to a single seven-day storm event exported up to 61% of the total annual farm P load. It is evident from this study that short-term, high-intensity storm events can skew annual P loads due to the export of significantly higher particulate matter from farm canals. Exported particulates rich in P can provide a supplemental source of nutrients if captured and replenished back into the farmlands, as a sustainable farming practice.

Targeting efflux pumps to overcome antifungal drug resistance

PubMed Central

Holmes, Ann R; Cardno, Tony S; Strouse, J Jacob; Ivnitski-Steele, Irena; Keniya, Mikhail V; Lackovic, Kurt; Monk, Brian C; Sklar, Larry A; Cannon, Richard D

2016-01-01

Resistance to antifungal drugs is an increasingly significant clinical problem. The most common antifungal resistance encountered is efflux pump-mediated resistance of Candida species to azole drugs. One approach to overcome this resistance is to inhibit the pumps and chemosensitize resistant strains to azole drugs. Drug discovery targeting fungal efflux pumps could thus result in the development of azole-enhancing combination therapy. Heterologous expression of fungal efflux pumps in Saccharomyces cerevisiae provides a versatile system for screening for pump inhibitors. Fungal efflux pumps transport a range of xenobiotics including fluorescent compounds. This enables the use of fluorescence-based detection, as well as growth inhibition assays, in screens to discover compounds targeting efflux-mediated antifungal drug resistance. A variety of medium- and high-throughput screens have been used to identify a number of chemical entities that inhibit fungal efflux pumps. PMID:27463566

Nutrient availability and the ultimate control of the biological carbon pump in the western tropical South Pacific Ocean

NASA Astrophysics Data System (ADS)

Moutin, Thierry; Wagener, Thibaut; Caffin, Mathieu; Fumenia, Alain; Gimenez, Audrey; Baklouti, Melika; Bouruet-Aubertot, Pascale; Pujo-Pay, Mireille; Leblanc, Karine; Lefevre, Dominique; Helias Nunige, Sandra; Leblond, Nathalie; Grosso, Olivier; de Verneil, Alain

2018-05-01

Surface waters (0-200 m) of the western tropical South Pacific (WTSP) were sampled along a longitudinal 4000 km transect (OUTPACE cruise, DOI: 10.17600/15000900) during the austral summer (stratified) period (18 February to 3 April 2015) between the Melanesian Archipelago (MA) and the western part of the SP gyre (WGY). Two distinct areas were considered for the MA, the western MA (WMA), and the eastern MA (EMA). The main carbon (C), nitrogen (N), and phosphorus (P) pools and fluxes provide a basis for the characterization of the expected trend from oligotrophy to ultra-oligotrophy, and the building of first-order budgets at the daily and seasonal timescales (using climatology). Sea surface chlorophyll a well reflected the expected oligotrophic gradient with higher values obtained at WMA, lower values at WGY, and intermediate values at EMA. As expected, the euphotic zone depth, the deep chlorophyll maximum, and nitracline depth deepen from west to east. Nevertheless, phosphaclines and nitraclines did not match. The decoupling between phosphacline and nitracline depths in the MA allows for excess P to be locally provided in the upper water by winter mixing. We found a significant biological soft tissue carbon pump in the MA sustained almost exclusively by dinitrogen (N2) fixation and essentially controlled by phosphate availability in this iron-rich environment. The MA appears to be a net sink for atmospheric CO2, while the WGY is in quasi-steady state. We suggest that the necessary excess P, allowing the success of nitrogen fixers and subsequent carbon production and export, is mainly brought to the upper surface by local deep winter convection at an annual timescale rather than by surface circulation. While the origin of the decoupling between phosphacline and nitracline remains uncertain, the direct link between local P upper water enrichment, N2 fixation, and organic carbon production and export, offers a possible shorter timescale than previously thought between N input by N2 fixation and carbon export. The low iron availability in the SP gyre and P availability in the MA during the stratified period may appear as the ultimate control of N input by N2 fixation. Because of the huge volume of water to consider, and because the SP Ocean is the place of intense denitrification in the east (N sink) and N2 fixation in the west (N source), precise seasonal C, N, P, and iron (Fe) budgets would be of prime interest to understand the efficiency, at the present time and in the future, of the oceanic biological carbon pump.

Seasonally different carbon flux changes in the Southern Ocean in response to the southern annular mode.

PubMed

Hauck, J; Völker, C; Wang, T; Hoppema, M; Losch, M; Wolf-Gladrow, D A

2013-12-01

Stratospheric ozone depletion and emission of greenhouse gases lead to a trend of the southern annular mode (SAM) toward its high-index polarity. The positive phase of the SAM is characterized by stronger than usual westerly winds that induce changes in the physical carbon transport. Changes in the natural carbon budget of the upper 100 m of the Southern Ocean in response to a positive SAM phase are explored with a coupled ecosystem-general circulation model and regression analysis. Previously overlooked processes that are important for the upper ocean carbon budget during a positive SAM period are identified, namely, export production and downward transport of carbon north of the polar front (PF) as large as the upwelling in the south. The limiting micronutrient iron is brought into the surface layer by upwelling and stimulates phytoplankton growth and export production but only in summer. This leads to a drawdown of carbon and less summertime outgassing (or more uptake) of natural CO 2 . In winter, biological mechanisms are inactive, and the surface ocean equilibrates with the atmosphere by releasing CO 2 . In the annual mean, the upper ocean region south of the PF loses more carbon by additional export production than by the release of CO 2 into the atmosphere, highlighting the role of the biological carbon pump in response to a positive SAM event.

Anticyclonic eddies are more productive than cyclonic eddies in subtropical gyres because of winter mixing.

PubMed

Dufois, François; Hardman-Mountford, Nick J; Greenwood, Jim; Richardson, Anthony J; Feng, Ming; Matear, Richard J

2016-05-01

Mesoscale eddies are ubiquitous features of ocean circulation that modulate the supply of nutrients to the upper sunlit ocean, influencing the rates of carbon fixation and export. The popular eddy-pumping paradigm implies that nutrient fluxes are enhanced in cyclonic eddies because of upwelling inside the eddy, leading to higher phytoplankton production. We show that this view does not hold for a substantial portion of eddies within oceanic subtropical gyres, the largest ecosystems in the ocean. Using space-based measurements and a global biogeochemical model, we demonstrate that during winter when subtropical eddies are most productive, there is increased chlorophyll in anticyclones compared with cyclones in all subtropical gyres (by 3.6 to 16.7% for the five basins). The model suggests that this is a consequence of the modulation of winter mixing by eddies. These results establish a new paradigm for anticyclonic eddies in subtropical gyres and could have important implications for the biological carbon pump and the global carbon cycle.

Sumoylation of Smad3 stimulates its nuclear export during PIASy-mediated suppression of TGF-{beta} signaling

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

Imoto, Seiyu; Ohbayashi, Norihiko; Ikeda, Osamu

2008-05-30

Sma- and MAD-related protein 3 (Smad3) plays crucial roles in the transforming growth factor-{beta} (TGF-{beta})-mediated signaling pathway, which produce a variety of cellular responses, including cell proliferation and differentiation. In our previous study, we demonstrated that protein inhibitor of activated STATy (PIASy) suppresses TGF-{beta} signaling by interacting with and sumoylating Smad3. In the present study, we examined the molecular mechanisms of Smad3 sumoylation during PIASy-mediated suppression of TGF-{beta} signaling. We found that small-interfering RNA-mediated reduction of endogenous PIASy expression enhanced TGF-{beta}-induced gene expression. Importantly, coexpression of Smad3 with PIASy and SUMO1 affected the DNA-binding activity of Smad3. Furthermore, coexpression ofmore » Smad3 with PIASy and SUMO1 stimulated the nuclear export of Smad3. Finally, fluorescence resonance energy transfer analyses revealed that Smad3 interacted with SUMO1 in the cytoplasm. These results suggest that PIASy regulates TGF-{beta}/Smad3-mediated signaling by stimulating sumoylation and nuclear export of Smad3.« less

Extracellular accumulation of recombinant protein by Escherichia coli in a defined medium.

PubMed

Fu, Xiang-Yang

2010-09-01

Extracellular accumulation of recombinant proteins in the culture medium of Escherichia coli is desirable but difficult to obtain. The inner or cytoplasmic membrane and the outer membrane of E. coli are two barriers for releasing recombinant proteins expressed in the cytoplasm into the culture medium. Even if recombinant proteins have been exported into the periplasm, a space between the outer membrane and the inner membrane, the outer membrane remains the last barrier for their extracellular release. However, when E. coli was cultured in a particular defined medium, recombinant proteins exported into the periplasm could diffuse into the culture medium automatically. If a nonionic detergent, Triton X-100, was added in the medium, recombinant proteins expressed in the cytoplasm could also be released into the culture medium. It was then that extracellular accumulation of recombinant proteins could be obtained by exporting them into the periplasm or releasing them from the cytoplasm with Triton X-100 addition. The tactics described herein provided simple and valuable methods for achieving extracellular production of recombinant proteins in E. coli.

Rerouting Carbon Flux To Enhance Photosynthetic Productivity

PubMed Central

Ducat, Daniel C.; Avelar-Rivas, J. Abraham; Way, Jeffrey C.

2012-01-01

The bioindustrial production of fuels, chemicals, and therapeutics typically relies upon carbohydrate inputs derived from agricultural plants, resulting in the entanglement of food and chemical commodity markets. We demonstrate the efficient production of sucrose from a cyanobacterial species, Synechococcus elongatus, heterologously expressing a symporter of protons and sucrose (cscB). cscB-expressing cyanobacteria export sucrose irreversibly to concentrations of >10 mM without culture toxicity. Moreover, sucrose-exporting cyanobacteria exhibit increased biomass production rates relative to wild-type strains, accompanied by enhanced photosystem II activity, carbon fixation, and chlorophyll content. The genetic modification of sucrose biosynthesis pathways to minimize competing glucose- or sucrose-consuming reactions can further improve sucrose production, allowing the export of sucrose at rates of up to 36.1 mg liter−1 h illumination−1. This rate of production exceeds that of previous reports of targeted, photobiological production from microbes. Engineered S. elongatus produces sucrose in sufficient quantities (up to ∼80% of total biomass) such that it may be a viable alternative to sugar synthesis from terrestrial plants, including sugarcane. PMID:22307292

Rerouting Carbon Flux To Enhance Photosynthetic Productivity

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

Ducat, DC; Avelar-Rivas, JA; Way, JC

2012-03-23

The bioindustrial production of fuels, chemicals, and therapeutics typically relies upon carbohydrate inputs derived from agricultural plants, resulting in the entanglement of food and chemical commodity markets. We demonstrate the efficient production of sucrose from a cyanobacterial species, Synechococcus elongatus, heterologously expressing a symporter of protons and sucrose (cscB). cscB-expressing cyanobacteria export sucrose irreversibly to concentrations of >10 mM without culture toxicity. Moreover, sucrose-exporting cyanobacteria exhibit increased biomass production rates relative to wild-type strains, accompanied by enhanced photosystem II activity, carbon fixation, and chlorophyll content. The genetic modification of sucrose biosynthesis pathways to minimize competing glucose-or sucrose-consuming reactions can furthermore » improve sucrose production, allowing the export of sucrose at rates of up to 36.1 mg liter(-1) h illumination(-1). This rate of production exceeds that of previous reports of targeted, photobiological production from microbes. Engineered S. elongatus produces sucrose in sufficient quantities (up to similar to 80% of total biomass) such that it may be a viable alternative to sugar synthesis from terrestrial plants, including sugarcane.« less

A conserved RNA structural element within the hepatitis B virus post-transcriptional regulatory element enhance nuclear export of intronless transcripts and repress the splicing mechanism.

PubMed

Visootsat, Akasit; Payungporn, Sunchai; T-Thienprasert, Nattanan P

2015-12-01

Hepatitis B virus (HBV) infection is a primary cause of hepatocellular carcinoma and liver cirrhosis worldwide. To develop novel antiviral drugs, a better understanding of HBV gene expression regulation is vital. One important aspect is to understand how HBV hijacks the cellular machinery to export unspliced RNA from the nucleus. The HBV post-transcriptional regulatory element (HBV PRE) has been proposed to be the HBV RNA nuclear export element. However, the function remains controversial, and the core element is unclear. This study, therefore, aimed to identify functional regulatory elements within the HBV PRE and investigate their functions. Using bioinformatics programs based on sequence conservation and conserved RNA secondary structures, three regulatory elements were predicted, namely PRE 1151-1410, PRE 1520-1620 and PRE 1650-1684. PRE 1151-1410 significantly increased intronless and unspliced luciferase activity in both HepG2 and COS-7 cells. Likewise, PRE 1151-1410 significantly elevated intronless and unspliced HBV surface transcripts in liver cancer cells. Moreover, motif analysis predicted that PRE 1151-1410 contains several regulatory motifs. This study reported the roles of PRE 1151-1410 in intronless transcript nuclear export and the splicing mechanism. Additionally, these results provide knowledge in the field of HBV RNA regulation. Moreover, PRE 1151-1410 may be used to enhance the expression of other mRNAs in intronless reporter plasmids.

Thermodynamic performance of multi-stage gradational lead screw vacuum pump

NASA Astrophysics Data System (ADS)

Zhao, Fan; Zhang, Shiwei; Sun, Kun; Zhang, Zhijun

2018-02-01

As a kind of dry mechanical vacuum pump, the twin-screw vacuum pump has an outstanding pumping performance during operation, widely used in the semiconductor industry. Compared with the constant lead screw (CLS) vacuum pump, the gradational lead screw (GLS) vacuum pump is more popularly applied in recent years. Nevertheless, not many comparative studies on the thermodynamic performance of GLS vacuum pump can be found in the literature. Our study focuses on one type of GLS vacuum pump, the multi-stage gradational lead screw (MGLS) vacuum pump, gives a detailed description of its construction and illustrates it with the drawing. Based on the structural analysis, the thermodynamic procedure is divided into four distinctive processes, including sucking process, transferring (compressing) process, backlashing process and exhausting process. The internal mechanism of each process is qualitatively illustrated and the mathematical expressions of seven thermodynamic parameters are given under the ideal situation. The performance curves of MGLS vacuum pump are plotted by MATLAB software and compared with those of the CLS vacuum pump in the same case. The results can well explain why the MGLS vacuum pump has more favorable pumping performance than the CLS vacuum pump in saving energy, reducing noise and heat dissipation.

Root bacterial endophytes confer drought resistance and enhance expression and activity of a vacuolar H+ -pumping pyrophosphatase in pepper plants.

PubMed

Vigani, Gianpiero; Rolli, Eleonora; Marasco, Ramona; Dell'Orto, Marta; Michoud, Grégoire; Soussi, Asma; Raddadi, Noura; Borin, Sara; Sorlini, Claudia; Zocchi, Graziano; Daffonchio, Daniele

2018-05-22

It has been previously shown that the transgenic overexpression of the plant root vacuolar proton pumps H + -ATPase (V-ATPase) and H + -PPase (V-PPase) confer tolerance to drought. Since plant-root endophytic bacteria can also promote drought tolerance, we hypothesize that such promotion can be associated to the enhancement of the host vacuolar proton pumps expression and activity. To test this hypothesis, we selected two endophytic bacteria endowed with an array of in vitro plant growth promoting traits. Their genome sequences confirmed the presence of traits previously shown to confer drought resistance to plants, such as the synthesis of nitric oxide and of organic volatile organic compounds. We used the two strains on pepper (Capsicuum annuum L.) because of its high sensitivity to drought. Under drought conditions, both strains stimulated a larger root system and enhanced the leaves' photosynthetic activity. By testing the expression and activity of the vacuolar proton pumps, H + -ATPase (V-ATPase) and H + -PPase (V-PPase), we found that bacterial colonization enhanced V-PPase only. We conclude that the enhanced expression and activity of V-PPase can be favoured by the colonization of drought-tolerance-inducing bacterial endophytes. This article is protected by copyright. All rights reserved. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

Long-Term Patterns in Production and Export of Fecal Pellets by Krill and Salps along the Western Antarctic Peninsula

NASA Astrophysics Data System (ADS)

Steinberg, D. K.; Ruck, K. E.; Cope, J. S.

2016-02-01

The Western Antarctic Peninsula (WAP) is one of the most rapidly warming regions on Earth, and where climate-induced changes in zooplankton abundance and species composition could dramatically affect the pelagic food web and biogeochemical cycling. We examined long-term (1993 to the present) and spatial trends in summer abundance of, and fecal pellet production (FPP) by, Antarctic krill (Euphausia superba) and gelatinous salps (Salpa thompsoni) and their relationship with physical and other environmental parameters. Zooplankton were collected as part of the Palmer, Antarctica Long-Term Ecological Research Program (PAL LTER) from the epipelagic zone in a region divided into latitudinal (North, South, and Far South) and cross-shelf (coastal, shelf, slope) sub-regions. Beginning in 2009, FPP and sinking rate experiments were conducted at representative stations along these gradients. FPP peaks occurred every 4-6 years in both species in the north and south, but alternated such that some years were characterized by high krill-mediated export, and others by high salp-mediated export. In the far south (where perennial sea ice still persists), and in both coastal and shelf sub-regions, krill FFP exceeded that of salps. Conversely, off the slope, salp FPP exceeded that of krill. Variability in krill FPP was strongly and positively influenced by primary production 2-years prior, and negatively correlated with sea surface temperature (no lag). Salp FPP was most significantly correlated with sea ice parameters, with highest FPP in years of lowest sea-ice extent, duration, and area. Warmer water and ice-free conditions favored salps over krill, which also increased overall potential export of fecal pellet carbon to depth. We discuss the implications of this potential increase in biological pump efficiency as the climate warms.

Mechanisms for the export of archaeal lipids down the water column in the upwelling area off Cape Blanc, North-West Africa

NASA Astrophysics Data System (ADS)

Ebersbach, Friederike; Goldenstein, Nadine; Iversen, Morten; Mollenhauer, Gesine; Hinrichs, Kai-Uwe

2016-04-01

Transport mechanisms of microbial membrane lipids from surface waters to the seafloor are poorly understood. In particular, pelagic archaeal glycerol dibiphytanyl glycerol tetraethers (GDGTs) from planktonic archaea are frequently used for reconstruction of ancient sea surface temperatures (Schouten et al. 2013). Because planktonic archaea are too small and neutrally buoyant to sink independently, transport vehicles for efficient export of fossil archaeal biomarkers to the sediment are required. The surface ocean is coupled with the deep ocean through biogenic sinking particles, a process known as the biological pump (Volk and Hoffert 1985). Two different pathways for particle formation, mainly taking place in the mesopelagic zone, are distinguished: Direct aggregation of phytoplankton blooms or grazing, resulting in phyto-detrital aggregates or reprocessed faecal material, respectively. Grazing and packaging into sinking particles is a possible export mechanism for GDGTs (Huguet et al. 2006). Moreover, it is assumed that phyto-detrital aggregates also play an important role in transporting GDGTs to the deep (Mollenhauer et al. 2015), but processes behind this pathway remain unclear. However, there are only few studies that link GDGT signals in sinking particles to the composition of the exported particulate matter (e.g. Yamamoto et al., 2012; Mollenhauer et al. 2015). Here we investigate sinking particles and suspended particulate matter (SPM) from spring blooms in 2012 and 2013 in the upwelling region in the Atlantic Ocean off Cape Blanc, Mauritania. We compare for the first time material from free-floating sediment traps (100, 200 and 400 m; purely sinking particles) with sinking particles and SPM from size fractionated in-situ pump (ISP) filters (several depths between 40 and 2350 m). This setup allows to relate the signal from archaeal lipids to (i) the flux of particulate organic carbon and the particle assemblages as revealed by the characterisation of thousands of individual particles collected in gels in addition to (ii) the sinking particles and SPM present in different particle size fractions on the filters. First results show that the large size fraction carries relatively more intact lipids indicating fresh material being attached to sinking particles rather than suspended in the water column. Furthermore, the distribution of 1G-GDGTs over depths differs from that of 2G- and HPH-GDGTs which might relate to different archaeal communities at different depths. Our findings contribute to the mechanistic understanding of the export of organic molecules through the water column and support the validation of lipid-based paleoceanographic proxies. References Huguet, C., Cartes, J.E., Sinninghe Damsté, J.S., Schouten, S., 2006. Marine crenarchaeotal membrane lipids in decapods: Implications for the TEX86 paleothermometer. Geochemistry, Geophysics, Geosystems 7 (11), Q11010. Mollenhauer, G., Basse, A., Kim, J.-H., Sinninghe Damsté, J.S., Fischer, G., 2015. A four-year record of UK'37- and TEX86-derived sea surface temperature estimates from sinking particles in the filamentous upwelling region off Cape Blanc, Mauritania. Deep Sea Research Part I: Oceanographic Research Papers 97, 67-79. doi:10.1016/j.dsr.2014.11.015 Schouten, S., Hopmans, E.C., Sinninghe Damsté, J.S., 2013. The organic geochemistry of glycerol dialkyl glycerol tetraether lipids: A review. Organic Geochemistry 54, 19-61. Volk, T., Hoffert, M.I., 1985. Ocean carbon pumps: analysis of relative strengths and efficiencies in ocean-driven atmospheric CO2 changes. Geophysical Monographs 32, 99-110. Yamamoto, M., Shimamoto, A., Fukuhara, T., Tanaka, Y., Ishizaka, J., 2012. Glycerol dialkyl glycerol tetraethers and TEX86 index in sinking particles in the western North Pacific. Organic Geochemistry 53 (0), 52-62.

IRON INCREASES EXPRESSION OF IRON-EXPORT PROTEIN MTP1 IN LUNG CELLS

EPA Science Inventory

Accumulation of reactive iron in acute and chronic lung disease suggests that iron-driven free radical formation could contribute to tissue injury. Safe transport and sequestration of this metal is likely to be of importance in lung defense. We provide evidence for the expression...

Bmp6 Expression in Murine Liver Non Parenchymal Cells: A Mechanism to Control their High Iron Exporter Activity and Protect Hepatocytes from Iron Overload?

PubMed Central

Rausa, Marco; Pagani, Alessia; Nai, Antonella; Campanella, Alessandro; Gilberti, Maria Enrica; Apostoli, Pietro; Camaschella, Clara; Silvestri, Laura

2015-01-01

Bmp6 is the main activator of hepcidin, the liver hormone that negatively regulates plasma iron influx by degrading the sole iron exporter ferroportin in enterocytes and macrophages. Bmp6 expression is modulated by iron but the molecular mechanisms are unknown. Although hepcidin is expressed almost exclusively by hepatocytes (HCs), Bmp6 is produced also by non-parenchymal cells (NPCs), mainly sinusoidal endothelial cells (LSECs). To investigate the regulation of Bmp6 in HCs and NPCs, liver cells were isolated from adult wild type mice whose diet was modified in iron content in acute or chronic manner and in disease models of iron deficiency (Tmprss6 KO mouse) and overload (Hjv KO mouse). With manipulation of dietary iron in wild-type mice, Bmp6 and Tfr1 expression in both HCs and NPCs was inversely related, as expected. When hepcidin expression is abnormal in murine models of iron overload (Hjv KO mice) and deficiency (Tmprss6 KO mice), Bmp6 expression in NPCs was not related to Tfr1. Despite the low Bmp6 in NPCs from Tmprss6 KO mice, Tfr1 mRNA was also low. Conversely, despite body iron overload and high expression of Bmp6 in NPCs from Hjv KO mice, Tfr1 mRNA and protein were increased. However, in the same cells ferritin L was only slightly increased, but the iron content was not, suggesting that Bmp6 in these cells reflects the high intracellular iron import and export. We propose that NPCs, sensing the iron flux, not only increase hepcidin through Bmp6 with a paracrine mechanism to control systemic iron homeostasis but, controlling hepcidin, they regulate their own ferroportin, inducing iron retention or release and further modulating Bmp6 production in an autocrine manner. This mechanism, that contributes to protect HC from iron loading or deficiency, is lost in disease models of hepcidin production. PMID:25860887

Gain statistics of a fiber optical parametric amplifier with a temporally incoherent pump.

PubMed

Xu, Y Q; Murdoch, S G

2010-03-15

We present an investigation of the statistics of the gain fluctuations of a fiber optical parametric amplifier pumped with a temporally incoherent pump. We derive a simple expression for the probability distribution of the gain of the amplified optical signal. The gain statistics are shown to be a strong function of the signal detuning and allow the possibility of generating optical gain distributions with controllable long-tails. Very good agreement is found between this theory and the experimentally measured gain distributions of an incoherently pumped amplifier.

Duration of breast milk expression among working mothers enrolled in an employer-sponsored lactation program.

PubMed

Ortiz, Joan; McGilligan, Kathryn; Kelly, Patricia

2004-01-01

Maternal employment has been one of the greatest barriers to breastfeeding. Women are increasingly solving this problem by expressing milk at work and taking it home to their infants. The objective was to determine duration of breast milk expression among working mothers enrolled in an employer-sponsored lactation program. Retrospective reviews were conducted on the lactation records of 462 women employed by 5 corporations in order to describe and characterize their experiences. The lactation program included the employees' choice of (a) a class on the benefits of breastfeeding; (b) services of a certified lactation consultant (CLC); and (c) private room in the workplace with equipment for pumping. Breastfeeding was initiated by 97.5% of the participants, with 57.8% continuing for at least 6 months. Of the 435 (94.2%) who returned to work after giving birth, 343 (78.9%) attempted pumping milk at work, and 336 (98%) were successful. They expressed milk in the workplace for a mean of 6.3 months (SD = 3.9, range 2 weeks to 21 months). The mean age of infants when the mothers stopped pumping at work was 9.1 months (SD = 4.1, range 1.9 to 25 months). Most of the women who pumped their milk at work were working full time (84.2%). The mean postnatal maternity leave was 2.8 months. The proportion of women who chose to pump at work was higher among women who were salaried than among those who were paid hourly wages (p < 0.01). Company-sponsored lactation programs can enable employed mothers to provide breast milk for their infants as long as they wish, thus helping the nation attain the Healthy People 2010 goals of 50% of mothers breastfeeding until their infants are 6-months-old.

Time dependent temperature distribution in pulsed Ti:sapphire lasers

NASA Technical Reports Server (NTRS)

Buoncristiani, A. Martin; Byvik, Charles E.; Farrukh, Usamah O.

1988-01-01

An expression is derived for the time dependent temperature distribution in a finite solid state laser rod for an end-pumped beam of arbitrary shape. The specific case of end pumping by circular (constant) or Gaussian beam is described. The temperature profile for a single pump pulse and for repetitive pulse operation is discussed. The particular case of the temperature distribution in a pulsed titanium:sapphire rod is considered.

Regulating Intracellular Calcium in Plants: From Molecular Genetics to Physiology

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

Heven Sze

To grow, develop, adapt, and reproduce, plants have evolved mechanisms to regulate the uptake, translocation and sorting of calcium ions into different cells and subcellular compartments. Yet how plants accomplish this remarkable feat is still poorly understood. The spatial and temporal changes in intracellular [Ca2+] during growth and during responses to hormonal and environmental stimuli indicate that Ca2+ influx and efflux transporters are diverse and tightly regulated in plants. The specific goals were to determine the biological roles of multiple Ca pumps (ECAs) in the model plant Arabidopsis thaliana. We had pioneered the use of K616 yeast strain to functionallymore » express plant Ca pumps, and demonstrated two distinct types of Ca pumps in plants (Sze et al., 2000. Annu Rev Plant Biol. 51,433). ACA2 represented one type that was auto-inhibited by the N-terminal region and stimulated by calmodulin. ECA1 represented another type that was not sensitive to calmodulin and phylogenetically distinct from ACAs. The goal to determine the biological roles of multiple ECA-type Ca pumps in Arabidopsis has been accomplished. Although we demonstrated ECA1 was a Ca pump by functional expression in yeast, the in vivo roles of ECAs was unclear. A few highlights are described. ECA1 and/or ECA4 are Ca/Mn pumps localized to the ER and are highly expressed in all cell types. Using homozygous T-DNA insertional mutants of eca1, we demonstrated that the ER-bound ECA1 supports growth and confers tolerance of plants growing on medium low in Ca or containing toxic levels of Mn. This is the first genetic study to determine the in vivo function of a Ca pump in plants. A phylogenetically distinct ECA3 is also a Ca/Mn pump that is localized to endosome, such as post-Golgi compartments. Although it is expressed at lower levels than ECA1, eca3 mutants are impaired in Ca-dependent root growth and in pollen tube elongation. Increased secretion of wall proteins in mutants suggests that Ca and Mn homeostasis in post-Golgi compartments are critical for secretory activities. Moreover, perturbation of the secretory machinery limits growth possibly by upsetting the synthesis, processing and assembly of cell wall components. Analyses of whole genome transcriptome of pollen shows that a subset of Ca pump genes are developmentally regulated. Each ECA Ca pump is localized to distinct endomembrane compartments and regulate Ca and Mn homeostasis required for optimal growth and for tolerance to high Mn stress. Ca and Mn levels within endomembrane lumen appear to be critical for activities of the secretory machinery including post-Golgi compartments that coordinate membrane traffic and sorting of materials to the vacuole and the cell wall. Significance: Thus sorting of Ca/Mn by ECA pumps in endomembranes is critical for membrane trafficking pattern which serves as a central coordinator of plant growth, development and adaptation to abiotic and biotic stress.« less

Selective export of autotaxin from the endoplasmic reticulum.

PubMed

Lyu, Lin; Wang, Baolu; Xiong, Chaoyang; Zhang, Xiaotian; Zhang, Xiaoyan; Zhang, Junjie

2017-04-28

Autotaxin (ATX) or ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2) is a secretory glycoprotein and functions as the key enzyme for lysophosphatidic acid generation. The mechanism of ATX protein trafficking is largely unknown. Here, we demonstrated that p23, a member of the p24 protein family, was the protein-sorting receptor required for endoplasmic reticulum (ER) export of ATX. A di-phenylalanine (Phe-838/Phe-839) motif in the human ATX C-terminal region was identified as a transport signal essential for the ATX-p23 interaction. Knockdown of individual Sec24 isoforms by siRNA revealed that ER export of ATX was impaired only if Sec24C was down-regulated. These results suggest that ATX is selectively exported from the ER through a p23, Sec24C-dependent pathway. In addition, it was found that AKT signaling played a role in ATX secretion regulation to facilitate ATX ER export by enhancing the nuclear factor of activated T cell-mediated p23 expression. Furthermore, the di-hydrophobic amino acid motifs (FY) also existed in the C-terminal regions of human ENPP1 and ENPP3. Such a p23, Sec24C-dependent selective ER export mechanism is conserved among these ENPP family members. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

AKT3 controls mitochondrial biogenesis and autophagy via regulation of the major nuclear export protein CRM-1

PubMed Central

Corum, Daniel G.; Tsichlis, Philip N.; Muise-Helmericks, Robin C.

2014-01-01

Our previous work has shown that Akt3 is required for mitochondrial biogenesis in primary human endothelial cells (ECs) and in Akt3-null mice; Akt3 affects subcellular localization of peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1α), the master regulator of mitochondrial biogenesis. The purpose of this study is to determine the mechanism by which Akt3 controls the subcellular distribution of PGC-1α and to explore the effect on mitochondrial biogenesis and turnover during angiogenesis. Here we use standard biochemical analyses and Akt3-knockdown strategies to show that Akt3 controls the stabilization of chromosome maintenance region-1 (CRM-1), the major nuclear export receptor. Site-directed mutagenesis and association analyses show that PGC-1α nuclear export is CRM-1 dependent. Akt3 knockdown and CRM-1 overexpression cause 3-fold reductions in PGC-1α target gene expression, compared to control levels. Akt3 inhibition causes autophagy, as measured by autophagosome formation, in a CRM-1-dependent, Akt1/mTOR-independent pathway. In vivo, Akt3-null and heterozygous mice show dose-dependent decreases in angiogenesis compared to wild-type littermates (∼5- and 2.5-fold decreases, respectively), as assessed by Matrigel plug assays. This correlates with an ∼1.5-fold decrease in mitochondrial Cox IV expression. Our studies suggest that Akt3 is a regulator of mitochondrial dynamics in the vasculature via regulation of CRM-1-dependent nuclear export.—Corum, D. G., Tsichlis, P. N., Muise-Helmericks, R. C. AKT3 controls mitochondrial biogenesis and autophagy via regulation of the major nuclear export protein CRM-1. PMID:24081905

Leukemia-Associated Nup214 Fusion Proteins Disturb the XPO1-Mediated Nuclear-Cytoplasmic Transport Pathway and Thereby the NF-κB Signaling Pathway.

PubMed

Saito, Shoko; Cigdem, Sadik; Okuwaki, Mitsuru; Nagata, Kyosuke

2016-07-01

Nuclear-cytoplasmic transport through nuclear pore complexes is mediated by nuclear transport receptors. Previous reports have suggested that aberrant nuclear-cytoplasmic transport due to mutations or overexpression of nuclear pore complexes and nuclear transport receptors is closely linked to diseases. Nup214, a component of nuclear pore complexes, has been found as chimeric fusion proteins in leukemia. Among various Nup214 fusion proteins, SET-Nup214 and DEK-Nup214 have been shown to be engaged in tumorigenesis, but their oncogenic mechanisms remain unclear. In this study, we examined the functions of the Nup214 fusion proteins by focusing on their effects on nuclear-cytoplasmic transport. We found that SET-Nup214 and DEK-Nup214 interact with exportin-1 (XPO1)/CRM1 and nuclear RNA export factor 1 (NXF1)/TAP, which mediate leucine-rich nuclear export signal (NES)-dependent protein export and mRNA export, respectively. SET-Nup214 and DEK-Nup214 decreased the XPO1-mediated nuclear export of NES proteins such as cyclin B and proteins involved in the NF-κB signaling pathway by tethering XPO1 onto nuclear dots where Nup214 fusion proteins are localized. We also demonstrated that SET-Nup214 and DEK-Nup214 expression inhibited NF-κB-mediated transcription by abnormal tethering of the complex containing p65 and its inhibitor, IκB, in the nucleus. These results suggest that SET-Nup214 and DEK-Nup214 perturb the regulation of gene expression through alteration of the nuclear-cytoplasmic transport system. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Nascent chain-monitored remodeling of the Sec machinery for salinity adaptation of marine bacteria

PubMed Central

Ishii, Eiji; Chiba, Shinobu; Hashimoto, Narimasa; Kojima, Seiji; Homma, Michio; Ito, Koreaki; Akiyama, Yoshinori; Mori, Hiroyuki

2015-01-01

SecDF interacts with the SecYEG translocon in bacteria and enhances protein export in a proton-motive-force-dependent manner. Vibrio alginolyticus, a marine-estuarine bacterium, contains two SecDF paralogs, V.SecDF1 and V.SecDF2. Here, we show that the export-enhancing function of V.SecDF1 requires Na+ instead of H+, whereas V.SecDF2 is Na+-independent, presumably requiring H+. In accord with the cation-preference difference, V.SecDF2 was only expressed under limited Na+ concentrations whereas V.SecDF1 was constitutive. However, it is not the decreased concentration of Na+ per se that the bacterium senses to up-regulate the V.SecDF2 expression, because marked up-regulation of the V.SecDF2 synthesis was observed irrespective of Na+ concentrations under certain genetic/physiological conditions: (i) when the secDF1VA gene was deleted and (ii) whenever the Sec export machinery was inhibited. VemP (Vibrio export monitoring polypeptide), a secretory polypeptide encoded by the upstream ORF of secDF2VA, plays the primary role in this regulation by undergoing regulated translational elongation arrest, which leads to unfolding of the Shine–Dalgarno sequence for translation of secDF2VA. Genetic analysis of V. alginolyticus established that the VemP-mediated regulation of SecDF2 is essential for the survival of this marine bacterium in low-salinity environments. These results reveal that a class of marine bacteria exploits nascent-chain ribosome interactions to optimize their protein export pathways to propagate efficiently under different ionic environments that they face in their life cycles. PMID:26392525

Mothers' experiences expressing breast milk for their preterm infants: does NICU design make a difference?

PubMed

Dowling, Donna A; Blatz, Mary Ann; Graham, Gregory

2012-12-01

This study examined differences in outcomes of provision of mothers' milk before and after implementation of a single-family room (SFR) neonatal intensive care unit (NICU) and described issues related to long-term milk expression. The sample included 40 mothers (15 in the original NICU and 25 in the SFR NICU). Descriptive comparative. Mothers were recruited 2 months before and 3 months after opening an SFR NICU. Nutritional data were collected throughout hospitalization. Mothers used a milk expression diary during hospitalization and completed a survey, "My Experiences With Milk Expression" immediately before infant discharge. Seventy-five percent of mothers planned to express breast milk or breastfeed before delivery. The majority of the mothers (55%) were most comfortable pumping in their own homes because of the increased privacy. There were no statistically significant differences between the 2 groups regarding the place where they were most comfortable pumping or where they usually pumped, although more mothers pumped in their babies' rooms in the SFR NICU. The majority of the mothers reported concern about their milk supply at some time during hospitalization and 47.5% reported having breast problems. At discharge, 71.8% of the total group was providing some breast milk and 44.7% of the total group was providing breast milk exclusively. There were no significant differences between the groups in outcomes concerning the provision of breast milk. Individual mother's needs for privacy need to be determined and interventions to support mothers' feeding plans throughout hospitalization and at discharge need to be developed.

A high-throughput screening system targeting the nuclear export pathway via the third nuclear export signal of influenza A virus nucleoprotein.

PubMed

Kakisaka, Michinori; Mano, Takafumi; Aida, Yoko

2016-06-02

Two classes of antiviral drugs, M2 channel inhibitors and neuraminidase (NA) inhibitors, are currently approved for the treatment of influenza; however, the development of resistance against these agents limits their efficacy. Therefore, the identification of new targets and the development of new antiviral drugs against influenza are urgently needed. The third nuclear export signal (NES3) of nucleoprotein (NP) is the most important for viral replication among seven NESs encoded by four viral proteins, NP, M1, NS1, and NS2. NP-NES3 is critical for the nuclear export of NP, and targeting NP-NES3 is therefore a promising strategy that may lead to the development of antiviral drugs. However, a high-throughput screening (HTS) system to identify inhibitors of NP nuclear export has not been established. Here, we developed a novel HTS system to evaluate the inhibitory effects of compounds on the nuclear export pathway mediated by NP-NES3 using a MDCK cell line stably expressing NP-NES3 fused to a green fluorescent protein from aequorea coerulescens (AcGFP-NP-NES3) and a cell imaging analyzer. This HTS system was used to screen a 9600-compound library, leading to the identification of several hit compounds with inhibitory activity against the nuclear export of AcGFP-NP-NES3. The present HTS system provides a useful strategy for the identification of inhibitors targeting the nuclear export of NP via its NES3 sequence. Copyright © 2016. Published by Elsevier B.V.

Glacial-interglacial atmospheric CO2 change: a possible "standing volume" effect on deep-ocean carbon sequestration

NASA Astrophysics Data System (ADS)

Skinner, L. C.

2009-09-01

So far, the exploration of possible mechanisms for glacial atmospheric CO2 drawdown and marine carbon sequestration has tended to focus on dynamic or kinetic processes (i.e. variable mixing-, equilibration- or export rates). Here an attempt is made to underline instead the possible importance of changes in the standing volumes of intra-oceanic carbon reservoirs (i.e. different water-masses) in influencing the total marine carbon inventory. By way of illustration, a simple mechanism is proposed for enhancing the marine carbon inventory via an increase in the volume of relatively cold and carbon-enriched deep water, analogous to modern Lower Circumpolar Deep Water (LCDW), filling the ocean basins. A set of simple box-model experiments confirm the expectation that a deep sea dominated by an expanded LCDW-like watermass holds more CO2, without any pre-imposed changes in ocean overturning rate, biological export or ocean-atmosphere exchange. The magnitude of this "standing volume effect" (which operates by boosting the solubility- and biological pumps) might be as large as the contributions that have previously been attributed to carbonate compensation, terrestrial biosphere reduction or ocean fertilisation for example. By providing a means of not only enhancing but also driving changes in the efficiency of the biological- and solubility pumps, this standing volume mechanism may help to reduce the amount of glacial-interglacial CO2 change that remains to be explained by other mechanisms that are difficult to assess in the geological archive, such as reduced mass transport or mixing rates in particular. This in turn could help narrow the search for forcing conditions capable of pushing the global carbon cycle between glacial and interglacial modes.

Structure based classification for bile salt export pump (BSEP) inhibitors using comparative structural modeling of human BSEP

NASA Astrophysics Data System (ADS)

Jain, Sankalp; Grandits, Melanie; Richter, Lars; Ecker, Gerhard F.

2017-06-01

The bile salt export pump (BSEP) actively transports conjugated monovalent bile acids from the hepatocytes into the bile. This facilitates the formation of micelles and promotes digestion and absorption of dietary fat. Inhibition of BSEP leads to decreased bile flow and accumulation of cytotoxic bile salts in the liver. A number of compounds have been identified to interact with BSEP, which results in drug-induced cholestasis or liver injury. Therefore, in silico approaches for flagging compounds as potential BSEP inhibitors would be of high value in the early stage of the drug discovery pipeline. Up to now, due to the lack of a high-resolution X-ray structure of BSEP, in silico based identification of BSEP inhibitors focused on ligand-based approaches. In this study, we provide a homology model for BSEP, developed using the corrected mouse P-glycoprotein structure (PDB ID: 4M1M). Subsequently, the model was used for docking-based classification of a set of 1212 compounds (405 BSEP inhibitors, 807 non-inhibitors). Using the scoring function ChemScore, a prediction accuracy of 81% on the training set and 73% on two external test sets could be obtained. In addition, the applicability domain of the models was assessed based on Euclidean distance. Further, analysis of the protein-ligand interaction fingerprints revealed certain functional group-amino acid residue interactions that could play a key role for ligand binding. Though ligand-based models, due to their high speed and accuracy, remain the method of choice for classification of BSEP inhibitors, structure-assisted docking models demonstrate reasonably good prediction accuracies while additionally providing information about putative protein-ligand interactions.

Conserved Allosteric Hot Spots in the Transmembrane Domains of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channels and Multidrug Resistance Protein (MRP) Pumps*

PubMed Central

Wei, Shipeng; Roessler, Bryan C.; Chauvet, Sylvain; Guo, Jingyu; Hartman, John L.; Kirk, Kevin L.

2014-01-01

ATP-binding cassette (ABC) transporters are an ancient family of transmembrane proteins that utilize ATPase activity to move substrates across cell membranes. The ABCC subfamily of the ABC transporters includes active drug exporters (the multidrug resistance proteins (MRPs)) and a unique ATP-gated ion channel (cystic fibrosis transmembrane conductance regulator (CFTR)). The CFTR channel shares gating principles with conventional ligand-gated ion channels, but the allosteric network that couples ATP binding at its nucleotide binding domains (NBDs) with conformational changes in its transmembrane helices (TMs) is poorly defined. It is also unclear whether the mechanisms that govern CFTR gating are conserved with the thermodynamically distinct MRPs. Here we report a new class of gain of function (GOF) mutation of a conserved proline at the base of the pore-lining TM6. Multiple substitutions of this proline promoted ATP-free CFTR activity and activation by the weak agonist, 5′-adenylyl-β,γ-imidodiphosphate (AMP-PNP). TM6 proline mutations exhibited additive GOF effects when combined with a previously reported GOF mutation located in an outer collar of TMs that surrounds the pore-lining TMs. Each TM substitution allosterically rescued the ATP sensitivity of CFTR gating when introduced into an NBD mutant with defective ATP binding. Both classes of GOF mutations also rescued defective drug export by a yeast MRP (Yor1p) with ATP binding defects in its NBDs. We conclude that the conserved TM6 proline helps set the energy barrier to both CFTR channel opening and MRP-mediated drug efflux and that CFTR channels and MRP pumps utilize similar allosteric mechanisms for coupling conformational changes in their translocation pathways to ATP binding at their NBDs. PMID:24876383

Conserved allosteric hot spots in the transmembrane domains of cystic fibrosis transmembrane conductance regulator (CFTR) channels and multidrug resistance protein (MRP) pumps.

PubMed

Wei, Shipeng; Roessler, Bryan C; Chauvet, Sylvain; Guo, Jingyu; Hartman, John L; Kirk, Kevin L

2014-07-18

ATP-binding cassette (ABC) transporters are an ancient family of transmembrane proteins that utilize ATPase activity to move substrates across cell membranes. The ABCC subfamily of the ABC transporters includes active drug exporters (the multidrug resistance proteins (MRPs)) and a unique ATP-gated ion channel (cystic fibrosis transmembrane conductance regulator (CFTR)). The CFTR channel shares gating principles with conventional ligand-gated ion channels, but the allosteric network that couples ATP binding at its nucleotide binding domains (NBDs) with conformational changes in its transmembrane helices (TMs) is poorly defined. It is also unclear whether the mechanisms that govern CFTR gating are conserved with the thermodynamically distinct MRPs. Here we report a new class of gain of function (GOF) mutation of a conserved proline at the base of the pore-lining TM6. Multiple substitutions of this proline promoted ATP-free CFTR activity and activation by the weak agonist, 5'-adenylyl-β,γ-imidodiphosphate (AMP-PNP). TM6 proline mutations exhibited additive GOF effects when combined with a previously reported GOF mutation located in an outer collar of TMs that surrounds the pore-lining TMs. Each TM substitution allosterically rescued the ATP sensitivity of CFTR gating when introduced into an NBD mutant with defective ATP binding. Both classes of GOF mutations also rescued defective drug export by a yeast MRP (Yor1p) with ATP binding defects in its NBDs. We conclude that the conserved TM6 proline helps set the energy barrier to both CFTR channel opening and MRP-mediated drug efflux and that CFTR channels and MRP pumps utilize similar allosteric mechanisms for coupling conformational changes in their translocation pathways to ATP binding at their NBDs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

[Design and verification of Luo-Ye pump-based stress formation for cultivation of tissue-engineered blood vessel].

PubMed

Liao, Wen-Jun; Chen, Wan-Wen; Wen, Zhang; Wu, Yue-Heng; Li, Dong-Feng; Zhou, Jia-Hui; Zheng, Jian-Yi; Lin, Zhan-Yi

2016-06-20

To improve Luo-Ye pump-based stress-forming system and optimize the stimulating effect on smooth muscle cells during cultivation of tissue-engineered blood vessels (TEBV). A new Luo-Ye pump-based TEBV 3D culture system was developed by adding an air pump to the output of the bioreactor. A pressure guide wire was used to measure the stress at different points of the silicone tube inside the TEBV bio-reactor, and fitting curves of the stress changes over time was created using Origin 8.0 software. The TEBVs were constructed by seeding vascular smooth muscle cells (VSMCs) isolated from human umbilical artery on polyglycolic acid (PGA) and cultured under dynamic conditions with 40 mmHg resistance (improved group), dynamic conditions without resistance (control group) or static condition (static group) for 4 weeks. The harvested TEBVs were then examined with HE staining, masson staining, α-SMA immunohistochemical staining, and scanning and transmission electron microscopy with semi-quantitative analysis of collagen content and α-SMA expression. The measured stress values and the fitting curves showed that the stress stimuli from the Luo-Ye pump were enhanced by adding an air pump to the output of the bioreactor. Histological analysis revealed improved VSMC density, collagen content and α-SMA expression in the TEBVs constructed with the improved method as compared with those in the control and static groups. Adding an air pump to the Luo-Ye pump significantly enhances the stress stimulation in the TEBV 3-D culture system to promote the secretion function of VSMCs.

Overexpression of the ABC transporter AvtAB increases avermectin production in Streptomyces avermitilis.

PubMed

Qiu, Jingfan; Zhuo, Ying; Zhu, Dongqing; Zhou, Xiufen; Zhang, Lixin; Bai, Linquan; Deng, Zixin

2011-10-01

Avermectins are 16-membered macrocyclic polyketides with potent antiparasitic activities, produced by Streptomyces avermitilis. Upstream of the avermectin biosynthetic gene cluster, there is the avtAB operon encoding the ABC transporter AvtAB, which is highly homologous to the mammalian multidrug efflux pump P-glycoprotein (Pgp). Inactivation of avtAB had no effect, but increasing the concentration of avtAB mRNA 30-500-fold, using a multi-copy plasmid in S. avermitilis, enhanced avermectin production about two-fold both in the wild-type and in a high-yield producer strain on agar plates. In liquid industrial fermentation medium, the overall productivity of avermectin B1a in the engineered high-yield producer was improved for about 50%, from 3.3 to 4.8 g/l. In liquid YMG medium, moreover, the ratio of intracellular to extracellular accumulation of avermectin B1a was dropped from 6:1 to 4.5:1 in response to multiple copies of avtAB. Additionally, the overexpression of avtAB did not cause any increased expression of the avermectin biosynthetic genes through RT-PCR analysis. We propose that the AvtAB transporter exports avermectin, and thus reduces the feedback inhibition on avermectin production inside the cell. This strategy may be useful for enhancing the production of other antibiotics.

Modelling of OPNMR phenomena using photon energy-dependent 〈Sz〉 in GaAs and InP

NASA Astrophysics Data System (ADS)

Wheeler, Dustin D.; Willmering, Matthew M.; Sesti, Erika L.; Pan, Xingyuan; Saha, Dipta; Stanton, Christopher J.; Hayes, Sophia E.

2016-12-01

We have modified the model for optically-pumped NMR (OPNMR) to incorporate a revised expression for the expectation value of the z-projection of the electron spin, 〈Sz 〉 and apply this model to both bulk GaAs and a new material, InP. This expression includes the photon energy dependence of the electron polarization when optically pumping direct-gap semiconductors in excess of the bandgap energy, Eg . Rather than using a fixed value arising from coefficients (the matrix elements) for the optical transitions at the k = 0 bandedge, we define a new parameter, Sopt (Eph) . Incorporating this revised element into the expression for 〈Sz 〉 , we have simulated the photon energy dependence of the OPNMR signals from bulk semi-insulating GaAs and semi-insulating InP. In earlier work, we matched calculations of electron spin polarization (alone) to features in a plot of OPNMR signal intensity versus photon energy for optical pumping (Ramaswamy et al., 2010). By incorporating an electron spin polarization which varies with pump wavelength into the penetration depth model of OPNMR signal, we are able to model features in both III-V semiconductors. The agreement between the OPNMR data and the corresponding model demonstrates that fluctuations in the OPNMR intensity have particular sensitivity to light hole-to-conduction band transitions in bulk systems. We provide detailed plots of the theoretical predictions for optical pumping transition probabilities with circularly-polarized light for both helicities of light, broken down into illustrative plots of optical magnetoabsorption and spin polarization, shown separately for heavy-hole and light-hole transitions. These plots serve as an effective roadmap of transitions, which are helpful to other researchers investigating optical pumping effects.

The MerR-like regulator BrlR confers biofilm tolerance by activating multidrug efflux pumps in Pseudomonas aeruginosa biofilms.

PubMed

Liao, Julie; Schurr, Michael J; Sauer, Karin

2013-08-01

A defining characteristic of biofilms is antibiotic tolerance that can be up to 1,000-fold greater than that of planktonic cells. In Pseudomonas aeruginosa, biofilm tolerance to antimicrobial agents requires the biofilm-specific MerR-type transcriptional regulator BrlR. However, the mechanism by which BrlR mediates biofilm tolerance has not been elucidated. Genome-wide transcriptional profiling indicated that brlR was required for maximal expression of genes associated with antibiotic resistance, in particular those encoding the multidrug efflux pumps MexAB-OprM and MexEF-OprN. Chromatin immunoprecipitation (ChIP) analysis revealed a direct regulation of these genes by BrlR, with DNA binding assays confirming BrlR binding to the promoter regions of the mexAB-oprM and mexEF-oprN operons. Quantitative reverse transcriptase PCR (qRT-PCR) analysis further indicated BrlR to be an activator of mexAB-oprM and mexEF-oprN gene expression. Moreover, immunoblot analysis confirmed increased MexA abundance in cells overexpressing brlR. Inactivation of both efflux pumps rendered biofilms significantly more susceptible to five different classes of antibiotics by affecting MIC but not the recalcitrance of biofilms to killing by bactericidal agents. Overexpression of either efflux pump in a ΔbrlR strain partly restored tolerance of ΔbrlR biofilms to antibiotics. Expression of brlR in mutant biofilms lacking both efflux pumps partly restored antimicrobial tolerance of biofilms to wild-type levels. Our results indicate that BrlR acts as an activator of multidrug efflux pumps to confer tolerance to P. aeruginosa biofilms and to resist the action of antimicrobial agents.

Modelling of OPNMR phenomena using photon energy-dependent 〈Sz〉 in GaAs and InP.

PubMed

Wheeler, Dustin D; Willmering, Matthew M; Sesti, Erika L; Pan, Xingyuan; Saha, Dipta; Stanton, Christopher J; Hayes, Sophia E

2016-12-01

We have modified the model for optically-pumped NMR (OPNMR) to incorporate a revised expression for the expectation value of the z-projection of the electron spin, 〈S z 〉 and apply this model to both bulk GaAs and a new material, InP. This expression includes the photon energy dependence of the electron polarization when optically pumping direct-gap semiconductors in excess of the bandgap energy, E g . Rather than using a fixed value arising from coefficients (the matrix elements) for the optical transitions at the k=0 bandedge, we define a new parameter, S opt (E ph ). Incorporating this revised element into the expression for 〈S z 〉, we have simulated the photon energy dependence of the OPNMR signals from bulk semi-insulating GaAs and semi-insulating InP. In earlier work, we matched calculations of electron spin polarization (alone) to features in a plot of OPNMR signal intensity versus photon energy for optical pumping (Ramaswamy et al., 2010). By incorporating an electron spin polarization which varies with pump wavelength into the penetration depth model of OPNMR signal, we are able to model features in both III-V semiconductors. The agreement between the OPNMR data and the corresponding model demonstrates that fluctuations in the OPNMR intensity have particular sensitivity to light hole-to-conduction band transitions in bulk systems. We provide detailed plots of the theoretical predictions for optical pumping transition probabilities with circularly-polarized light for both helicities of light, broken down into illustrative plots of optical magnetoabsorption and spin polarization, shown separately for heavy-hole and light-hole transitions. These plots serve as an effective roadmap of transitions, which are helpful to other researchers investigating optical pumping effects. Copyright © 2016 Elsevier Inc. All rights reserved.

The manifold phospholipases A of Legionella pneumophila - identification, export, regulation, and their link to bacterial virulence.

PubMed

Banerji, Sangeeta; Aurass, Philipp; Flieger, Antje

2008-04-01

The intracellular lung pathogen Legionella pneumophila expresses secreted and cell-associated phospholipase A (PLA) and lysophospholipase A (LPLA) activities belonging to at least three enzyme families. The first family consists of three secreted PLA and LPLA activities displaying the amino acid signature motif 'GDSL'; PlaA, PlaC and PlaD. The second group contains the cell-associated and very potent PLA/LPLA, PlaB. The third group, the patatin-like proteins, comprises 11 members. One patatin-like protein, PatA/VipD, shows LPLA and PLA activities and interferes with vesicular trafficking when expressed in yeast and therefore is possibly involved in the intracellular infection process. Likewise, members of the first two phospholipase families have roles in bacterial virulence because phospholipases are important virulence factors that have been shown to promote bacterial survival, spread and host cell modification/damage. The GDSL enzyme PlaA detoxifies cytolytic lysophospholipids, and PlaB shows contact-dependent haemolytic activity. PlaC acylates cholesterol, a lipid present in eukaryotic hosts but not in the bacterium. Many of the L. pneumophila PLAs are exported by the type II Lsp or the type IVB Dot/Icm secretion systems involved in virulence factor export. Moreover, the regulation of lipolytic activities depends on the transcriptional regulators LetA/S and RpoS, inducing the expression of virulence traits, and on posttranscriptional activators like the zinc metalloprotease ProA.

Carbon export in the naturally iron-fertilized Kerguelen area of the Southern Ocean based on the 234Th approach

NASA Astrophysics Data System (ADS)

Planchon, F.; Ballas, D.; Cavagna, A.-J.; Bowie, A. R.; Davies, D.; Trull, T.; Laurenceau-Cornec, E. C.; Van Der Merwe, P.; Dehairs, F.

2015-06-01

This study examined upper-ocean particulate organic carbon (POC) export using the 234Th approach as part of the second KErguelen Ocean and Plateau compared Study expedition (KEOPS2). Our aim was to characterize the spatial and the temporal variability of POC export during austral spring (October-November 2011) in the Fe-fertilized area of the Kerguelen Plateau region. POC export fluxes were estimated at high productivity sites over and downstream of the plateau and compared to a high-nutrient low-chlorophyll (HNLC) area upstream of the plateau in order to assess the impact of iron-induced productivity on the vertical export of carbon. Deficits in 234Th activities were observed at all stations in surface waters, indicating early scavenging by particles in austral spring. 234Th export was lowest at the reference station R-2 and highest in the recirculation region (E stations) where a pseudo-Lagrangian survey was conducted. In comparison 234Th export over the central plateau and north of the polar front (PF) was relatively limited throughout the survey. However, the 234Th results support that Fe fertilization increased particle export in all iron-fertilized waters. The impact was greatest in the recirculation feature (3-4 fold at 200 m depth, relative to the reference station), but more moderate over the central Kerguelen Plateau and in the northern plume of the Kerguelen bloom (~2-fold at 200 m depth). The C : Th ratio of large (>53 μm) potentially sinking particles collected via sequential filtration using in situ pumping (ISP) systems was used to convert the 234Th flux into a POC export flux. The C : Th ratios of sinking particles were highly variable (3.1 ± 0.1 to 10.5 ± 0.2 μmol dpm-1) with no clear site-related trend, despite the variety of ecosystem responses in the fertilized regions. C : Th ratios showed a decreasing trend between 100 and 200 m depth suggesting preferential carbon loss relative to 234Th possibly due to heterotrophic degradation and/or grazing activity. C : Th ratios of sinking particles sampled with drifting sediment traps in most cases showed very good agreement with ratios for particles collected via ISP deployments (>53 μm particles). Carbon export production varied between 3.5 ± 0.9 and 11.8 ± 1.3 mmol m-2 d-1 from the upper 100 m and between 1.8 ± 0.9 and 8.2 ± 0.9 mmol m-2 d-1 from the upper 200 m. The highest export production was found inside the PF meander with a range of 5.3 ± 1.0 to 11.8 ± 1.1 mmol m-2 d-1 over the 19-day survey period. The impact of Fe fertilization is highest inside the PF meander with 2.9-4.5-fold higher carbon flux at 200 m depth in comparison to the HNLC control station. The impact of Fe fertilization was significantly less over the central plateau (stations A3 and E-4W) and in the northern branch of the bloom (station F-L) with 1.6-2.0-fold higher carbon flux compared to the reference station R. Export efficiencies (ratio of export to primary production and ratio of export to new production) were particularly variable with relatively high values in the recirculation feature (6 to 27 %, respectively) and low values (1 to 5 %, respectively) over the central plateau (station A3) and north of the PF (station F-L), indicating spring biomass accumulation. Comparison with KEOPS1 results indicated that carbon export production is much lower during the onset of the bloom in austral spring than during the peak and declining phases in late summer.

Spatiotemporal Variability in Particulate Organic Carbon Export Observed Using Bio-Optical Profiling Floats

NASA Astrophysics Data System (ADS)

Estapa, M. L.

2016-02-01

Autonomous, bio-optical profiling floats are poised to broaden the number and spatiotemporal resolution of observations of the ocean's biological pump. Here, we used multiple optical sensors aboard two bio-optical profiling floats (Navis BGCi, Sea-Bird) deployed in the Sargasso Sea to derive in situ proxies for particulate carbon (PC) flux, sub-mixed layer net community production (NCP) and to drive a model of net primary production (NPP). Profiles were collected at approximately 2-day resolution, and drift-phase PC flux observations were collected at subdaily resolution at a rotating cycle of observation depths between 150 and 1000 m. The magnitudes of NPP, PC flux, and their annually-averaged ratio were generally consistent with observations at the nearby Bermuda Atlantic Timeseries Study (BATS) site. PC flux and the export ratio were enhanced in the autumn as well as in the spring, and varied over short timescales possibly due to the influence of mesoscale eddies. The relatively shallow park depths and short profile cycle lengths allow us to identify ephemeral, subsurface bio-optical features and compare them to measured fluxes and satellite-observed surface properties.

Restoration of chemosensitivity in cancer cells with MDR phenotype by deoxyribozyme, compared with ribozyme.

PubMed

Xing, Ai-Yan; Shi, Duan-bo; Liu, Wei; Chen, Xu; Sun, Yan-Lin; Wang, Xiao; Zhang, Jian-ping; Gao, Peng

2013-06-01

One of the main mechanisms for multidrug resistance (MDR) involves multidrug resistance gene 1 (MDR1) which encodes P-glycoprotein (Pgp). Pgp acts as a drug efflux pump and exports chemotherapeutic agents from cancer cells. Specific inhibition of Pgp expression by gene therapy is considered a well-respective strategy having less innate toxicities. At present, the investigation of DRz in reversal MDR is scarce. In the study, phosphorothioate DRz that targets to the translation initiation codon AUG was synthesized and transfected into breast cancer cells and leukemia cells with MDR phenotype. ASODN (antisense oligonucleotide) and ribozyme targets to the same region were also synthesized for comparison analysis. Alterations in MDR1 mRNA and Pgp were determined by RT-PCR, Northern blot, flow cytometry and Rh123 retention tests. Chemosensitivity of the treated cells was determined by MTT assay. The results showed that DRz could significantly suppress expression of MDR1 mRNA and inhibit synthesis of Pgp. The efflux activity of Pgp was inhibited accordingly. Chemosensitivity assay showed that a 21-fold reduction in drug resistance for Adriamycin and a 45-fold reduction in drug resistance for Vinblastine were found in the treated cells 36h after transfection. These data suggest that DRz targeted to the translation initiation codon AUG can reverse MDR phenotype in cancer cells and restore their chemosensitivity. Moreover, the reversal efficiency of DRz is better than that of ribozyme and ASODN targets to the same region of MDR1 mRNA. Copyright © 2013 Elsevier Inc. All rights reserved.

Defective canalicular transport and toxicity of dietary ursodeoxycholic acid in the abcb11-/- mouse: transport and gene expression studies.

PubMed

Wang, Renxue; Liu, Lin; Sheps, Jonathan A; Forrest, Dana; Hofmann, Alan F; Hagey, Lee R; Ling, Victor

2013-08-15

The bile salt export pump (BSEP), encoded by the abcb11 gene, is the major canalicular transporter of bile acids from the hepatocyte. BSEP malfunction in humans causes bile acid retention and progressive liver injury, ultimately leading to end-stage liver failure. The natural, hydrophilic, bile acid ursodeoxycholic acid (UDCA) is efficacious in the treatment of cholestatic conditions, such as primary biliary cirrhosis and cholestasis of pregnancy. The beneficial effects of UDCA include promoting bile flow, reducing hepatic inflammation, preventing apoptosis, and maintaining mitochondrial integrity in hepatocytes. However, the role of BSEP in mediating UDCA efficacy is not known. Here, we used abcb11 knockout mice (abcb11-/-) to test the effects of acute and chronic UDCA administration on biliary secretion, bile acid composition, liver histology, and liver gene expression. Acutely infused UDCA, or its taurine conjugate (TUDC), was taken up by the liver but retained, with negligible biliary output, in abcb11-/- mice. Feeding UDCA to abcb11-/- mice led to weight loss, retention of bile acids, elevated liver enzymes, and histological damage to the liver. Semiquantitative RT-PCR showed that genes encoding Mdr1a and Mdr1b (canalicular) as well as Mrp4 (basolateral) transporters were upregulated in abcb11-/- mice. We concluded that infusion of UDCA and TUDC failed to induce bile flow in abcb11-/- mice. UDCA fed to abcb11-/- mice caused liver damage and the appearance of biliary tetra- and penta-hydroxy bile acids. Supplementation with UDCA in the absence of Bsep caused adverse effects in abcb11-/- mice.

RNA Nuclear Export: From Neurological Disorders to Cancer.

PubMed

Hautbergue, Guillaume M

2017-01-01

The presence of a nuclear envelope, also known as nuclear membrane, defines the structural framework of all eukaryotic cells by separating the nucleus, which contains the genetic material, from the cytoplasm where the synthesis of proteins takes place. Translation of proteins in Eukaryotes is thus dependent on the active transport of DNA-encoded RNA molecules through pores embedded within the nuclear membrane. Several mechanisms are involved in this process generally referred to as RNA nuclear export or nucleocytoplasmic transport of RNA. The regulated expression of genes requires the nuclear export of protein-coding messenger RNA molecules (mRNAs) as well as non-coding RNAs (ncRNAs) together with proteins and pre-assembled ribosomal subunits. The nuclear export of mRNAs is intrinsically linked to the co-transcriptional processing of nascent transcripts synthesized by the RNA polymerase II. This functional coupling is essential for the survival of cells allowing for timely nuclear export of fully processed transcripts, which could otherwise cause the translation of abnormal proteins such as the polymeric repeat proteins produced in some neurodegenerative diseases. Alterations of the mRNA nuclear export pathways can also lead to genome instability and to various forms of cancer. This chapter will describe the molecular mechanisms driving the nuclear export of RNAs with a particular emphasis on mRNAs. It will also review their known alterations in neurological disorders and cancer, and the recent opportunities they offer for the potential development of novel therapeutic strategies.

A Truncated AdeS Kinase Protein Generated by ISAba1 Insertion Correlates with Tigecycline Resistance in Acinetobacter baumannii

PubMed Central

Sun, Jun-Ren; Perng, Cherng-Lih; Chan, Ming-Chin; Morita, Yuji; Lin, Jung-Chung; Su, Chih-Mao; Wang, Wei-Yao; Chang, Tein-Yao; Chiueh, Tzong-Shi

2012-01-01

Over-expression of AdeABC efflux pump stimulated continuously by the mutated AdeRS two component system has been found to result in antimicrobial resistance, even tigecycline (TGC) resistance, in multidrug-resistant Acinetobacter baumannii (MRAB). Although the insertion sequence, ISAba1, contributes to one of the AdeRS mutations, the detail mechanism remains unclear. In the present study we collected 130 TGC-resistant isolates from 317 carbapenem resistant MRAB (MRAB-C) isolates, and 38 of them were characterized with ISAba1 insertion in the adeS gene. The relationship between the expression of AdeABC efflux pump and TGC resistant was verified indirectly by successfully reducing TGC resistance with NMP, an efflux pump inhibitor. Further analysis showed that the remaining gene following the ISAba1 insertion was still transcribed to generate a truncated AdeS protein by the Pout promoter on ISAba1 instead of frame shift or pre-termination. Through introducing a series of recombinant adeRS constructs into a adeRS knockout strain, we demonstrated the truncated AdeS protein was constitutively produced and stimulating the expression of AdeABC efflux pump via interaction with AdeR. Our findings suggest a mechanism of antimicrobial resistance induced by an aberrant cytoplasmic sensor derived from an insertion element. PMID:23166700

Alternative splicing of Staufen2 creates the nuclear export signal for CRM1 (Exportin 1).

PubMed

Miki, Takashi; Yoneda, Yoshihiro

2004-11-12

Mammalian Staufen2 (Stau2), a brain-specific double-stranded RNA-binding protein, is involved in the localization of mRNA in neurons. To gain insights into the function of Stau2, the subcellular localization of Stau2 isoforms fused to the green fluorescence protein was examined. Fluorescence microscopic analysis showed that Stau2 functions as a nucleocytoplasmic shuttle protein. The nuclear export of the 62-kDa isoform of Stau2 (Stau2(62)) is mediated by the double-stranded RNA-binding domain 3 (RBD3) because a mutation to RBD3 led to nuclear accumulation. On the other hand, the shorter isoform of Stau2, Stau2(59), is exported from the nucleus by two distinct pathways, one of which is RBD3-mediated and the other of which is CRM1 (exportin 1)-dependent. The nuclear export signal recognized by CRM1 was found to be located in the N-terminal region of Stau2(59). These results suggest that Stau2 may carry a variety of RNAs out of the nucleus, using the two export pathways. The present study addresses the issue of why plural Stau2 isoforms are expressed in neurons.

Modified host cells with efflux pumps

DOEpatents

Dunlop, Mary J.; Keasling, Jay D.; Mukhopadhyay, Aindrila

2016-08-30

The present invention provides for a modified host cell comprising a heterologous expression of an efflux pump capable of transporting an organic molecule out of the host cell wherein the organic molecule at a sufficiently high concentration reduces the growth rate of or is lethal to the host cell.

AKT3 controls mitochondrial biogenesis and autophagy via regulation of the major nuclear export protein CRM-1.

PubMed

Corum, Daniel G; Tsichlis, Philip N; Muise-Helmericks, Robin C

2014-01-01

Our previous work has shown that Akt3 is required for mitochondrial biogenesis in primary human endothelial cells (ECs) and in Akt3-null mice; Akt3 affects subcellular localization of peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1α), the master regulator of mitochondrial biogenesis. The purpose of this study is to determine the mechanism by which Akt3 controls the subcellular distribution of PGC-1α and to explore the effect on mitochondrial biogenesis and turnover during angiogenesis. Here we use standard biochemical analyses and Akt3-knockdown strategies to show that Akt3 controls the stabilization of chromosome maintenance region-1 (CRM-1), the major nuclear export receptor. Site-directed mutagenesis and association analyses show that PGC-1α nuclear export is CRM-1 dependent. Akt3 knockdown and CRM-1 overexpression cause 3-fold reductions in PGC-1α target gene expression, compared to control levels. Akt3 inhibition causes autophagy, as measured by autophagosome formation, in a CRM-1-dependent, Akt1/mTOR-independent pathway. In vivo, Akt3-null and heterozygous mice show dose-dependent decreases in angiogenesis compared to wild-type littermates (~5- and 2.5-fold decreases, respectively), as assessed by Matrigel plug assays. This correlates with an ~1.5-fold decrease in mitochondrial Cox IV expression. Our studies suggest that Akt3 is a regulator of mitochondrial dynamics in the vasculature via regulation of CRM-1-dependent nuclear export.

Broad host range vectors for expression of proteins with (Twin-) Strep-tag, His-tag and engineered, export optimized yellow fluorescent protein

PubMed Central

2013-01-01

Background In current protein research, a limitation still is the production of active recombinant proteins or native protein associations to assess their function. Especially the localization and analysis of protein-complexes or the identification of modifications and small molecule interaction partners by co-purification experiments requires a controllable expression of affinity- and/or fluorescence tagged variants of a protein of interest in its native cellular background. Advantages of periplasmic and/or homologous expressions can frequently not be realized due to a lack of suitable tools. Instead, experiments are often limited to the heterologous production in one of the few well established expression strains. Results Here, we introduce a series of new RK2 based broad host range expression plasmids for inducible production of affinity- and fluorescence tagged proteins in the cytoplasm and periplasm of a wide range of Gram negative hosts which are designed to match the recently suggested modular Standard European Vector Architecture and database. The vectors are equipped with a yellow fluorescent protein variant which is engineered to fold and brightly fluoresce in the bacterial periplasm following Sec-mediated export, as shown from fractionation and imaging studies. Expression of Strep-tag®II and Twin-Strep-tag® fusion proteins in Pseudomonas putida KT2440 is demonstrated for various ORFs. Conclusion The broad host range constructs we have produced enable good and controlled expression of affinity tagged protein variants for single-step purification and qualify for complex co-purification experiments. Periplasmic export variants enable production of affinity tagged proteins and generation of fusion proteins with a novel engineered Aequorea-based yellow fluorescent reporter protein variant with activity in the periplasm of the tested Gram-negative model bacteria Pseudomonas putida KT2440 and Escherichia coli K12 for production, localization or co-localization studies. In addition, the new tools facilitate metabolic engineering and yield assessment for cytoplasmic or periplasmic protein production in a number of different expression hosts when yields in one initially selected are insufficient. PMID:23687945

Two Outer Membrane Proteins Contribute to Caulobacter crescentus Cellular Fitness by Preventing Intracellular S-Layer Protein Accumulation

DOE PAGES

Overton, K. Wesley; Park, Dan M.; Yung, Mimi C.; ...

2016-09-23

Surface layers, or S-layers, are two-dimensional protein arrays that form the outermost layer of many bacteria and archaea. They serve several functions, including physical protection of the cell from environmental threats. The high abundance of S-layer proteins necessitates a highly efficient export mechanism to transport the S-layer protein from the cytoplasm to the cell exterior.Caulobacter crescentusis unique in that it has two homologous, seemingly redundant outer membrane proteins, RsaF aand RsaF b, which together with other components form a type I protein translocation pathway for S-layer export. These proteins have homology toEscherichia coliTolC, the outer membrane channel of multidrug effluxmore » pumps. Here we provide evidence that, unlike TolC, RsaF aand RsaF bare not involved in either the maintenance of membrane stability or the active export of antimicrobial compounds. Rather, RsaF aand RsaF bare required to prevent intracellular accumulation and aggregation of the S-layer protein RsaA; deletion of RsaF aand RsaF bled to a general growth defect and lowered cellular fitness. Using Western blotting, transmission electron microscopy, and transcriptome sequencing (RNA-seq), we show that loss of both RsaF aand RsaF bled to accumulation of insoluble RsaA in the cytoplasm, which in turn caused upregulation of a number of genes involved in protein misfolding and degradation pathways. These findings provide new insight into the requirement for RsaF aand RsaF bin cellular fitness and tolerance to antimicrobial agents and further our understanding of the S-layer export mechanism on both the transcriptional and translational levels inC. crescentus. IMPORTANCEDecreased growth rate and reduced cell fitness are common side effects of protein production in overexpression systems. Inclusion bodies typically form inside the cell, largely due to a lack of sufficient export machinery to transport the overexpressed proteins to the extracellular environment. This phenomenon can conceivably also occur in natural systems. As one example of a system evolved to prevent intracellular protein accumulation, our study demonstrates thatCaulobacter crescentushas two homologous outer membrane transporter proteins that are involved in S-layer export. This is an interesting case study that demonstrates how bacteria can evolve redundancy to ensure adequate protein export functionality and maintain high cellular fitness. Moreover, we provide evidence that these two outer membrane proteins, although being the closestC. crescentushomologs to TolC inE. coli, do not process TolC functionality inC. crescentus.« less

Two Outer Membrane Proteins Contribute to Caulobacter crescentus Cellular Fitness by Preventing Intracellular S-Layer Protein Accumulation

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

Overton, K. Wesley; Park, Dan M.; Yung, Mimi C.

ABSTRACT Surface layers, or S-layers, are two-dimensional protein arrays that form the outermost layer of many bacteria and archaea. They serve several functions, including physical protection of the cell from environmental threats. The high abundance of S-layer proteins necessitates a highly efficient export mechanism to transport the S-layer protein from the cytoplasm to the cell exterior.Caulobacter crescentusis unique in that it has two homologous, seemingly redundant outer membrane proteins, RsaF aand RsaF b, which together with other components form a type I protein translocation pathway for S-layer export. These proteins have homology toEscherichia coliTolC, the outer membrane channel of multidrugmore » efflux pumps. Here we provide evidence that, unlike TolC, RsaF aand RsaF bare not involved in either the maintenance of membrane stability or the active export of antimicrobial compounds. Rather, RsaF aand RsaF bare required to prevent intracellular accumulation and aggregation of the S-layer protein RsaA; deletion of RsaF aand RsaF bled to a general growth defect and lowered cellular fitness. Using Western blotting, transmission electron microscopy, and transcriptome sequencing (RNA-seq), we show that loss of both RsaF aand RsaF bled to accumulation of insoluble RsaA in the cytoplasm, which in turn caused upregulation of a number of genes involved in protein misfolding and degradation pathways. These findings provide new insight into the requirement for RsaF aand RsaF bin cellular fitness and tolerance to antimicrobial agents and further our understanding of the S-layer export mechanism on both the transcriptional and translational levels inC. crescentus. IMPORTANCEDecreased growth rate and reduced cell fitness are common side effects of protein production in overexpression systems. Inclusion bodies typically form inside the cell, largely due to a lack of sufficient export machinery to transport the overexpressed proteins to the extracellular environment. This phenomenon can conceivably also occur in natural systems. As one example of a system evolved to prevent intracellular protein accumulation, our study demonstrates thatCaulobacter crescentushas two homologous outer membrane transporter proteins that are involved in S-layer export. This is an interesting case study that demonstrates how bacteria can evolve redundancy to ensure adequate protein export functionality and maintain high cellular fitness. Moreover, we provide evidence that these two outer membrane proteins, although being the closestC. crescentushomologs to TolC inE. coli, do not process TolC functionality inC. crescentus.« less

Two Outer Membrane Proteins Contribute to Caulobacter crescentus Cellular Fitness by Preventing Intracellular S-Layer Protein Accumulation

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

Overton, K. Wesley; Park, Dan M.; Yung, Mimi C.

Surface layers, or S-layers, are two-dimensional protein arrays that form the outermost layer of many bacteria and archaea. They serve several functions, including physical protection of the cell from environmental threats. The high abundance of S-layer proteins necessitates a highly efficient export mechanism to transport the S-layer protein from the cytoplasm to the cell exterior.Caulobacter crescentusis unique in that it has two homologous, seemingly redundant outer membrane proteins, RsaF aand RsaF b, which together with other components form a type I protein translocation pathway for S-layer export. These proteins have homology toEscherichia coliTolC, the outer membrane channel of multidrug effluxmore » pumps. Here we provide evidence that, unlike TolC, RsaF aand RsaF bare not involved in either the maintenance of membrane stability or the active export of antimicrobial compounds. Rather, RsaF aand RsaF bare required to prevent intracellular accumulation and aggregation of the S-layer protein RsaA; deletion of RsaF aand RsaF bled to a general growth defect and lowered cellular fitness. Using Western blotting, transmission electron microscopy, and transcriptome sequencing (RNA-seq), we show that loss of both RsaF aand RsaF bled to accumulation of insoluble RsaA in the cytoplasm, which in turn caused upregulation of a number of genes involved in protein misfolding and degradation pathways. These findings provide new insight into the requirement for RsaF aand RsaF bin cellular fitness and tolerance to antimicrobial agents and further our understanding of the S-layer export mechanism on both the transcriptional and translational levels inC. crescentus. IMPORTANCEDecreased growth rate and reduced cell fitness are common side effects of protein production in overexpression systems. Inclusion bodies typically form inside the cell, largely due to a lack of sufficient export machinery to transport the overexpressed proteins to the extracellular environment. This phenomenon can conceivably also occur in natural systems. As one example of a system evolved to prevent intracellular protein accumulation, our study demonstrates thatCaulobacter crescentushas two homologous outer membrane transporter proteins that are involved in S-layer export. This is an interesting case study that demonstrates how bacteria can evolve redundancy to ensure adequate protein export functionality and maintain high cellular fitness. Moreover, we provide evidence that these two outer membrane proteins, although being the closestC. crescentushomologs to TolC inE. coli, do not process TolC functionality inC. crescentus.« less

A novel ABCB11 mutation in an Iranian girl with progressive familial intrahepatic cholestasis

PubMed Central

Saber, Sassan; Vazifehmand, Reza; Bagherizadeh, Iman; Kasiri, Mahbubeh

2013-01-01

Progressive familial intrahepatic cholestasis is an autosomal recessive liver disorder caused by (biallelic) mutations in the ATP8B1 of ABCB11 gene. A nine-year-old girl with cholestasis was referred for genetic counseling. She had a family history of cholestasis in two previous expired siblings. Genetic analysis of the ABCB11 gene led to the identification of a novel homozygous mutation in exon 25. The mutation 3593- A > G lead to a missense mutation at the amino acid level (His1198Arg). This mutation caused PFIC2 due to abnormal function in the bile salt export pump protein (BSEP). PMID:24339557

Direct observation of nucleocytoplasmic transport by microinjection of GFP-tagged proteins in living cells.

PubMed

Rosorius, O; Heger, P; Stelz, G; Hirschmann, N; Hauber, J; Stauber, R H

1999-08-01

We established a straightforward experimental system to investigate directly the requirements for nucleocytoplasmic transport in live cells. For this purpose, substrates were created containing nuclear localization signals (NLS) or nuclear export signals (NES) linked to a chimeric protein composed of the glutathione S-transferase (GST) fused to the green fluorescent protein (GFP). The combination of GST/GFP-tagging allowed us to control protein expression in bacteria and to monitor protein purification during chromatography. Following microinjection into somatic cells, nuclear export/import of the highly fluorescent substrates could be observed directly by fluorescence microscopy. This system sets the stage to quantitate, in real time, the kinetics of nuclear import/export in living cells and to evaluate qualitative differences in various NLS/NES signals and pathways.

Identification of Bicarbonate as a Trigger and Genes Involved with Extracellular DNA Export in Mycobacterial Biofilms

PubMed Central

Rose, Sasha J.

2016-01-01

ABSTRACT Extracellular DNA (eDNA) is an integral biofilm matrix component of numerous pathogens, including nontuberculous mycobacteria (NTM). Cell lysis is the source of eDNA in certain bacteria, but the source of eDNA remains unidentified for NTM, as well as for other eDNA-containing bacterial species. In this study, conditions affecting eDNA export were examined, and genes involved with the eDNA export mechanism were identified. After a method for monitoring eDNA in real time in undisturbed biofilms was established, different conditions affecting eDNA were investigated. Bicarbonate positively influenced eDNA export in a pH-independent manner in Mycobacterium avium, M. abscessus, and M. chelonae. The surface-exposed proteome of M. avium in eDNA-containing biofilms revealed abundant carbonic anhydrases. Chemical inhibition of carbonic anhydrases with ethoxzolamide significantly reduced eDNA export. An unbiased transposon mutant library screen for eDNA export in M. avium identified many severely eDNA-attenuated mutants, including one not expressing a unique FtsK/SpoIIIE-like DNA-transporting pore, two with inactivation of carbonic anhydrases, and nine with inactivation of genes belonging to a unique genomic region, as well as numerous mutants involved in metabolism and energy production. Complementation of nine mutants that included the FtsK/SpoIIIE and carbonic anhydrase significantly restored eDNA export. Interestingly, several attenuated eDNA mutants have mutations in genes encoding proteins that were found with the surface proteomics, and many more mutations are localized in operons potentially encoding surface proteins. Collectively, our data strengthen the evidence of eDNA export being an active mechanism that is activated by the bacterium responding to bicarbonate. PMID:27923918

Enhanced immunogenicity of HPV 16 E7 fusion proteins in DNA vaccination.

PubMed

Michel, Nico; Osen, Wolfram; Gissmann, Lutz; Schumacher, Ton N M; Zentgraf, Hanswalter; Müller, Martin

2002-03-01

DNA vaccination is a promising approach for inducing both humoral and cellular immune responses. For immunotherapy of HPV-16-associated diseases the E7 protein is considered a prime candidate, as it is expressed in all HPV-16-positive tumors. Unfortunately, the E7 protein is a very poor inducer of a cytotoxic T-cell response, when being used as antigen in DNA vaccination. Here we demonstrate that after fusion to protein export/import signals such as the herpes simplex virus ferry protein VP22, E7 can translocate in vitro from VP22-E7-expressing cells to neighboring cells that do not carry the VP22-E7 gene. In vivo, the VP22-E7 fusion shows significantly increased efficiency in inducing a cytotoxic T-cell response. Our data suggest that the export function of VP22 plays a major role in this phenomenon, since VP22 can be replaced by classical protein export signals, without impairing the induction of the E7-specific cellular immune response. However, all E7 fusion constructs showed significantly elevated protein steady-state levels, which might also account for the observed boost in immunogenicity. (C)2002 Elsevier Science (USA).

Effect of dexamethasone on skeletal muscle Na+,K+ pump subunit specific expression and K+ homeostasis during exercise in humans

PubMed Central

Nordsborg, Nikolai; Ovesen, Jakob; Thomassen, Martin; Zangenberg, Mathias; Jøns, Christian; Iaia, F Marcello; Nielsen, Jens Jung; Bangsbo, Jens

2008-01-01

The effect of dexamethasone on Na+,K+ pump subunit expression and muscle exchange of K+ during exercise in humans was investigated. Nine healthy male subjects completed a randomized double blind placebo controlled protocol, with ingestion of dexamethasone (Dex: 2 × 2 mg per day) or placebo (Pla) for 5 days. Na+,K+ pump catalytic α1 and α2 subunit expression was ∼17% higher (P < 0.05) and the structural β1 and β2 subunit expression was ∼6–8% higher (P < 0.05) after Dex compared with Pla. During one-legged knee-extension for 10 min at low intensity (LI; 18.6 ± 1.0 W), two moderate intensity (51.7 ± 2.4 W) exercise bouts (MI1: 5 min; 2 min recovery; MI2: exhaustive) and two high-intensity (71.7 ± 2.5 W) exercise bouts (HI1: 1 min 40 s; 2 min recovery; HI2: exhaustive), femoral venous K+ was lower (P < 0.05) in Dex compared with Pla. Thigh K+ release was lower (P < 0.05) in Dex compared with Pla in LI and MI, but not in HI. Time to exhaustion in MI2 tended to improve (393 ± 50 s versus 294 ± 41 s; P = 0.07) in Dex compared with Pla, whereas no difference was detected in HI2 (106 ± 10 s versus 108 ± 9 s). The results indicate that an increased Na+,K+ pump expression per se is of importance for thigh K+ reuptake at the onset of low and moderate intensity exercise, but less important during high intensity exercise. PMID:18174214

Enhancement of survival and electricity production in an engineered bacterium by light-driven proton pumping.

PubMed

Johnson, Ethan T; Baron, Daniel B; Naranjo, Belén; Bond, Daniel R; Schmidt-Dannert, Claudia; Gralnick, Jeffrey A

2010-07-01

Microorganisms can use complex photosystems or light-dependent proton pumps to generate membrane potential and/or reduce electron carriers to support growth. The discovery that proteorhodopsin is a light-dependent proton pump that can be expressed readily in recombinant bacteria enables development of new strategies to probe microbial physiology and to engineer microbes with new light-driven properties. Here, we describe functional expression of proteorhodopsin and light-induced changes in membrane potential in the bacterium Shewanella oneidensis strain MR-1. We report that there were significant increases in electrical current generation during illumination of electrochemical chambers containing S. oneidensis expressing proteorhodopsin. We present evidence that an engineered strain is able to consume lactate at an increased rate when it is illuminated, which is consistent with the hypothesis that proteorhodopsin activity enhances lactate uptake by increasing the proton motive force. Our results demonstrate that there is coupling of a light-driven process to electricity generation in a nonphotosynthetic engineered bacterium. Expression of proteorhodopsin also preserved the viability of the bacterium under nutrient-limited conditions, providing evidence that fulfillment of basic energy needs of organisms may explain the widespread distribution of proteorhodopsin in marine environments.

Contribution of urban runoff in Taipei metropolitan area to dissolved inorganic nitrogen export in the Danshui River, Taiwan.

PubMed

Kuo, Nae-Wen; Jien, Shih-Hao; Hong, Nien-Ming; Chen, Yao-Te; Lee, Tsung-Yu

2017-01-01

A previous study has demonstrated that Danshui River has almost the highest dissolved inorganic nitrogen (DIN) yield in the world and exports most of the DIN in the form of ammonium unlike the world's large rivers. However, the DIN sources are poorly constrained. In this study, the contributions of major sources in the Taipei metropolitan area to the DIN export in the Danshui River were investigated. It is observed that ammonium is the major DIN species in the downstream reaches, resulting from the ammonium-dominated inputs of the effluents of wastewater treatment plants (WWTP) and rain water pumping stations (RWPS). DIN concentrations in the downstream (urban) reaches are substantially elevated. The upstream tributaries annually discharge ∼2709 t DIN to the downstream reaches. However, the DIN discharge off the downstream reaches rises to ∼17,918 t, resulting from the contribution of RWPS-collected water, i.e., ∼14,632 t, and the effluents of two WWTP, i.e., ∼577 t. RWPS-collected water inherently contains the contribution of atmospheric deposition, ∼2937 t DIN. This finding implies that ∼11,695 t (∼66 % of the downstream output) DIN flux off the Danshui River is from urban runoff and can be attributed to human activities in the Taipei metropolitan area. To improve the water quality in the Danshui River, water quality controls in urban runoff are important.

Electrical controllable spin pump based on a zigzag silicene nanoribbon junction.

PubMed

Zhang, Lin; Tong, Peiqing

2017-12-13

We propose a possible electrical controllable spin pump based on a zigzag silicene nanoribbon ferromagnetic junction by applying two time-dependent perpendicular electric fields. By using the Keldysh Green's function method, we derive the analytic expression of the spin-resolved current at the adiabatic approximation and demonstrate that two asymmetric spin up and spin down currents can be pumped out in the device without an external bias. The pumped currents mainly come from the interplay between the photon-assisted spin pump effect and the electrically-modulated energy band structure of the tunneling junction. The spin valve phenomena are not only related to the energy gap opened by two perpendicular staggered potentials, but also dependent on the system parameters such as the pumping frequency, the pumping phase difference, the spin-orbit coupling and the Fermi level, which can be tuned by the electrical methods. The proposed device can also be used to produce a pure spin current and a 100% polarized spin current through the photon-assisted pumping process. Our investigations may provide an electrical manipulation of spin-polarized electrons in graphene-like pumping devices.

A double-edged sword: lactation consultants' perceptions of the impact of breast pumps on the practice of breastfeeding.

PubMed

Buckley, Kathleen M

2009-01-01

This qualitative descriptive study examined the beliefs and experiences of 12 lactation consultants regarding the impact of breast pumps on breastfeeding practices. Interview topics on breast pumps included types and patterns of use, mothers' experiences, and advantages and risks. The lactation consultants reported an increase in the use of breast pumps due to improved marketing, a change in society's view of pumps as a necessity rather than a luxury, and the impact of birthing technology. Reasons given for this increased use were mothers' need to have greater control over the breastfeeding process and to quantify the amount of breastmilk. Concerns were expressed regarding an overdependency on breastfeeding technology by some lactation consultants and mothers.

A Double-Edged Sword: Lactation Consultants' Perceptions of the Impact of Breast Pumps on the Practice of Breastfeeding

PubMed Central

Buckley, Kathleen M

2009-01-01

This qualitative descriptive study examined the beliefs and experiences of 12 lactation consultants regarding the impact of breast pumps on breastfeeding practices. Interview topics on breast pumps included types and patterns of use, mothers' experiences, and advantages and risks. The lactation consultants reported an increase in the use of breast pumps due to improved marketing, a change in society's view of pumps as a necessity rather than a luxury, and the impact of birthing technology. Reasons given for this increased use were mothers' need to have greater control over the breastfeeding process and to quantify the amount of breastmilk. Concerns were expressed regarding an overdependency on breastfeeding technology by some lactation consultants and mothers. PMID:20190850

Analytical solution of the transient temperature profile in gain medium of passively Q-switched microchip laser.

PubMed

Han, Xiahui; Li, Jianlang

2014-11-01

The transient temperature evolution in the gain medium of a continuous wave (CW) end-pumped passively Q-switched microchip (PQSM) laser is analyzed. By approximating the time-dependent population inversion density as a sawtooth function of time and treating the time-dependent pump absorption of a CW end-pumped PQSM laser as the superposition of an infinite series of short pumping pulses, the analytical expressions of transient temperature evolution and distribution in the gain medium for four- and three-level laser systems, respectively, are given. These analytical solutions are applied to evaluate the transient temperature evolution and distribution in the gain medium of CW end-pumped PQSM Nd:YAG and Yb:YAG lasers.

The Role of the Transcription Factors MtrR and MtrA in the Fitness of the Pathogen Neisseria gonorrhoeae

DTIC Science & Technology

2007-10-19

MacA -MacB efflux pump is expressed at very low levels in N. gonorrhoeae; however, experimental alteration of the promoter sequence showed that the...specificity (132). The gonococcus expresses four efflux pump systems, namely MtrC-Mtr-D-MtrE (64), FarA-FarB-MtrE (97), NorM (158), and MacA -MacB (161...2005. Characterization of the MacA -MacB efflux system in Neisseria gonorrhoeae. The Journal of antimicrobial chemotherapy 56:856-860. 162. Rouquette, C

Full counting statistics of a charge pump in the Coulomb blockade regime

NASA Astrophysics Data System (ADS)

Andreev, A. V.; Mishchenko, E. G.

2001-12-01

We study full charge counting statistics (FCCS) of a charge pump based on a nearly open single electron transistor. The problem is mapped onto an exactly soluble problem of a nonequilibrium g=1/2 Luttinger liquid with an impurity. We obtain an analytic expression for the generating function of the transmitted charge for an arbitrary pumping strength. Although this model contains fractionally charged excitations only integer transmitted charges can be observed. In the weak pumping limit FCCS correspond to a Poissonian transmission of particles with charge e*=e/2 from which all events with odd numbers of transferred particles are excluded.

Expression and purification of a new recombinant camel hepcidin able to promote the degradation of the iron exporter ferroportin1.

PubMed

Boumaiza, Mohamed; Jaouen, Maryse; Deschemin, Jean-Christophe; Ezzine, Aymen; Ben Khalaf, Noureddine; Vaulont, Sophie; Marzouki, Mohamed Nèjib; Sari, Marie Agnès

2015-11-01

Hepcidin, a 25-amino-acid and highly disulfide bonded antimicrobial peptide, is the central regulator of iron homeostasis. This hormone is expressed in response to iron and inflammation and interacts with ferroportin1 (FPN1), the only known iron exporter in vertebrates, inducing its internalization and degradation. Thus, the export of iron from cells to plasma will be significantly diminished. Thereby, hepcidin has become the target of intense research studies due to its profound biomedical significance. This study describes the functional expression of recombinant camel hepcidin in Escherichia coli. Biologically active recombinant camel hepcidin was obtained thanks to the production of a hepcidin-thioredoxin fusion protein (TRX-HepcD) and a purified camel hepcidin, with an extra methionine at the N-terminus, was obtained after enterokinase cleavage of the fusion protein. Presence of the four disulfide bridges was verified using MALDI-ToF spectrometry. The recombinant camel hepcidin was compared to related synthetic bioactive peptides, including human hepcidin, and was found equally able to promote ferroportin degradation of mouse macrophages. Furthermore, camel hepcidins exhibits a high capacity to inhibit the growth of Leishmania major promastigotes. These results proved that production of functional camel hepcidin can be achieved in E. coli, this is a major interest for the production of cysteine rich peptides or proteins that can be purified under their functional form without the need of a refolding process. Copyright © 2015 Elsevier Inc. All rights reserved.

Structure of a prokaryotic virtual proton pump at 3.2 Å resolution

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

Fang, Yiling; Jayaram, Hariharan; Shane, Tania

2009-09-15

To reach the mammalian gut, enteric bacteria must pass through the stomach. Many such organisms survive exposure to the harsh gastric environment (pH 1.5-4) by mounting extreme acid-resistance responses, one of which, the arginine-dependent system of Escherichia coli, has been studied at levels of cellular physiology, molecular genetics and protein biochemistry. This multiprotein system keeps the cytoplasm above pH 5 during acid challenge by continually pumping protons out of the cell using the free energy of arginine decarboxylation. At the heart of the process is a 'virtual proton pump' in the inner membrane, called AdiC, that imports L-arginine from themore » gastric juice and exports its decarboxylation product agmatine. AdiC belongs to the APC superfamily of membrane proteins, which transports amino acids, polyamines and organic cations in a multitude of biological roles, including delivery of arginine for nitric oxide synthesis, facilitation of insulin release from pancreatic {beta}-cells, and, when inappropriately overexpressed, provisioning of certain fast-growing neoplastic cells with amino acids. High-resolution structures and detailed transport mechanisms of APC transporters are currently unknown. Here we describe a crystal structure of AdiC at 3.2 {angstrom} resolution. The protein is captured in an outward-open, substrate-free conformation with transmembrane architecture remarkably similar to that seen in four other families of apparently unrelated transport proteins.« less

Transfection of influenza A virus nuclear export protein induces the expression of tumor necrosis factor alpha.

PubMed

Lara-Sampablo, Alejandra; Flores-Alonso, Juan Carlos; De Jesús-Ortega, Nereyda; Santos-López, Gerardo; Vallejo-Ruiz, Verónica; Rosas-Murrieta, Nora; Reyes-Carmona, Sandra; Herrera-Camacho, Irma; Reyes-Leyva, Julio

2014-06-24

Influenza A virus genomic segments eight codes for non-structural 1 (NS1) protein that is involved in evasion of innate antiviral response, and nuclear export protein (NEP) that participates in the export of viral ribonucleoprotein (RNP) complexes, transcription and replication. Tumor necrosis factor alpha (TNF-α) is highly expressed during influenza virus infections and is considered an anti-infective cytokine. NS1 and NEP proteins were overexpressed and their role on TNF-α expression was evaluated. Both TNF-α mRNA and protein increased in cells transfected with NEP but not with NS1. We further investigate if NS1 or NEP regulates the activity of TNF-α promoter. In the presence of NEP the activity of TNF-α promoter increased significantly compared with the control (83.5±2.9 vs. 30.9±2.8, respectively; p=0.001). This effect decreased 15-fold when the TNF-α promoter distal region was deleted, suggesting the involvement of mitogen-activated protein kinases (MAPK) and NF-kB response elements. This was corroborated by testing the effect produced on TNF-α promoter by the treatment with Raf/MEK/ERK (U0126), NF-kB (Bay-11-7082) and PI3K (Ly294-002) cell signaling inhibitors. Treatment with U0126 and Bay-117082 reduced the activity of TNF-α promoter mediated by NEP (41.5±3.2, 70% inhibition; and 80.6±7.4, 35% inhibition, respectively) compared to mock-treated control. The results suggest a new role for NEP protein that participates in the transcriptional regulation of human TNF-α expression. Copyright © 2014 Elsevier B.V. All rights reserved.

An NXF1 mRNA with a retained intron is expressed in hippocampal and neocortical neurons and is translated into a protein that functions as an Nxf1 cofactor.

PubMed

Li, Ying; Bor, Yeou-Cherng; Fitzgerald, Mark P; Lee, Kevin S; Rekosh, David; Hammarskjold, Marie-Louise

2016-12-01

The Nxf1 protein is a major nuclear export receptor for the transport of mRNA, and it also is essential for export of retroviral mRNAs with retained introns. In the latter case, it binds to RNA elements known as constitutive transport elements (CTEs) and functions in conjunction with a cofactor known as Nxt1. The NXF1 gene also regulates expression of its own intron-containing RNA through the use of a functional CTE within intron 10. mRNA containing this intron is exported to the cytoplasm, where it can be translated into the 356-amino acid short Nxf1(sNxf1) protein, despite the fact that it is a prime candidate for nonsense-mediated decay (NMD). Here we demonstrate that sNxf1 is highly expressed in nuclei and dendrites of hippocampal and neocortical neurons in rodent brain. Additionally, we show that sNxf1 localizes in RNA granules in neurites of differentiated N2a mouse neuroblastoma cells, where it shows partial colocalization with Staufen2 isoform SS, a protein known to play a role in dendritic mRNA trafficking. We also show that sNxf1 forms heterodimers in conjunction with the full-length Nxf1 and that sNxf1 can replace Nxt1 to enhance the expression of CTE-containing mRNA and promote its association with polyribosomes. © 2016 Li et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Expression of a novel gene, gluP, is essential for normal Bacillus subtilis cell division and contributes to glucose export

PubMed Central

Mesak, Lili R; Mesak, Felix M; Dahl, Michael K

2004-01-01

Background The Bacillus subtilis glucokinase operon was predicted to be comprised of the genes, yqgP (now named gluP), yqgQ, and glcK. We have previously established a role for glcK in glucose metabolism. In the absence of enzymes that phosphorylate glucose, such as GlcK and/or enzyme IIGlc, accumulated cytoplasmic glucose can be transported out of the cell. Genes within the glucokinase operon were not previously known to play a role in glucose transport. Here we describe the expression of gluP and its function in glucose transport. Results We found that transcription of the glucokinase operon was regulated, putatively, by two promoters: σA and σH. Putative σA and σH-recognition sites were located upstream of and within gluP, respectively. Transcriptional glucokinase operon – lacZ fusions and Northern blotting were used to analyze the expression of gluP. GluP was predicted to be an integral membrane protein. Moreover, the prediction of GluP structure revealed interesting signatures: a rhomboid domain and two tetracopeptide repeat (TPR) motifs. Microscopic analysis showed that GluP minus cells were unable to divide completely, resulting in a filamentous phenotype. The cells were grown in either rich or minimal medium. We found GluP may be involved in glucose transport. [14C]-glucose uptake by the GluP minus strain was slightly less than in the wild type. On the other hand, trehalose-derived glucose in the growth medium of the GluP minus strain was detected in very low amounts. Experimental controls comprised of single or multiple genes mutations within the glucose transporting phosphotransferase system. Conclusions gluP seems to be regulated only by a putative σA-dependent promoter. The glucose uptake and export assays suggest that GluP is important for glucose export and may act as an exporter. This also supports the role of the glucokinase operon in glucose utilization. PMID:15050034

Cardiac glycoside ouabain induces activation of ATF-1 and StAR expression by interacting with the α4 isoform of the sodium pump in Sertoli cells.

PubMed

Dietze, Raimund; Konrad, Lutz; Shihan, Mazen; Kirch, Ulrike; Scheiner-Bobis, Georgios

2013-03-01

Sertoli cells express α1 and α4 isoforms of the catalytic subunit of Na(+),K(+)-ATPase (sodium pump). Our recent findings demonstrated that interactions of the α4 isoform with cardiotonic steroids (CTS) like ouabain induce signaling cascades that resemble the so-called non-classical testosterone pathway characterized by activation of the c-Src/c-Raf/Erk1/2/CREB signaling cascade. Here we investigate a possible physiological significance of the activated cascade. The results obtained in the current investigation show that the ouabain-induced signaling cascade also leads to the activation of the CREB-related activating transcription factor 1 (ATF-1) in the Sertoli cell line 93RS2 in a concentration- and time-dependent manner, as demonstrated by detection of ATF-1 phosphorylated on Ser63 in western blots. The ouabain-activated ATF-1 protein was found to localize to the cell nuclei. The sodium pump α4 isoform mediates this activation, as it is ablated when cells are incubated with siRNA to the α4 isoform. Ouabain also leads to increased expression of steroidogenic acute regulator (StAR) protein, which has been shown to be a downstream consequence of CREB/ATF-1 activation. Taking into consideration that CTS are most likely produced endogenously, the demonstrated induction of StAR expression by ouabain establishes a link between CTS, the α4 isoform of the sodium pump, and steroidogenesis crucial for male fertility and reproduction. Copyright © 2012 Elsevier B.V. All rights reserved.

21 CFR 884.5150 - Nonpowered breast pump.

Code of Federal Regulations, 2013 CFR

2013-04-01

... device used to express milk from the breast. (b) Classification. Class I. The device is exempt from the... 250 mm Hg suction, and the device materials that contact breast or breast milk do not produce... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Nonpowered breast pump. 884.5150 Section 884.5150...

21 CFR 884.5150 - Nonpowered breast pump.

Code of Federal Regulations, 2010 CFR

2010-04-01

... device used to express milk from the breast. (b) Classification. Class I. The device is exempt from the... 250 mm Hg suction, and the device materials that contact breast or breast milk do not produce... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Nonpowered breast pump. 884.5150 Section 884.5150...

21 CFR 884.5150 - Nonpowered breast pump.

Code of Federal Regulations, 2014 CFR

2014-04-01

... device used to express milk from the breast. (b) Classification. Class I. The device is exempt from the... 250 mm Hg suction, and the device materials that contact breast or breast milk do not produce... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Nonpowered breast pump. 884.5150 Section 884.5150...

21 CFR 884.5150 - Nonpowered breast pump.

Code of Federal Regulations, 2012 CFR

2012-04-01

... device used to express milk from the breast. (b) Classification. Class I. The device is exempt from the... 250 mm Hg suction, and the device materials that contact breast or breast milk do not produce... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Nonpowered breast pump. 884.5150 Section 884.5150...

21 CFR 884.5150 - Nonpowered breast pump.

Code of Federal Regulations, 2011 CFR

2011-04-01

... device used to express milk from the breast. (b) Classification. Class I. The device is exempt from the... 250 mm Hg suction, and the device materials that contact breast or breast milk do not produce... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Nonpowered breast pump. 884.5150 Section 884.5150...

Conjecture regarding posttranslational modifications to the arabidopsis type I proton-pumping pyrophosphatase (AVP1)

USDA-ARS?s Scientific Manuscript database

Agbiotechnology uses genetic engineering to improve the output and value of crops. Altering the expression of the plant Type I Proton-pumping Pyrophosphatase (H+-PPase) has already proven to be a useful tool to enhance crop productivity. Despite the effective use of this gene in translational resear...

Proton-pumping rhodopsins are abundantly expressed by microbial eukaryotes in a high-Arctic fjord.

PubMed

Vader, Anna; Laughinghouse, Haywood D; Griffiths, Colin; Jakobsen, Kjetill S; Gabrielsen, Tove M

2018-02-01

Proton-pumping rhodopsins provide an alternative pathway to photosynthesis by which solar energy can enter the marine food web. Rhodopsin genes are widely found in marine bacteria, also in the Arctic, and were recently reported from several eukaryotic lineages. So far, little is known about rhodopsin expression in Arctic eukaryotes. In this study, we used metatranscriptomics and 18S rDNA tag sequencing to examine the mid-summer function and composition of marine protists (size 0.45-10 µm) in the high-Arctic Billefjorden (Spitsbergen), especially focussing on the expression of microbial proton-pumping rhodopsins. Rhodopsin transcripts were highly abundant, at a level similar to that of genes involved in photosynthesis. Phylogenetic analyses placed the environmental rhodopsins within disparate eukaryotic lineages, including dinoflagellates, stramenopiles, haptophytes and cryptophytes. Sequence comparison indicated the presence of several functional types, including xanthorhodopsins and a eukaryotic clade of proteorhodopsin. Transcripts belonging to the proteorhodopsin clade were also abundant in published metatranscriptomes from other oceanic regions, suggesting a global distribution. The diversity and abundance of rhodopsins show that these light-driven proton pumps play an important role in Arctic microbial eukaryotes. Understanding this role is imperative to predicting the future of the Arctic marine ecosystem faced by a changing light climate due to diminishing sea-ice. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

FAX1, a Novel Membrane Protein Mediating Plastid Fatty Acid Export

PubMed Central

Li, Nannan; Gügel, Irene Luise; Giavalisco, Patrick; Zeisler, Viktoria; Schreiber, Lukas; Soll, Jürgen; Philippar, Katrin

2015-01-01

Fatty acid synthesis in plants occurs in plastids, and thus, export for subsequent acyl editing and lipid assembly in the cytosol and endoplasmatic reticulum is required. Yet, the transport mechanism for plastid fatty acids still remains enigmatic. We isolated FAX1 (fatty acid export 1), a novel protein, which inserts into the chloroplast inner envelope by α-helical membrane-spanning domains. Detailed phenotypic and ultrastructural analyses of FAX1 mutants in Arabidopsis thaliana showed that FAX1 function is crucial for biomass production, male fertility and synthesis of fatty acid-derived compounds such as lipids, ketone waxes, or pollen cell wall material. Determination of lipid, fatty acid, and wax contents by mass spectrometry revealed that endoplasmatic reticulum (ER)-derived lipids decreased when FAX1 was missing, but levels of several plastid-produced species increased. FAX1 over-expressing lines showed the opposite behavior, including a pronounced increase of triacyglycerol oils in flowers and leaves. Furthermore, the cuticular layer of stems from fax1 knockout lines was specifically reduced in C29 ketone wax compounds. Differential gene expression in FAX1 mutants as determined by DNA microarray analysis confirmed phenotypes and metabolic imbalances. Since in yeast FAX1 could complement for fatty acid transport, we concluded that FAX1 mediates fatty acid export from plastids. In vertebrates, FAX1 relatives are structurally related, mitochondrial membrane proteins of so-far unknown function. Therefore, this protein family might represent a powerful tool not only to increase lipid/biofuel production in plants but also to explore novel transport systems involved in vertebrate fatty acid and lipid metabolism. PMID:25646734

PigZ, a TetR/AcrR family repressor, modulates secondary metabolism via the expression of a putative four-component resistance-nodulation-cell-division efflux pump, ZrpADBC, in Serratia sp. ATCC 39006.

PubMed

Gristwood, Tamzin; Fineran, Peter C; Everson, Lee; Salmond, George P C

2008-07-01

The Gram-negative enterobacterium, Serratia sp. ATCC 39006 synthesizes several secondary metabolites, including prodigiosin (Pig) and a carbapenem antibiotic (Car). A complex hierarchical network of regulatory proteins control Pig and Car production. In this study we characterize a TetR family regulator, PigZ, which represses transcription of a divergently transcribed putative resistance-nodulation-cell-division (RND) efflux pump, encoded by zrp (PigZ repressed pump) ADBC, via direct binding to the zrpA-pigZ intergenic region. Unusually, this putative RND pump contains two predicted membrane fusion proteins (MFPs), ZrpA and ZrpD. A mutation in pigZ resulted in multiple phenotypic changes, including exoenzyme production, motility and differential regulation of Pig and Car production. A polar suppressor mutation, within zrpA, restored all tested phenotypes to parental strain levels, indicating that the changes observed are due to the increase in expression of ZrpADBC in the absence of the repressor, PigZ. Genomic deletions of zrpA and zrpD indicate that the MFP ZrpD, but not ZrpA, is essential for activity of the putative pump. Bioinformatic analysis revealed that putative RND efflux pumps encoding two MFP components are not uncommon, particularly among plant-associated, Gram-negative bacteria. In addition, based on phylogenetic analysis, we propose that these pairs of MFPs consist of two distinct subtypes.

"Breastfeeding" without baby: A longitudinal, qualitative investigation of how mothers perceive, feel about, and practice human milk expression.

PubMed

Felice, Julia P; Geraghty, Sheela R; Quaglieri, Caroline W; Yamada, Rei; Wong, Adriana J; Rasmussen, Kathleen M

2017-07-01

Most American mothers who produce human milk (HM) now pump in place of some or all feeding at the breast, and most American infants are now fed pumped HM. We aimed to investigate mothers' perceptions of, attitudes toward, and practices for pumping and providing pumped HM. Results related to pumping are reported here. We conducted in-depth, semi-structured interviews among a diverse sample of 20 mothers who pumped, following each from pregnancy through infant HM-feeding cessation up to 1 year postpartum. Data were analyzed using thematic analysis with Atlas.ti. Mothers' reasons for pumping changed over time and reflected their needs and desires (e.g., latch difficulty, return to work, and increasing their milk supply). Mothers reported that pump type and quality were important to pumping success and that pumping was time-consuming, costly, and unpleasant compared to feeding at the breast. Regardless of how often mothers pumped, most felt pumping was necessary to meet their infant HM-feeding goals and was a welcome means of sharing with other caregivers the bonding opportunity and tasks they associated with feeding infants. Mothers interpreted output from pumping sessions to understand their ability to provide enough milk to meet their infants' needs. Mothers' reasons for pumping may signal constraints to infant HM feeding that may be addressed with policy changes. Mothers' attitudes and perceptions toward pumping indicate that, although pumping fills important and welcome roles for many mothers, the reality of its practice may make it an unacceptable or infeasible substitute for some. © 2017 John Wiley & Sons Ltd.

The Quiet Revolution: Breastfeeding Transformed With the Use of Breast Pumps

PubMed Central

Geraghty, Sheela R.

2011-01-01

A quiet revolution has been taking place in the feeding of US infants in the form of women using electric breast pumps. This revolution in milk expression may be a boon for both mothers and infants if more infants are fed human milk or if they receive human milk for a longer period. Milk expression may also be problematic for mothers, and it may be particularly problematic for infants if they are fed too much, fed milk of an inappropriate composition, or fed milk that is contaminated. As a result, the time has come to determine the prevalence of exclusive and periodic breast milk expression and the consequences of these behaviors for the health of mothers and their infants. PMID:21680919

Significantly higher activity of a cytoplasmic hammerhead ribozyme than a corresponding nuclear counterpart: engineered tRNAs with an extended 3′ end can be exported efficiently and specifically to the cytoplasm in mammalian cells

PubMed Central

Kuwabara, Tomoko; Warashina, Masaki; Koseki, Shiori; Sano, Masayuki; Ohkawa, Jun; Nakayama, Kazuhisa; Taira, Kazunari

2001-01-01

Hammerhead ribozymes were expressed under the control of similar tRNA promoters, localizing transcripts either in the cytoplasm or the nucleus. The tRNAVal-driven ribozyme (tRNA-Rz; tRNA with extra sequences at the 3′ end) that has been used in our ribozyme studies was exported efficiently into the cytoplasm and ribozyme activity was detected only in the cytoplasmic fraction. Both ends of the transported tRNA-Rz were characterized comprehensively and the results confirmed that tRNA-Rz had unprocessed 5′ and 3′ ends. Furthermore, it was also demonstrated that the activity of the exported ribozyme was significantly higher than that of the ribozyme which remained in the nucleus. We suggest that it is possible to engineer tRNA-Rz, which can be exported to the cytoplasm based on an understanding of secondary structures, and then tRNA-driven ribozymes may be co-localized with their target mRNAs in the cytoplasm of mammalian cells. PMID:11433023

Small RNA sequencing in cells and exosomes identifies eQTLs and 14q32 as a region of active export

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

Tsang, Emily K.; Abell, Nathan S.; Li, Xin

Exosomes are small extracellular vesicles that carry heterogeneous cargo, including RNA, between cells. Increasing evidence suggests that exosomes are important mediators of intercellular communication and biomarkers of disease. Despite this, the variability of exosomal RNA between individuals has not been well quantified. To assess this variability, we sequenced the small RNA of cells and exosomes from a 17-member family. Across individuals, we show that selective export of miRNAs occurs not only at the level of specific transcripts, but that a cluster of 74 mature miRNAs on chromosome 14q32 is massively exported in exosomes while mostly absent from cells. We alsomore » observe more interindividual variability between exosomal samples than between cellular ones and identify four miRNA expression quantitative trait loci shared between cells and exosomes. Lastly, our findings indicate that genomically colocated miRNAs can be exported together and highlight the variability in exosomal miRNA levels between individuals as relevant for exosome use as diagnostics.« less

Small RNA sequencing in cells and exosomes identifies eQTLs and 14q32 as a region of active export

DOE PAGES

Tsang, Emily K.; Abell, Nathan S.; Li, Xin; ...

2016-10-31

Exosomes are small extracellular vesicles that carry heterogeneous cargo, including RNA, between cells. Increasing evidence suggests that exosomes are important mediators of intercellular communication and biomarkers of disease. Despite this, the variability of exosomal RNA between individuals has not been well quantified. To assess this variability, we sequenced the small RNA of cells and exosomes from a 17-member family. Across individuals, we show that selective export of miRNAs occurs not only at the level of specific transcripts, but that a cluster of 74 mature miRNAs on chromosome 14q32 is massively exported in exosomes while mostly absent from cells. We alsomore » observe more interindividual variability between exosomal samples than between cellular ones and identify four miRNA expression quantitative trait loci shared between cells and exosomes. Lastly, our findings indicate that genomically colocated miRNAs can be exported together and highlight the variability in exosomal miRNA levels between individuals as relevant for exosome use as diagnostics.« less

Induction of endoplasmic reticulum calcium pump expression during early leukemic B cell differentiation.

PubMed

Aït Ghezali, Lamia; Arbabian, Atousa; Roudot, Hervé; Brouland, Jean-Philippe; Baran-Marszak, Fanny; Salvaris, Evelyn; Boyd, Andrew; Drexler, Hans G; Enyedi, Agnes; Letestu, Remi; Varin-Blank, Nadine; Papp, Bela

2017-06-26

Endoplasmic reticulum (ER) calcium storage and release play important roles in B lymphocyte maturation, survival, antigen-dependent cell activation and immunoglobulin synthesis. Calcium is accumulated in the endoplasmic reticulum (ER) by Sarco/Endoplasmic Reticulum Calcium ATPases (SERCA enzymes). Because lymphocyte function is critically dependent on SERCA activity, it is important to understand qualitative and quantitative changes of SERCA protein expression that occur during B lymphoid differentiation and leukemogenesis. In this work we investigated the modulation of SERCA expression during the pharmacologically induced differentiation of leukemic precursor B lymphoblast cell lines that carry the E2A-PBX1 fusion oncoprotein. Changes of SERCA levels during differentiation were determined and compared to those of established early B lymphoid differentiation markers. SERCA expression of the cells was compared to that of mature B cell lines as well, and the effect of the direct inhibition of SERCA-dependent calcium transport on the differentiation process was investigated. We show that E2A-PBX1 + leukemia cells simultaneously express SERCA2 and SERCA3-type calcium pumps; however, their SERCA3 expression is markedly inferior to that of mature B cells. Activation of protein kinase C enzymes by phorbol ester leads to phenotypic differentiation of the cells, and this is accompanied by the induction of SERCA3 expression. Direct pharmacological inhibition of SERCA-dependent calcium transport during phorbol ester treatment interferes with the differentiation process. These data show that the calcium pump composition of the ER is concurrent with increased SERCA3 expression during the differentiation of precursor B acute lymphoblastic leukemia cells, that a cross-talk exists between SERCA function and the control of differentiation, and that SERCA3 may constitute an interesting new marker for the study of early B cell phenotype.

Multiple alterations of canalicular membrane transport activities in rats with CCl(4)-induced hepatic injury.

PubMed

Song, Im-Sook; Lee, Young-Mi; Chung, Suk-Jae; Shim, Chang-Koo

2003-04-01

The influence of CCl(4)-induced experimental hepatic injury (CCl(4)-EHI) on the expression and transport activities of primary active transporters on the canalicular membrane, including P-glycoprotein (P-gp), a bile salt export pump (Bsep) and a multidrug resistance associated protein2 (Mrp2), was assessed. CCl(4)-EHI was induced by an intraperitoneal injection of CCl(4) to rats at a dose of 1 ml/kg 24 h prior to the preparation of canalicular liver plasma membrane (cLPM) vesicles and pharmacokinetic studies. The expression of each transporter was measured for the vesicles via Western blot analysis at 6, 12, 24, 36, and 48 h after the injection of CCl(4). The in vivo canalicular excretion clearance (CL(exc)) of [(3)H]daunomycin, [(3)H]taurocholate and [(3)H]17beta-estradiol-17beta-D-glucuronide (E(2)17betaG), representative substrates of P-gp, Bsep, and Mrp2, respectively, was determined following an i.v. infusion to rats. The uptake of each substrate into cLPM vesicles in the presence of ATP was also measured by a rapid filtration technique. As the result of the CCl(4)-EHI, the protein level of transporters was altered as a function of time in multiple manners; it was increased by 3.6-fold for P-gp, unchanged for Bsep, and decreased by 73% for Mrp2 at 24 h. The in vivo CL(exc) and the intrinsic uptake clearance into cLPM vesicles (CL(int)) at 24 h after the CCl(4) injection (CCl(4)-EHI(24 h)) were also influenced by the EHI in a similar manner; they were increased by 1.8- and 1.9-fold for daunomycin, unchanged for taurocholate, and decreased by 41 and 39% for E(2)17betaG, respectively, consistent with multiple alterations in the expression of the relevant transporters.

Climate change impacts on net primary production (NPP) and export production (EP) regulated by increasing stratification and phytoplankton community structure in the CMIP5 models

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

Fu, Weiwei; Randerson, James T.; Moore, J. Keith

We examine climate change impacts on net primary production (NPP) and export production (sinking particulate flux; EP) with simulations from nine Earth system models (ESMs) performed in the framework of the fifth phase of the Coupled Model Intercomparison Project (CMIP5). Global NPP and EP are reduced by the end of the century for the intense warming scenario of Representative Concentration Pathway (RCP) 8.5. Relative to the 1990s, NPP in the 2090s is reduced by 2–16% and EP by 7–18%. The models with the largest increases in stratification (and largest relative declines in NPP and EP) also show the largest positivemore » biases in stratification for the contemporary period, suggesting overestimation of climate change impacts on NPP and EP. All of the CMIP5 models show an increase in stratification in response to surface–ocean warming and freshening, which is accompanied by decreases in surface nutrients, NPP and EP. There is considerable variability across the models in the magnitudes of NPP, EP, surface nutrient concentrations and their perturbations by climate change. The negative response of NPP and EP to increasing stratification reflects primarily a bottom-up control, as upward nutrient flux declines at the global scale. Models with dynamic phytoplankton community structure show larger declines in EP than in NPP. This pattern is driven by phytoplankton community composition shifts, with reductions in productivity by large phytoplankton as smaller phytoplankton (which export less efficiently) are favored under the increasing nutrient stress. Thus, the projections of the NPP response to climate change are critically dependent on the simulated phytoplankton community structure, the efficiency of the biological pump and the resulting levels of regenerated production, which vary widely across the models. In conclusion, community structure is represented simply in the CMIP5 models, and should be expanded to better capture the spatial patterns and climate-driven changes in export efficiency.« less

Climate change impacts on net primary production (NPP) and export production (EP) regulated by increasing stratification and phytoplankton community structure in the CMIP5 models

DOE PAGES

Fu, Weiwei; Randerson, James T.; Moore, J. Keith

2016-09-16

We examine climate change impacts on net primary production (NPP) and export production (sinking particulate flux; EP) with simulations from nine Earth system models (ESMs) performed in the framework of the fifth phase of the Coupled Model Intercomparison Project (CMIP5). Global NPP and EP are reduced by the end of the century for the intense warming scenario of Representative Concentration Pathway (RCP) 8.5. Relative to the 1990s, NPP in the 2090s is reduced by 2–16% and EP by 7–18%. The models with the largest increases in stratification (and largest relative declines in NPP and EP) also show the largest positivemore » biases in stratification for the contemporary period, suggesting overestimation of climate change impacts on NPP and EP. All of the CMIP5 models show an increase in stratification in response to surface–ocean warming and freshening, which is accompanied by decreases in surface nutrients, NPP and EP. There is considerable variability across the models in the magnitudes of NPP, EP, surface nutrient concentrations and their perturbations by climate change. The negative response of NPP and EP to increasing stratification reflects primarily a bottom-up control, as upward nutrient flux declines at the global scale. Models with dynamic phytoplankton community structure show larger declines in EP than in NPP. This pattern is driven by phytoplankton community composition shifts, with reductions in productivity by large phytoplankton as smaller phytoplankton (which export less efficiently) are favored under the increasing nutrient stress. Thus, the projections of the NPP response to climate change are critically dependent on the simulated phytoplankton community structure, the efficiency of the biological pump and the resulting levels of regenerated production, which vary widely across the models. In conclusion, community structure is represented simply in the CMIP5 models, and should be expanded to better capture the spatial patterns and climate-driven changes in export efficiency.« less

A linkage between Asian dust, dissolved iron and marine export production in the deep ocean

NASA Astrophysics Data System (ADS)

Han, Yongxiang; Zhao, Tianliang; Song, Lianchun; Fang, Xiaomin; Yin, Yan; Deng, Zuqin; Wang, Suping; Fan, Shuxian

2011-08-01

Iron-addition experiments have revealed that iron supply exerts controls on biogeochemical cycles in the ocean and ultimately influences the Earth's climate system. The iron hypothesis in its broad outlines has been proved to be correct. However, the hypothesis needs to be verified with an observable biological response to specific dust deposition events. Plankton growth following the Asian dust storm over Ocean Station PAPA (50°N, 145°W) in the North Pacific Ocean in April 2001 was the first supportive evidence of natural aeolian iron inputs to ocean; The data were obtained through the SeaWiFS satellite and robot carbon explorers by Bishop et al. Using the NARCM modeling results in this study, the calculated total dust deposition flux was 35 mg m -2 per day in PAPA region from the dust storm of 11-13 April, 2001 into 0.0615 mg m -2 d -1 (about 1100 nM) soluble iron in the surface layer at Station PAPA. It was enough for about 1100 nM to enhance the efficiency of the marine biological pump and trigger the rapid increase of POC and chlorophyll. The iron fertilization hypothesis therefore is plausible. However, even if this specific dust event can support the iron fertilization hypothesis, long-term observation data are lacking in marine export production and continental dust. In this paper, we also conducted a simple correlation analysis between the diatoms and foraminifera at about 3000 m and 4000 m at two subarctic Pacific stations and the dust aerosol production from China's mainland. The correlation coefficient between marine export production and dust storm frequency in the core area of the dust storms was significantly high, suggesting that aerosols generated by Asian dust storm are the source of iron for organic matter fixation in the North Pacific Ocean. These results suggest that there could be an interlocking chain for the change of atmospheric dust aerosol-soluble iron-marine export production.

The glacial iron cycle from source to export

NASA Astrophysics Data System (ADS)

Hawkings, J.; Wadham, J. L.; Tranter, M.; Raiswell, R.; Benning, L. G.; Statham, P. J.; Tedstone, A. J.; Nienow, P. W.; Telling, J.; Bagshaw, E.; Simmons, S. L.

2014-12-01

Nutrient availability limits primary production in large sectors of the world's oceans. Iron is the major limiting nutrient in around one third of the oceanic euphotic zone, most significantly in the Southern Ocean proximal to Antarctica. In these areas the availability of bioavailable iron can influence the amount of primary production, and thus the strength of the biological pump and associated carbon drawdown from the atmosphere. Despite experiencing widespread iron limitation, the Polar oceans are among the most productive on Earth. Due to the extreme cold, remoteness and their perceived "stasis", ice sheets have previously been though of as insignificant in global biogeochemical cycles. However, large marine algal blooms have been observed in iron-limited areas where glacial influence is large, and it is possible that these areas are stimulated by glacial bioavailable iron input. Here we discuss the importance of the Greenland and Antarctic ice sheets in the global iron cycle. Using field collected trace element data, bulk meltwater chemistry and mineralogical analysis, including photomicrographs, EELS and XANES, we present, for the first time, a conceptual model of the glacial iron cycle from source to export. Using this data we discuss the sources of iron in glacial meltwater, transportation and alteration through the glacial system, and subsequent export to downstream environments. Data collected in 2012 and 2013 from two different Greenlandic glacial catchments are shown, with the most detailed breakdown of iron speciation and concentrations in glacial areas yet reported. Furthermore, the first data from Greenlandic icebergs is presented, allowing meltwater-derived and iceberg-derived iron export to be compared, and the influence of both in marine productivity to be estimated. Using our conceptual model and flux estimates from our dataset, glacial iron delivery in both the northern and southern hemisphere is discussed. Finally, we compare our flux estimates to other major iron sources to the polar regions such as aeolian dust, and discuss potential implications of increased melting of the ice sheets on the global iron cycle in the future.

Detergent Screening and Purification of the Human Liver ABC Transporters BSEP (ABCB11) and MDR3 (ABCB4) Expressed in the Yeast Pichia pastoris

PubMed Central

Stindt, Jan; Smits, Sander H. J.; Schmitt, Lutz

2013-01-01

The human liver ATP-binding cassette (ABC) transporters bile salt export pump (BSEP/ABCB11) and the multidrug resistance protein 3 (MDR3/ABCB4) fulfill the translocation of bile salts and phosphatidylcholine across the apical membrane of hepatocytes. In concert with ABCG5/G8, these two transporters are responsible for the formation of bile and mutations within these transporters can lead to severe hereditary diseases. In this study, we report the heterologous overexpression and purification of human BSEP and MDR3 as well as the expression of the corresponding C-terminal GFP-fusion proteins in the yeast Pichia pastoris. Confocal laser scanning microscopy revealed that BSEP-GFP and MDR3-GFP are localized in the plasma membrane of P. pastoris. Furthermore, we demonstrate the first purification of human BSEP and MDR3 yielding ∼1 mg and ∼6 mg per 100 g of wet cell weight, respectively. By screening over 100 detergents using a dot blot technique, we found that only zwitterionic, lipid-like detergents such as Fos-cholines or Cyclofos were able to extract both transporters in sufficient amounts for subsequent functional analysis. For MDR3, fluorescence-detection size exclusion chromatography (FSEC) screens revealed that increasing the acyl chain length of Fos-Cholines improved monodispersity. BSEP purified in n-dodecyl-β-D-maltoside or Cymal-5 after solubilization with Fos-choline 16 from P. pastoris membranes showed binding to ATP-agarose. Furthermore, detergent-solubilized and purified MDR3 showed a substrate-inducible ATPase activity upon addition of phosphatidylcholine lipids. These results form the basis for further biochemical analysis of human BSEP and MDR3 to elucidate the function of these clinically relevant ABC transporters. PMID:23593265

Vectorial transport of unconjugated and conjugated bile salts by monolayers of LLC-PK1 cells doubly transfected with human NTCP and BSEP or with rat Ntcp and Bsep.

PubMed

Mita, Sachiko; Suzuki, Hiroshi; Akita, Hidetaka; Hayashi, Hisamitsu; Onuki, Reiko; Hofmann, Alan F; Sugiyama, Yuichi

2006-03-01

Na(+)-taurocholate-cotransporting peptide (NTCP)/SLC10A1 and bile salt export pump (BSEP)/ABCB11 synergistically play an important role in the transport of bile salts by the hepatocyte. In this study, we transfected human NTCP and BSEP or rat Ntcp and Bsep into LLC-PK1 cells, a cell line devoid of bile salts transporters. Transport by these cells was characterized with a focus on substrate specificity between rats and humans. The basal to apical flux of taurocholate across NTCP- and BSEP-expressing LLC-PK1 monolayers was 10 times higher than that in the opposite direction, whereas the flux across the monolayer of control and NTCP or BSEP single-expressing cells did not show any vectorial transport. The basal to apical flux of taurocholate was saturated with a K(m) value of 20 microM. Vectorial transcellular transport was also observed for cholate, chenodeoxycholate, ursodeoxycholate, their taurine and glycine conjugates, and taurodeoxycholate and glycodeoxycholate, whereas no transport of lithocholate was detected. To evaluate the respective functions of NTCP and BSEP and to compare them with those of rat Ntcp and Bsep, we calculated the clearance by each transporter in this system. A good correlation in the clearance of the examined bile salts (cholate, chenodeoxycholate, ursodeoxycholate, and their taurine or glycine conjugates) was observed between transport by human and that of rat transporters in terms of their rank order: for NTCP, taurine conjugates > glycine conjugates > unconjugated bile salts, and for BSEP, unconjugated bile salts and glycine conjugates > taurine conjugates. In conclusion, the substrate specificity of human and rat NTCP and BSEP appear to be very similar at least for monovalent bile salts under physiological conditions.

PRIMING THE PUMP AND CONTROLLING THE FLOW.

ERIC Educational Resources Information Center

BUCHAN, VIVIAN

THE BEGINNING WRITER NEEDS BOTH ENCOURAGEMENT AND DIRECTION. ONCE A STUDENT, NONVERBAL OR FLUENT, HAS EXPRESSED AN OPINION, SIGNIFICANT OR TRIVIAL, THE PUMP CAN BE PRIMED BY ASKING HIM "WHY," AND HIS FLOW OF "BECAUSES" CAN BE CONTROLLED BY CHANNELING THEM INTO A SIMPLE PATTERN. THE NONVERBAL STUDENT IS ENCOURAGED TO WRITE WHEN HE LEARNS THAT A…

PI3K class II α regulates δ-opioid receptor export from the trans-Golgi network.

PubMed

Shiwarski, Daniel J; Darr, Marlena; Telmer, Cheryl A; Bruchez, Marcel P; Puthenveedu, Manojkumar A

2017-08-01

The interplay between signaling and trafficking by G protein-coupled receptors (GPCRs) has focused mainly on endocytic trafficking. Whether and how surface delivery of newly synthesized GPCRs is regulated by extracellular signals is less understood. Here we define a signaling-regulated checkpoint at the trans -Golgi network (TGN) that controls the surface delivery of the delta opioid receptor (δR). In PC12 cells, inhibition of phosphoinositide-3 kinase (PI3K) activity blocked export of newly synthesized δR from the Golgi and delivery to the cell surface, similar to treatment with nerve growth factor (NGF). Depletion of class II phosphoinositide-3 kinase α (PI3K C2A), but not inhibition of class I PI3K, blocked δR export to comparable levels and attenuated δR-mediated cAMP inhibition. NGF treatment displaced PI3K C2A from the Golgi and optogenetic recruitment of the PI3K C2A kinase domain to the TGN-induced δR export downstream of NGF. Of importance, PI3K C2A expression promotes export of endogenous δR in primary trigeminal ganglion neurons. Taken together, our results identify PI3K C2A as being required and sufficient for δR export and surface delivery in neuronal cells and suggest that it could be a key modulator of a novel Golgi export checkpoint that coordinates GPCR delivery to the surface. © 2017 Shiwarski et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Heterologous Expression and Isolation of Influenza A Virus Nuclear Export Protein NEP.

PubMed

Golovko, A O; Koroleva, O N; Drutsa, V L

2017-12-01

Influenza A virus nuclear export protein NEP (NS2, 14.4 kDa) plays a key role in various steps of the virus life cycle. Highly purified protein preparations are required for structural and functional studies. In this study, we designed a series of Escherichia coli plasmid constructs for highly efficient expression of the NEP gene under control of the constitutive trp promoter. An efficient method for extraction of NEP from inclusion bodies based on dodecyl sulfate treatment was developed. Preparations of purified NEP with either N- or C-terminal (His) 6 -tag were obtained using Ni-NTA agarose affinity chromatography with yield of more than 20 mg per liter of culture. According to CD data, the secondary structure of the proteins matched that of natural NEP. A high propensity of NEP to aggregate over a wide range of conditions was observed.

Identification of a functional nuclear export signal in the green fluorescent protein asFP499

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

Mustafa, Huseyin; Strasser, Bernd; Rauth, Sabine

2006-04-21

The green fluorescent protein (GFP) asFP499 from Anemonia sulcata is a distant homologue of the GFP from Aequorea victoria. We cloned the asFP499 gene into a mammalian expression vector and showed that this protein was expressed in the human lymphoblast cell line Ramos RA1 and in the embryonic kidney 293T cell line (HEK 293T). In HEK 293T cells, asFP499 was localized mainly in the cytoplasm, suggesting that the protein was excluded from the nucleus. We identified {sub 194}LRMEKLNI{sub 201} as a candidate nuclear export signal in asFP499 and mutated the isoleucine at position 201 to an alanine. Unlike the wildtypemore » form, the mutant protein was distributed throughout the cytoplasm and nucleus. This is First report of a GFP that contains a functional NES.« less

Identification of Antibody and Small Molecule Antagonists of Ferroportin-Hepcidin Interaction

PubMed Central

Ross, Sandra L.; Biswas, Kaustav; Rottman, James; Allen, Jennifer R.; Long, Jason; Miranda, Les P.; Winters, Aaron; Arvedson, Tara L.

2017-01-01

The iron exporter ferroportin and its ligand, the hormone hepcidin, control fluxes of stored and recycled iron for use in a variety of essential biochemical processes. Inflammatory disorders and malignancies are often associated with high hepcidin levels, leading to ferroportin down-regulation, iron sequestration in tissue macrophages and subsequent anemia. The objective of this research was to develop reagents to characterize the expression of ferroportin, the interaction between ferroportin and hepcidin, as well as to identify novel ferroportin antagonists capable of maintaining iron export in the presence of hepcidin. Development of investigative tools that enabled cell-based screening assays is described in detail, including specific and sensitive monoclonal antibodies that detect endogenously-expressed human and mouse ferroportin and fluorescently-labeled chemically-synthesized human hepcidin. Large and small molecule antagonists inhibiting hepcidin-mediated ferroportin internalization were identified, and unique insights into the requirements for interaction between these two key iron homeostasis molecules are provided. PMID:29209212

Replacement of the yeast TRP4 3' untranslated region by a hammerhead ribozyme results in a stable and efficiently exported mRNA that lacks a poly(A) tail.

PubMed Central

Düvel, Katrin; Valerius, Oliver; Mangus, David A; Jacobson, Allan; Braus, Gerhard H

2002-01-01

The mRNA poly(A) tail serves different purposes, including the facilitation of nuclear export, mRNA stabilization, efficient translation, and, finally, specific degradation. The posttranscriptional addition of a poly(A) tail depends on sequence motifs in the 3' untranslated region (3' UTR) of the mRNA and a complex trans-acting protein machinery. In this study, we have replaced the 3' UTR of the yeast TRP4 gene with sequences encoding a hammerhead ribozyme that efficiently cleaves itself in vivo. Expression of the TRP4-ribozyme allele resulted in the accumulation of a nonpolyadenylated mRNA. Cells expressing the TRP4-ribozyme mRNA showed a reduced growth rate due to a reduction in Trp4p enzyme activity. The reduction in enzyme activity was not caused by inefficient mRNA export from the nucleus or mRNA destabilization. Rather, analyses of mRNA association with polyribosomes indicate that translation of the ribozyme-containing mRNA is impaired. This translational defect allows sufficient synthesis of Trp4p to support growth of trp4 cells, but is, nevertheless, of such magnitude as to activate the general control network of amino acid biosynthesis. PMID:12003493

Design and Specification of Low Pressure Sewer Systems for Recreation Areas.

DTIC Science & Technology

1985-02-01

holding tank receives wastewater Iflows by gravity. Level scitsors act ivate the grinder pump operation pump at preset levels. Emergency 40 overf low aid...above the septic 0 Lank floor, the septic tank should be secured against flotation . Test in. 70. Septir t.ank s shouldt Ihe wat’rtight and must be...8217 , pump i rig je,j , aind powt, r cost . Annual power cost can he estimated using the fo I I owi rig express ion: APC =ae PC (4) whe -e 0 APIC annual

The amino-terminal hydrophilic region of the vacuolar transporter Avt3p is dispensable for the vacuolar amino acid compartmentalization of Schizosaccharomyces pombe.

PubMed

Kawano-Kawada, Miyuki; Chardwiriyapreecha, Soracom; Manabe, Kunio; Sekito, Takayuki; Akiyama, Koichi; Takegawa, Kaoru; Kakinuma, Yoshimi

2016-12-01

Avt3p, a vacuolar amino acid exporter (656 amino acid residues) that is important for vacuolar amino acid compartmentalization as well as spore formation in Schizosaccharomyces pombe, has an extremely long hydrophilic region (approximately 290 amino acid residues) at its N-terminus. Because known functional domains have not been found in this region, its functional role was examined with a deletion mutant avt3 (∆1-270) expressed in S. pombe avt3∆ cells. The deletion of this region did not affect its intracellular localization or vacuolar contents of basic amino acids as well as neutral ones. The defect of avt3Δ cells in spore formation was rescued by the expression of avt3 + but was not completely rescued by the expression of avt3 (∆1-270) . The N-terminal region is thus dispensable for the function of Avt3p as an amino acid exporter, but it is likely to be involved in the role of Avt3p under nutritional starvation conditions.

The Nuclear Pore-Associated TREX-2 Complex Employs Mediator to Regulate Gene Expression

PubMed Central

Schneider, Maren; Hellerschmied, Doris; Schubert, Tobias; Amlacher, Stefan; Vinayachandran, Vinesh; Reja, Rohit; Pugh, B. Franklin; Clausen, Tim; Köhler, Alwin

2015-01-01

Summary Nuclear pore complexes (NPCs) influence gene expression besides their established function in nuclear transport. The TREX-2 complex localizes to the NPC basket and affects gene-NPC interactions, transcription, and mRNA export. How TREX-2 regulates the gene expression machinery is unknown. Here, we show that TREX-2 interacts with the Mediator complex, an essential regulator of RNA Polymerase (Pol) II. Structural and biochemical studies identify a conserved region on TREX-2, which directly binds the Mediator Med31/Med7N submodule. TREX-2 regulates assembly of Mediator with the Cdk8 kinase and is required for recruitment and site-specific phosphorylation of Pol II. Transcriptome and phenotypic profiling confirm that TREX-2 and Med31 are functionally interdependent at specific genes. TREX-2 additionally uses its Mediator-interacting surface to regulate mRNA export suggesting a mechanism for coupling transcription initiation and early steps of mRNA processing. Our data provide mechanistic insight into how an NPC-associated adaptor complex accesses the core transcription machinery. PMID:26317468

Hepatic 3D spheroid models for the detection and study of compounds with cholestatic liability

PubMed Central

Hendriks, Delilah F. G.; Fredriksson Puigvert, Lisa; Messner, Simon; Mortiz, Wolfgang; Ingelman-Sundberg, Magnus

2016-01-01

Drug-induced cholestasis (DIC) is poorly understood and its preclinical prediction is mainly limited to assessing the compound’s potential to inhibit the bile salt export pump (BSEP). Here, we evaluated two 3D spheroid models, one from primary human hepatocytes (PHH) and one from HepaRG cells, for the detection of compounds with cholestatic liability. By repeatedly co-exposing both models to a set of compounds with different mechanisms of hepatotoxicity and a non-toxic concentrated bile acid (BA) mixture for 8 days we observed a selective synergistic toxicity of compounds known to cause cholestatic or mixed cholestatic/hepatocellular toxicity and the BA mixture compared to exposure to the compounds alone, a phenomenon that was more pronounced after extending the exposure time to 14 days. In contrast, no such synergism was observed after both 8 and 14 days of exposure to the BA mixture for compounds that cause non-cholestatic hepatotoxicity. Mechanisms behind the toxicity of the cholestatic compound chlorpromazine were accurately detected in both spheroid models, including intracellular BA accumulation, inhibition of ABCB11 expression and disruption of the F-actin cytoskeleton. Furthermore, the observed synergistic toxicity of chlorpromazine and BA was associated with increased oxidative stress and modulation of death receptor signalling. Combined, our results demonstrate that the hepatic spheroid models presented here can be used to detect and study compounds with cholestatic liability. PMID:27759057

Transgenic Overexpression of Abcb11 Enhances Biliary Bile Salt Outputs, But Does Not Affect Cholesterol Cholelithogenesis in Mice

PubMed Central

Wang, Helen H.; Lammert, Frank; Schmitz, Anne; Wang, David Q.-H.

2010-01-01

Background Cholesterol gallstone disease is a complex genetic trait and induced by multiple but as yet unknown genes. A major Lith gene, Lith1 was first identified on chromosome 2 in gallstone-susceptible C57L mice compared with resistant AKR mice. Abcb11, encoding the canalicular bile salt export pump in the hepatocyte, co-localizes with the Lith1 QTL region and its hepatic expression is significantly higher in C57L mice than in AKR mice. Material and methods To investigate whether Abcb11 influences cholesterol gallstone formation, we created an Abcb11 transgenic strain on the AKR genetic background and fed these mice with a lithogenic diet for 56 days. Result We excluded functionally relevant polymorphisms of the Abcb11 gene and its promoter region between C57L and AKR mice. Overexpression of Abcb11 significantly promoted biliary bile salt secretion and increased circulating bile salt pool size and bile salt-dependent bile flow rate. However, biliary cholesterol and phospholipid secretion, as well as gallbladder size and contractility were comparable in transgenic and wild-type mice. At 56 days on the lithogenic diet, cholesterol saturation indexes of gallbladder biles and gallstone prevalence rates were essentially similar in these two groups of mice. Conclusion Overexpression of Abcb11 augments biliary bile salt secretion, but does not affect cholelithogenesis in mice. PMID:20456485

Deep Transcriptomic Analysis of Black Rockfish (Sebastes schlegelii) Provides New Insights on Responses to Acute Temperature Stress.

PubMed

Lyu, Likang; Wen, Haishen; Li, Yun; Li, Jifang; Zhao, Ji; Zhang, Simin; Song, Min; Wang, Xiaojie

2018-06-14

In the present study, we conducted an RNA-Seq analysis to characterize the genes and pathways involved in acute thermal and cold stress responses in the liver of black rockfish, a viviparous teleost that has the ability to cope with a wide range of temperature changes. A total of 584 annotated differentially expressed genes (DEGs) were identified in all three comparisons (HT vs NT, HT vs LT and LT vs NT). Based on an enrichment analysis, DEGs with a potential role in stress accommodation were classified into several categories, including protein folding, metabolism, immune response, signal transduction, molecule transport, membrane, and cell proliferation/apoptosis. Considering that thermal stress has a greater effect than cold stress in black rockfish, 24 shared DEGs in the intersection of the HT vs LT and HT vs NT groups were enriched in 2 oxidation-related gene ontology (GO) terms. Nine important heat-stress-reducing pathways were significantly identified and classified into 3 classes: immune and infectious diseases, organismal immune system and endocrine system. Eight DEGs (early growth response protein 1, bile salt export pump, abcb11, hsp70a, rtp3, 1,25-dihydroxyvitamin d(3) 24-hydroxylase, apoa4, transcription factor jun-b-like and an uncharacterized gene) were observed among all three comparisons, strongly implying their potentially important roles in temperature stress responses.

Conjecture Regarding Posttranslational Modifications to the Arabidopsis Type I Proton-Pumping Pyrophosphatase (AVP1)

PubMed Central

Pizzio, Gaston A.; Hirschi, Kendal D.; Gaxiola, Roberto A.

2017-01-01

Agbiotechnology uses genetic engineering to improve the output and value of crops. Altering the expression of the plant Type I Proton-pumping Pyrophosphatase (H+-PPase) has already proven to be a useful tool to enhance crop productivity. Despite the effective use of this gene in translational research, information regarding the intracellular localization and functional plasticity of the pump remain largely enigmatic. Using computer modeling several putative phosphorylation, ubiquitination and sumoylation target sites were identified that may regulate Arabidopsis H+-PPase (AVP1- Arabidopsis Vacuolar Proton-pump 1) subcellular trafficking and activity. These putative regulatory sites will direct future research that specifically addresses the partitioning and transport characteristics of this pump. We posit that fine-tuning H+-PPases activity and cellular distribution will facilitate rationale strategies for further genetic improvements in crop productivity. PMID:28955362

Predictable and efficient carbon sequestration in the North Pacific Ocean supported by symbiotic nitrogen fixation

PubMed Central

Karl, David M.; Church, Matthew J.; Dore, John E.; Letelier, Ricardo M.; Mahaffey, Claire

2012-01-01

The atmospheric and deep sea reservoirs of carbon dioxide are linked via physical, chemical, and biological processes. The last of these include photosynthesis, particle settling, and organic matter remineralization, and are collectively termed the “biological carbon pump.” Herein, we present results from a 13-y (1992–2004) sediment trap experiment conducted in the permanently oligotrophic North Pacific Subtropical Gyre that document a large, rapid, and predictable summertime (July 15–August 15) pulse in particulate matter export to the deep sea (4,000 m). Peak daily fluxes of particulate matter during the summer export pulse (SEP) average 408, 283, 24.1, 1.1, and 67.5 μmol·m−2·d−1 for total carbon, organic carbon, nitrogen, phosphorus (PP), and biogenic silica, respectively. The SEP is approximately threefold greater than mean wintertime particle fluxes and fuels more efficient carbon sequestration because of low remineralization during downward transit that leads to elevated total carbon/PP and organic carbon/PP particle stoichiometry (371:1 and 250:1, respectively). Our long-term observations suggest that seasonal changes in the microbial assemblage, namely, summertime increases in the biomass and productivity of symbiotic nitrogen-fixing cyanobacteria in association with diatoms, are the main cause of the prominent SEP. The recurrent SEP is enigmatic because it is focused in time despite the absence of any obvious predictable stimulus or habitat condition. We hypothesize that changes in day length (photoperiodism) may be an important environmental cue to initiate aggregation and subsequent export of organic matter to the deep sea. PMID:22308450

Predictable and efficient carbon sequestration in the North Pacific Ocean supported by symbiotic nitrogen fixation.

PubMed

Karl, David M; Church, Matthew J; Dore, John E; Letelier, Ricardo M; Mahaffey, Claire

2012-02-07

The atmospheric and deep sea reservoirs of carbon dioxide are linked via physical, chemical, and biological processes. The last of these include photosynthesis, particle settling, and organic matter remineralization, and are collectively termed the "biological carbon pump." Herein, we present results from a 13-y (1992-2004) sediment trap experiment conducted in the permanently oligotrophic North Pacific Subtropical Gyre that document a large, rapid, and predictable summertime (July 15-August 15) pulse in particulate matter export to the deep sea (4,000 m). Peak daily fluxes of particulate matter during the summer export pulse (SEP) average 408, 283, 24.1, 1.1, and 67.5 μmol·m(-2)·d(-1) for total carbon, organic carbon, nitrogen, phosphorus (PP), and biogenic silica, respectively. The SEP is approximately threefold greater than mean wintertime particle fluxes and fuels more efficient carbon sequestration because of low remineralization during downward transit that leads to elevated total carbon/PP and organic carbon/PP particle stoichiometry (371:1 and 250:1, respectively). Our long-term observations suggest that seasonal changes in the microbial assemblage, namely, summertime increases in the biomass and productivity of symbiotic nitrogen-fixing cyanobacteria in association with diatoms, are the main cause of the prominent SEP. The recurrent SEP is enigmatic because it is focused in time despite the absence of any obvious predictable stimulus or habitat condition. We hypothesize that changes in day length (photoperiodism) may be an important environmental cue to initiate aggregation and subsequent export of organic matter to the deep sea.

Contribution of Efflux Pumps, Porins, and β-Lactamases to Multidrug Resistance in Clinical Isolates of Acinetobacter baumannii

PubMed Central

Rumbo, C.; Gato, E.; López, M.; Ruiz de Alegría, C.; Fernández-Cuenca, F.; Martínez-Martínez, L.; Vila, J.; Pachón, J.; Cisneros, J. M.; Rodríguez-Baño, J.; Pascual, A.

2013-01-01

We investigated the mechanisms of resistance to carbapenems, aminoglycosides, glycylcyclines, tetracyclines, and quinolones in 90 multiresistant clinical strains of Acinetobacter baumannii isolated from two genetically unrelated A. baumannii clones: clone PFGE-ROC-1 (53 strains producing the OXA-58 β-lactamase enzyme and 18 strains with the OXA-24 β-lactamase) and clone PFGE-HUI-1 (19 strains susceptible to carbapenems). We used real-time reverse transcriptase PCR to correlate antimicrobial resistance (MICs) with expression of genes encoding chromosomal β-lactamases (AmpC and OXA-51), porins (OmpA, CarO, Omp33, Dcap-like, OprB, Omp25, OprC, OprD, and OmpW), and proteins integral to six efflux systems (AdeABC, AdeIJK, AdeFGH, CraA, AbeM, and AmvA). Overexpression of the AdeABC system (level of expression relative to that by A. baumannii ATCC 17978, 30- to 45-fold) was significantly associated with resistance to tigecycline, minocycline, and gentamicin and other biological functions. However, hyperexpression of the AdeIJK efflux pump (level of expression relative to that by A. baumannii ATCC 17978, 8- to 10-fold) was significantly associated only with resistance to tigecycline and minocycline (to which the TetB efflux system also contributed). TetB and TetA(39) efflux pumps were detected in clinical strains and were associated with resistance to tetracyclines and doxycycline. The absence of the AdeABC system and the lack of expression of other mechanisms suggest that tigecycline-resistant strains of the PFGE-HUI-1 clone may be associated with a novel resistance-nodulation-cell efflux pump (decreased MICs in the presence of the inhibitor Phe-Arg β-naphthylamide dihydrochloride) and the TetA(39) system. PMID:23939894

Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element.

PubMed

Lim, Chun Shen; Brown, Chris M

2016-09-01

Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel.

Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element

PubMed Central

Lim, Chun Shen; Brown, Chris M.

2016-01-01

ABSTRACT Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel. PMID:27031749

Regulation of thermogenesis in plants: the interaction of alternative oxidase and plant uncoupling mitochondrial protein.

PubMed

Zhu, Yan; Lu, Jianfei; Wang, Jing; Chen, Fu; Leng, Feifan; Li, Hongyu

2011-01-01

Thermogenesis is a process of heat production in living organisms. It is rare in plants, but it does occur in some species of angiosperm. The heat is generated via plant mitochondrial respiration. As possible involvement in thermogenesis of mitochondrial factors, alternative oxidases (AOXs) and plant uncoupling mitochondrial proteins (PUMPs) have been well studied. AOXs and PUMPs are ubiquitously present in the inner membrane of plant mitochondria. They serve as two major energy dissipation systems that balance mitochondrial respiration and uncoupled phosphorylation by dissipating the H+ redox energy and proton electrochemical gradient (ΔμH+) as heat, respectively. AOXs and PUMPs exert similar physiological functions during homeothermic heat production in thermogenic plants. AOXs have five isoforms, while PUMPs have six. Both AOXs and PUMPs are encoded by small nuclear multigene families. Multiple isoforms are expressed in different tissues or organs. Extensive studies have been done in the area of thermogenesis in higher plants. In this review, we focus on the involvement and regulation of AOXs and PUMPs in thermogenesis.

Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae.

PubMed

Chardwiriyapreecha, Soracom; Manabe, Kunio; Iwaki, Tomoko; Kawano-Kawada, Miyuki; Sekito, Takayuki; Lunprom, Siriporn; Akiyama, Koichi; Takegawa, Kaoru; Kakinuma, Yoshimi

2015-01-01

In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles.

HIV-1 Nef-associated Factor 1 Enhances Viral Production by Interacting with CRM1 to Promote Nuclear Export of Unspliced HIV-1 gag mRNA.

PubMed

Ren, Xiao-Xin; Wang, Hai-Bo; Li, Chuan; Jiang, Jin-Feng; Xiong, Si-Dong; Jin, Xia; Wu, Li; Wang, Jian-Hua

2016-02-26

HIV-1 depends on host-cell-encoded factors to complete its life cycle. A comprehensive understanding of how HIV-1 manipulates host machineries during viral infection can facilitate the identification of host targets for antiviral drugs or gene therapy. The cellular protein Naf1 (HIV-1 Nef-associated factor 1) is a CRM1-dependent nucleo-cytoplasmic shuttling protein, and has been identified to regulate multiple receptor-mediated signal pathways in inflammation. The cytoplasm-located Naf1 can inhibit NF-κB activation through binding to A20, and the loss of Naf1 controlled NF-κB activation is associated with multiple autoimmune diseases. However, the effect of Naf1 on HIV-1 mRNA expression has not been characterized. In this study we found that the nucleus-located Naf1 could promote nuclear export of unspliced HIV-1 gag mRNA. We demonstrated that the association between Naf1 and CRM1 was required for this function as the inhibition or knockdown of CRM1 expression significantly impaired Naf1-promoted HIV-1 production. The mutation of Naf1 nuclear export signals (NESs) that account for CRM1 recruitment for nuclear export decreased Naf1 function. Additionally, the mutation of the nuclear localization signal (NLS) of Naf1 diminished its ability to promote HIV-1 production, demonstrating that the shuttling property of Naf1 is required for this function. Our results reveal a novel role of Naf1 in enhancing HIV-1 production, and provide a potential therapeutic target for controlling HIV-1 infection. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Characterization of glycosylphosphatidylinositol-anchored lipid transfer protein 2 (LTPG2) and overlapping function between LTPG/LTPG1 and LTPG2 in cuticular wax export or accumulation in Arabidopsis thaliana.

PubMed

Kim, Hyojin; Lee, Saet Buyl; Kim, Hae Jin; Min, Myung Ki; Hwang, Inhwan; Suh, Mi Chung

2012-08-01

Cuticular waxes are synthesized by the extensive export of intracellular lipids from epidermal cells. However, it is still not known how hydrophobic cuticular lipids are exported to the plant surface through the hydrophilic cell wall. The LTPG2 gene was isolated based on Arabidopsis microarray analysis; this gene is predominantly expressed in stem epidermal peels as compared with in stems. The expression of LTPG2 transcripts was observed in various organs, including stem epidermis and silique walls. The composition of the cuticular wax was significantly altered in the stems and siliques of the ltpg2 mutant and ltpg1 ltpg2 double mutant. In particular, the reduced level of the C29 alkane, which is the major component of cuticular waxes in ltpg1 ltpg2 stems and siliques, was similar to the sum of reduced values of either parent. The total cuticular wax load was reduced by approximately 13% and 20% in both ltpg2 and ltpg1 ltpg2 siliques, respectively, and by approximately 14% in ltpg1 ltpg2 stems when compared with the wild-type. Similarly, severe alterations in the cuticular layer structure of epidermal cells of ltpg2 and ltpg1 ltpg2 stems and silique walls were observed. In tobacco epidermal cells, intracellular trafficking of the fluorescent LTPG/LTPG1 and LTPG2 to the plasma membrane was prevented by a dominant-negative mutant form of ADP-ribosylation factor 1, ARF1(T31N). Taken together, these results indicate that LTPG2 is functionally overlapped with LTPG/LTPG1 during cuticular wax export or accumulation and LTPG/LTPG1 and LTPG2 are targeted to the plasma membrane via the vesicular trafficking system.

Data Sufficiency Assessment and Pumping Test Design for Groundwater Prediction Using Decision Theory and Genetic Algorithms

NASA Astrophysics Data System (ADS)

McPhee, J.; William, Y. W.

2005-12-01

This work presents a methodology for pumping test design based on the reliability requirements of a groundwater model. Reliability requirements take into consideration the application of the model results in groundwater management, expressed in this case as a multiobjective management model. The pumping test design is formulated as a mixed-integer nonlinear programming (MINLP) problem and solved using a combination of genetic algorithm (GA) and gradient-based optimization. Bayesian decision theory provides a formal framework for assessing the influence of parameter uncertainty over the reliability of the proposed pumping test. The proposed methodology is useful for selecting a robust design that will outperform all other candidate designs under most potential 'true' states of the system

Understanding the recent changes in the Southern Ocean carbon cycle: A multidisciplinary approach

NASA Astrophysics Data System (ADS)

Manizza, M.; Kahru, M.; Menemenlis, D.; Nevison, C. D.; Mitchell, B. G.; Keeling, R. F.

2016-12-01

The Southern Ocean represents a key area of the global ocean for the uptake of the CO2 originating from fossil fuels emissions. In these waters, cold temperatures combined with high rates of biological production drive the carbon uptake that accounts for about one-third of the global ocean uptake.Recent studies showed that changes in the Southern Annular Mode (SAM) index, mainly a proxy of the intensity of westerly winds, had a significant impact on the temporal variability of the CO2 uptake in the Southern Ocean. In order to shed light on this problem we propose to use both satellite-derived estimates of ocean productivity and carbon export in combinations of ocean physical and biogeochemical state estimates focusing on the 2006-2013 period. While the estimates of carbon fixation and export based on remote sensing will provide key information on the spatial and temporal variations of the biological carbon pump, the ocean state estimates will provide additional information on physical and carbon cycle processes, including the air-sea CO2 fluxes of the Southern Ocean in the 2006-2013 period where model solutions have been optimized.These physical estimates will be used to force an ocean biogeochemical model (ECCO2-Darwin) that will compute the CO2 uptake for each year. The physical model, forced with optimized atmospheric forcing, aims to realistically simulate interannual ocean climate variability that drives changes in both physical and biogeochemical processes ultimately impacting the carbon uptake of the Southern Ocean, and potentially responding to the SAM index variations.Although in this study great emphasis is given to the role of physical climate variations at driving the CO2 uptake of these polar waters, we will integrate model results with estimates from remote sensing techniques to better understand role of the biological carbon pump and its variability potentially responding to the SAM index changes.

Regulation of Plasma Membrane Localization of the Na+-Taurocholate Cotransporting Polypeptide (Ntcp) by Hyperosmolarity and Tauroursodeoxycholate*

PubMed Central

Sommerfeld, Annika; Mayer, Patrick G. K.; Cantore, Miriam; Häussinger, Dieter

2015-01-01

In perfused rat liver, hepatocyte shrinkage induces a Fyn-dependent retrieval of the bile salt export pump (Bsep) and multidrug resistance-associated protein 2 (Mrp2) from the canalicular membrane (Cantore, M., Reinehr, R., Sommerfeld, A., Becker, M., and Häussinger, D. (2011) J. Biol. Chem. 286, 45014–45029) leading to cholestasis. However little is known about the effects of hyperosmolarity on short term regulation of the Na+-taurocholate cotransporting polypeptide (Ntcp), the major bile salt uptake system at the sinusoidal membrane of hepatocytes. The aim of this study was to analyze hyperosmotic Ntcp regulation and the underlying signaling events. Hyperosmolarity induced a significant retrieval of Ntcp from the basolateral membrane, which was accompanied by an activating phosphorylation of the Src kinases Fyn and Yes but not of c-Src. Hyperosmotic internalization of Ntcp was sensitive to SU6656 and PP-2, suggesting that Fyn mediates Ntcp retrieval from the basolateral membrane. Hyperosmotic internalization of Ntcp was also found in livers from wild-type mice but not in p47phox knock-out mice. Tauroursodeoxycholate (TUDC) and cAMP reversed hyperosmolarity-induced Fyn activation and triggered re-insertion of the hyperosmotically retrieved Ntcp into the membrane. This was associated with dephosphorylation of the Ntcp on serine residues. Insertion of Ntcp by TUDC was sensitive to the integrin inhibitory hexapeptide GRGDSP and inhibition of protein kinase A. TUDC also reversed the hyperosmolarity-induced retrieval of bile salt export pump from the canalicular membrane. These findings suggest a coordinated and oxidative stress- and Fyn-dependent retrieval of sinusoidal and canalicular bile salt transport systems from the corresponding membranes. Ntcp insertion was also identified as a novel target of β1-integrin-dependent TUDC action, which is frequently used in the treatment of cholestatic liver disease. PMID:26306036

Regulation of plasma membrane localization of the Na+-taurocholate cotransporting polypeptide (Ntcp) by hyperosmolarity and tauroursodeoxycholate.

PubMed

Sommerfeld, Annika; Mayer, Patrick G K; Cantore, Miriam; Häussinger, Dieter

2015-10-02

In perfused rat liver, hepatocyte shrinkage induces a Fyn-dependent retrieval of the bile salt export pump (Bsep) and multidrug resistance-associated protein 2 (Mrp2) from the canalicular membrane (Cantore, M., Reinehr, R., Sommerfeld, A., Becker, M., and Häussinger, D. (2011) J. Biol. Chem. 286, 45014-45029) leading to cholestasis. However little is known about the effects of hyperosmolarity on short term regulation of the Na(+)-taurocholate cotransporting polypeptide (Ntcp), the major bile salt uptake system at the sinusoidal membrane of hepatocytes. The aim of this study was to analyze hyperosmotic Ntcp regulation and the underlying signaling events. Hyperosmolarity induced a significant retrieval of Ntcp from the basolateral membrane, which was accompanied by an activating phosphorylation of the Src kinases Fyn and Yes but not of c-Src. Hyperosmotic internalization of Ntcp was sensitive to SU6656 and PP-2, suggesting that Fyn mediates Ntcp retrieval from the basolateral membrane. Hyperosmotic internalization of Ntcp was also found in livers from wild-type mice but not in p47(phox) knock-out mice. Tauroursodeoxycholate (TUDC) and cAMP reversed hyperosmolarity-induced Fyn activation and triggered re-insertion of the hyperosmotically retrieved Ntcp into the membrane. This was associated with dephosphorylation of the Ntcp on serine residues. Insertion of Ntcp by TUDC was sensitive to the integrin inhibitory hexapeptide GRGDSP and inhibition of protein kinase A. TUDC also reversed the hyperosmolarity-induced retrieval of bile salt export pump from the canalicular membrane. These findings suggest a coordinated and oxidative stress- and Fyn-dependent retrieval of sinusoidal and canalicular bile salt transport systems from the corresponding membranes. Ntcp insertion was also identified as a novel target of β1-integrin-dependent TUDC action, which is frequently used in the treatment of cholestatic liver disease. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Energy landscape of the reactions governing the Na+ deeply occluded state of the Na+/K+-ATPase in the giant axon of the Humboldt squid

PubMed Central

Castillo, Juan P.; De Giorgis, Daniela; Basilio, Daniel; Gadsby, David C.; Rosenthal, Joshua J. C.; Latorre, Ramon; Holmgren, Miguel; Bezanilla, Francisco

2011-01-01

The Na+/K+ pump is a nearly ubiquitous membrane protein in animal cells that uses the free energy of ATP hydrolysis to alternatively export 3Na+ from the cell and import 2K+ per cycle. This exchange of ions produces a steady-state outwardly directed current, which is proportional in magnitude to the turnover rate. Under certain ionic conditions, a sudden voltage jump generates temporally distinct transient currents mediated by the Na+/K+ pump that represent the kinetics of extracellular Na+ binding/release and Na+ occlusion/deocclusion transitions. For many years, these events have escaped a proper thermodynamic treatment due to the relatively small electrical signal. Here, taking the advantages offered by the large diameter of the axons from the squid Dosidicus gigas, we have been able to separate the kinetic components of the transient currents in an extended temperature range and thus characterize the energetic landscape of the pump cycle and those transitions associated with the extracellular release of the first Na+ from the deeply occluded state. Occlusion/deocclusion transition involves large changes in enthalpy and entropy as the ion is exposed to the external milieu for release. Binding/unbinding is substantially less costly, yet larger than predicted for the energetic cost of an ion diffusing through a permeation pathway, which suggests that ion binding/unbinding must involve amino acid side-chain rearrangements at the site. PMID:22143771

Energy landscape of the reactions governing the Na+ deeply occluded state of the Na+/K+-ATPase in the giant axon of the Humboldt squid.

PubMed

Castillo, Juan P; De Giorgis, Daniela; Basilio, Daniel; Gadsby, David C; Rosenthal, Joshua J C; Latorre, Ramon; Holmgren, Miguel; Bezanilla, Francisco

2011-12-20

The Na(+)/K(+) pump is a nearly ubiquitous membrane protein in animal cells that uses the free energy of ATP hydrolysis to alternatively export 3Na(+) from the cell and import 2K(+) per cycle. This exchange of ions produces a steady-state outwardly directed current, which is proportional in magnitude to the turnover rate. Under certain ionic conditions, a sudden voltage jump generates temporally distinct transient currents mediated by the Na(+)/K(+) pump that represent the kinetics of extracellular Na(+) binding/release and Na(+) occlusion/deocclusion transitions. For many years, these events have escaped a proper thermodynamic treatment due to the relatively small electrical signal. Here, taking the advantages offered by the large diameter of the axons from the squid Dosidicus gigas, we have been able to separate the kinetic components of the transient currents in an extended temperature range and thus characterize the energetic landscape of the pump cycle and those transitions associated with the extracellular release of the first Na(+) from the deeply occluded state. Occlusion/deocclusion transition involves large changes in enthalpy and entropy as the ion is exposed to the external milieu for release. Binding/unbinding is substantially less costly, yet larger than predicted for the energetic cost of an ion diffusing through a permeation pathway, which suggests that ion binding/unbinding must involve amino acid side-chain rearrangements at the site.

Identification of Novel Saccharomyces cerevisiae Proteins with Nuclear Export Activity: Cell Cycle-Regulated Transcription Factor Ace2p Shows Cell Cycle-Independent Nucleocytoplasmic Shuttling

PubMed Central

Jensen, Torben Heick; Neville, Megan; Rain, Jean Christophe; McCarthy, Terri; Legrain, Pierre; Rosbash, Michael

2000-01-01

Nuclear export of proteins containing leucine-rich nuclear export signals (NESs) is mediated by the NES receptor CRM1/Crm1p. We have carried out a yeast two-hybrid screen with Crm1p as a bait. The Crm1p-interacting clones were subscreened for nuclear export activity in a visual assay utilizing the Crm1p-inhibitor leptomycin B (LMB). This approach identified three Saccharomyces cerevisiae proteins not previously known to have nuclear export activity. These proteins are the 5′ RNA triphosphatase Ctl1p, the cell cycle-regulated transcription factor Ace2p, and a protein encoded by the previously uncharacterized open reading frame YDR499W. Mutagenesis analysis show that YDR499Wp contains an NES that conforms to the consensus sequence for leucine-rich NESs. Mutagenesis of Ctl1p and Ace2p were unable to identify specific NES residues. However, a 29-amino-acid region of Ace2p, rich in hydrophobic residues, contains nuclear export activity. Ace2p accumulates in the nucleus at the end of mitosis and activates early-G1-specific genes. We now provide evidence that Ace2p is nuclear not only in late M-early G1 but also during other stages of the cell cycle. This feature of Ace2p localization explains its ability to activate genes such as CUP1, which are not expressed in a cell cycle-dependent manner. PMID:11027275

PAUSED encodes the Arabidopsis exportin-t ortholog.

PubMed

Hunter, Christine A; Aukerman, Milo J; Sun, Hui; Fokina, Maria; Poethig, R Scott

2003-08-01

Los1p/exportin-t (XPOT) mediates the nuclear export of tRNAs in yeast and mammals. The requirements for this transport pathway are unclear, however, because los1 mutations do not affect yeast growth, and the phenotype of XPOT mutations in mammals is unknown. Here, we show that PAUSED (PSD) is the Arabidopsis ortholog of LOS1/XPOT and is capable of rescuing the tRNA export defect of los1 in Brewer's yeast (Saccharomyces cerevisiae), suggesting that its function has been conserved. Putative null alleles of PSD disrupt the initiation of the shoot apical meristem and delay leaf initiation after germination, the emergence of the radicle and lateral roots, and the transition to flowering. Plants doubly mutant for psd and hasty, the Arabidopsis ortholog of exportin 5, are viable but have a more severe phenotype than either single mutant. These results suggest that PSD plays a role in tRNA export in Arabidopsis, but that at least one-and perhaps several-additional tRNA export pathways also exist. The PSD transcript is broadly expressed during development and is alternatively spliced in the 3'-untranslated region. No temporal or spatial difference in the abundance of different splice forms was observed. We propose that the mutant phenotype of psd reflects defects in developmental events and cell/tissue types that require elevated levels of protein synthesis and are therefore acutely sensitive to a reduction in tRNA export.

Regulation of monocarboxylate transporter MCT1 expression by p53 mediates inward and outward lactate fluxes in tumors.

PubMed

Boidot, Romain; Végran, Frédérique; Meulle, Aline; Le Breton, Aude; Dessy, Chantal; Sonveaux, Pierre; Lizard-Nacol, Sarab; Feron, Olivier

2012-02-15

The monocarboxylate transporter (MCT) family member MCT1 can transport lactate into and out of tumor cells. Whereas most oxidative cancer cells import lactate through MCT1 to fuel mitochondrial respiration, the role of MCT1 in glycolysis-derived lactate efflux remains less clear. In this study, we identified a direct link between p53 function and MCT1 expression. Under hypoxic conditions, p53 loss promoted MCT1 expression and lactate export produced by elevated glycolytic flux, both in vitro and in vivo. p53 interacted directly with the MCT1 gene promoter and altered MCT1 mRNA stabilization. In hypoxic p53(-/-) tumor cells, NF-κB further supported expression of MCT1 to elevate its levels. Following glucose deprivation, upregulated MCT1 in p53(-/-) cells promoted lactate import and favored cell proliferation by fuelling mitochondrial respiration. We also found that MCT1 expression was increased in human breast tumors harboring p53 mutations and coincident features of hypoxia, with higher MCT1 levels associated with poorer clinical outcomes. Together, our findings identify MCT1 as a target for p53 repression and they suggest that MCT1 elevation in p53-deficient tumors allows them to adapt to metabolic needs by facilitating lactate export or import depending on the glucose availability.

Optical silencing of body wall muscles induces pumping inhibition in Caenorhabditis elegans

PubMed Central

Takahashi, Megumi

2017-01-01

Feeding, a vital behavior in animals, is modulated depending on internal and external factors. In the nematode Caenorhabditis elegans, the feeding organ called the pharynx ingests food by pumping driven by the pharyngeal muscles. Here we report that optical silencing of the body wall muscles, which drive the locomotory movement of worms, affects pumping. In worms expressing the Arch proton pump or the ACR2 anion channel in the body wall muscle cells, the pumping rate decreases after activation of Arch or ACR2 with light illumination, and recovers gradually after terminating illumination. Pumping was similarly inhibited by illumination in locomotion-defective mutants carrying Arch, suggesting that perturbation of locomotory movement is not critical for pumping inhibition. Analysis of mutants and cell ablation experiments showed that the signals mediating the pumping inhibition response triggered by activation of Arch with weak light are transferred mainly through two pathways: one involving gap junction-dependent mechanisms through pharyngeal I1 neurons, which mediate fast signals, and the other involving dense-core vesicle-dependent mechanisms, which mediate slow signals. Activation of Arch with strong light inhibited pumping strongly in a manner that does not rely on either gap junction-dependent or dense-core vesicle-dependent mechanisms. Our study revealed a new aspect of the neural and neuroendocrine controls of pumping initiated from the body wall muscles. PMID:29281635

Empirical evaluation of pump inlet compliance

NASA Technical Reports Server (NTRS)

Ghahremani, F. G.; Rubin, S.

1972-01-01

Cavitation compliance was determined experimentally from pulsing tests on a number of rocket turbopumps. The primary test data used for this study are those for the Rocketdyne H-1, F-1, and J-2 oxidizer and fuel pumps employed on Saturn vehicles. The study shows that these data can be correlated by a particular form of nondimensionalization, the key feature of which is to divide the operating cavitation number or suction specific speed by its value at head breakdown. An expression is obtained for a best-fit curve for these data. Another set of test data for the Aerojet LR87 and 91 pumps can be correlated by a somewhat different nondimensional pump performance parameter, specifically by relating the cavitation number to its position between the head breakdown point and the point of zero slope of the head coefficient versus cavitation number. Recommendations are given for the estimation of the cavitation compliance for new designs in the Rocketdyne family of pumps.

Genetic characterization of tigecycline resistance in clinical isolates of Enterobacter cloacae and Enterobacter aerogenes.

PubMed

Veleba, Mark; De Majumdar, Shyamasree; Hornsey, Michael; Woodford, Neil; Schneiders, Thamarai

2013-05-01

The intrinsically encoded ramA gene has been linked to tigecycline resistance through the up-regulation of efflux pump AcrAB in Enterobacter cloacae. The molecular basis for increased ramA expression in E. cloacae and Enterobacter aerogenes, as well as the role of AraC regulator rarA, has not yet been shown. To ascertain the intrinsic molecular mechanism(s) involved in tigecycline resistance in Enterobacter spp., we analysed the expression levels of ramA and rarA and corresponding efflux pump genes acrAB and oqxAB in Enterobacter spp. clinical isolates. The expression levels of ramA, rarA, oqxA and acrA were tested by quantitative real-time RT-PCR. The ramR open reading frames of the ramA-overexpressing strains were sequenced; strains harbouring mutations were transformed with wild-type ramR to study altered ramA expression and tigecycline susceptibility. Tigecycline resistance was mediated primarily by increased ramA expression in E. cloacae and E. aerogenes. Only the ramA-overexpressing E. cloacae isolates showed increased rarA and oqxA expression. Upon complementation with wild-type ramR, all Enterobacter spp. containing ramR mutations exhibited decreased ramA and acrA expression and increased tigecycline susceptibility. Exceptions were one E. cloacae strain and one E. aerogenes strain, where a decrease in ramA levels was not accompanied by lower acrA expression. Increased ramA expression due to ramR deregulation is the primary mediator of tigecycline resistance in clinical isolates of E. cloacae and E. aerogenes. However, some ramA-overexpressing isolates do not show changes in ramR, suggesting alternate pathways of ramA regulation; the rarA regulator and the oqxAB efflux pump may also play a role in tigecycline resistance in E. cloacae.

Distinct temporal changes in host cell lncRNA expression during the course of an adenovirus infection

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

Zhao, Hongxing, E-mail: Hongxing.Zhao@igp.uu.se; Chen, Maoshan; Lind, Sara Bergström

The deregulation of cellular long non-coding RNA (lncRNA) expression during a human adenovirus infection was studied by deep sequencing. Expression of lncRNAs increased substantially following the progression of the infection. Among 645 significantly expressed lncRNAs, the expression of 398 was changed more than 2-fold. More than 80% of them were up-regulated and 80% of them were detected during the late phase. Based on the genomic locations of the deregulated lncRNAs in relation to known mRNAs and miRNAs, they were predicted to be involved in growth, structure, apoptosis and wound healing in the early phase, cell proliferation in the intermediate phasemore » and protein synthesis, modification and transport in the late phase. The most significant functions of cellular RNA-binding proteins, previously shown to interact with the deregulated lncRNAs identified here, are involved in RNA splicing, nuclear export and translation events. We hypothesize that adenoviruses exploit the lncRNA network to optimize their reproduction. - Highlights: • The expression of 398 lncRNAs showed a distinct temporal pattern during Ad2 infection. • 80% of the deregulated lncRNAs were up-regulated during the late phase of infection. • The deregulated lncRNAs potentiallyinteract with 33 cellular RNA binding proteins. • These RBPs are involved in RNA splicing, nuclear export and translation. • Adenovirus exploits the cellular lncRNA network to optimize its replication.« less

Translations on Western Europe. No. 1029

DTIC Science & Technology

1977-03-03

HELLENEWS-EXPRESS, 3 Feb 77) 41 Greek-Syrian Ferry Link Agreed (HELLENEWS-EXPRESS, 27 Jan 77) 43 Knitwear Exporters Charge French Obstruction...In addition the preparatory technology for extracting the ore grains from the mud (for which a new process of "finest grain flotation" was developed... fully curbing its prices: it has an infla- tion rate of about 3 percent. Nevertheless, Belgium did make some gains over last year when the

CD30 downregulation, MMAE resistance, and MDR1 upregulation are all associated with resistance to brentuximab vedotin

PubMed Central

Chen, Robert; Hou, Jessie; Newman, Edward; Kim, Young; Donohue, Cecile; Liu, Xueli; Thomas, Sandra H.; Forman, Stephen J.; Kane, Susan E.

2015-01-01

Brentuximab vedotin (BV) is an antibody-drug conjugate that specifically delivers the potent cytotoxic drug MMAE to CD30-positive cells. BV is FDA-approved for treatment of relapsed/refractory Hodgkin lymphoma (HL) and anaplastic large cell lymphoma (ALCL); however, many patients do not achieve complete remission and develop BV resistant disease. We selected for BV-resistant HL (L428) and ALCL (Karpas-299) cell lines using either constant (ALCL) or pulsatile (HL) exposure to BV. We confirmed drug resistance by MTS assay, and analyzed CD30 expression in resistant cells by flow cytometry, qRT-PCR, and Western blotting. We also measured drug exporter expression, MMAE resistance, and intracellular MMAE concentrations in BV-resistant cells. Additionally, tissue biopsy samples from 10 HL and 5 ALCL patients who had relapsed or progressed after BV treatment were analyzed by immunohistocytochemistry for CD30 expression. The resistant ALCL cell line, but not the HL cell line, demonstrated downregulated CD30 expression compared to the parental cell line. In contrast, the HL cell line, but not the ALCL cell line, exhibited MMAE resistance and increased expression of the MDR1 drug exporter compared to the parental line. For both HL and ALCL, samples from patients relapsed/resistant on BV persistently expressed CD30 by immunohistocytochemistry. One HL patient sample expressed MDR1 by immunohistocytochemistry. Although loss of CD30 expression is a possible mode of BV resistance in ALCL in vitro models, this has not been confirmed in patients. MMAE resistance and MDR1 expression are possible modes of BV resistance for HL both in vitro and in patients. PMID:25840583

Autophagy requires poly(adp-ribosyl)ation-dependent AMPK nuclear export

PubMed Central

Rodríguez-Vargas, José M; Rodríguez, María I; Majuelos-Melguizo, Jara; García-Diaz, Ángel; González-Flores, Ariannys; López-Rivas, Abelardo; Virág, László; Illuzzi, Giuditta; Schreiber, Valerie; Dantzer, Françoise; Oliver, F Javier

2016-01-01

AMPK is a central energy sensor linking extracellular milieu fluctuations with the autophagic machinery. In the current study we uncover that Poly(ADP-ribosyl)ation (PARylation), a post-translational modification (PTM) of proteins, accounts for the spatial and temporal regulation of autophagy by modulating AMPK subcellular localisation and activation. More particularly, we show that the minority AMPK pool needs to be exported to the cytosol in a PARylation-dependent manner for optimal induction of autophagy, including ULK1 phosphorylation and mTORC1 inactivation. PARP-1 forms a molecular complex with AMPK in the nucleus in non-starved cells. In response to nutrient deprivation, PARP-1 catalysed PARylation, induced the dissociation of the PARP-1/AMPK complex and the export of free PARylated nuclear AMPK to the cytoplasm to activate autophagy. PARP inhibition, its silencing or the expression of PARylation-deficient AMPK mutants prevented not only the AMPK nuclear-cytosolic export but also affected the activation of the cytosolic AMPK pool and autophagosome formation. These results demonstrate that PARylation of AMPK is a key early signal to efficiently convey extracellular nutrient perturbations with downstream events needed for the cell to optimize autophagic commitment before autophagosome formation. PMID:27689873

CopZ from Bacillus subtilis interacts in vivo with a copper exporting CPx-type ATPase CopA.

PubMed

Radford, David S; Kihlken, Margaret A; Borrelly, Gilles P M; Harwood, Colin R; Le Brun, Nick E; Cavet, Jennifer S

2003-03-14

The structure of the hypothetical copper-metallochaperone CopZ from Bacillus subtilis and its predicted partner CopA have been studied but their respective contributions to copper export, -import, -sequestration and -supply are unknown. DeltacopA was hypersensitive to copper and contained more copper atoms cell(-1) than wild-type. Expression from the copA operator-promoter increased in elevated copper (not other metals), consistent with a role in copper export. A bacterial two-hybrid assay revealed in vivo interaction between CopZ and the N-terminal domain of CopA but not that of a related transporter, YvgW, involved in cadmium-resistance. Activity of copper-requiring cytochrome caa(3) oxidase was retained in deltacopZ and deltacopA. DeltacopZ was only slightly copper-hypersensitive but deltacopZ/deltacopA was more sensitive than deltacopA, implying some action of CopZ that is independent of CopA. Significantly, deltacopZ contained fewer copper atoms cell(-1) than wild-type under these conditions. CopZ makes a net contribution to copper sequestration and/or recycling exceeding any donation to CopA for export.

RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs.

PubMed

Huang, Feng; Zhang, Junsong; Zhang, Yijun; Geng, Guannan; Liang, Juanran; Li, Yingniang; Chen, Jingliang; Liu, Chao; Zhang, Hui

2015-12-01

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-box of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. Copyright © 2015 Elsevier Inc. All rights reserved.

Gastrin Induces Nuclear Export and Proteasome Degradation of Menin in Enteric Glial Cells.

PubMed

Sundaresan, Sinju; Meininger, Cameron A; Kang, Anthony J; Photenhauer, Amanda L; Hayes, Michael M; Sahoo, Nirakar; Grembecka, Jolanta; Cierpicki, Tomasz; Ding, Lin; Giordano, Thomas J; Else, Tobias; Madrigal, David J; Low, Malcolm J; Campbell, Fiona; Baker, Ann-Marie; Xu, Haoxing; Wright, Nicholas A; Merchant, Juanita L

2017-12-01

The multiple endocrine neoplasia, type 1 (MEN1) locus encodes the nuclear protein and tumor suppressor menin. MEN1 mutations frequently cause neuroendocrine tumors such as gastrinomas, characterized by their predominant duodenal location and local metastasis at time of diagnosis. Diffuse gastrin cell hyperplasia precedes the appearance of MEN1 gastrinomas, which develop within submucosal Brunner's glands. We investigated how menin regulates expression of the gastrin gene and induces generation of submucosal gastrin-expressing cell hyperplasia. Primary enteric glial cultures were generated from the VillinCre:Men1 FL/FL :Sst -/- mice or C57BL/6 mice (controls), with or without inhibition of gastric acid by omeprazole. Primary enteric glial cells from C57BL/6 mice were incubated with gastrin and separated into nuclear and cytoplasmic fractions. Cells were incubated with forskolin and H89 to activate or inhibit protein kinase A (a family of enzymes whose activity depends on cellular levels of cyclic AMP). Gastrin was measured in blood, tissue, and cell cultures using an ELISA. Immunoprecipitation with menin or ubiquitin was used to demonstrate post-translational modification of menin. Primary glial cells were incubated with leptomycin b and MG132 to block nuclear export and proteasome activity, respectively. We obtained human duodenal, lymph node, and pancreatic gastrinoma samples, collected from patients who underwent surgery from 1996 through 2007 in the United States or the United Kingdom. Enteric glial cells that stained positive for glial fibrillary acidic protein (GFAP+) expressed gastrin de novo through a mechanism that required PKA. Gastrin-induced nuclear export of menin via cholecystokinin B receptor (CCKBR)-mediated activation of PKA. Once exported from the nucleus, menin was ubiquitinated and degraded by the proteasome. GFAP and other markers of enteric glial cells (eg, p75 and S100B), colocalized with gastrin in human duodenal gastrinomas. MEN1-associated gastrinomas, which develop in the submucosa, might arise from enteric glial cells through hormone-dependent PKA signaling. This pathway disrupts nuclear menin function, leading to hypergastrinemia and associated sequelae. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Differential IFN-gamma stimulation of HLA-A gene expression through CRM-1-dependent nuclear RNA export.

PubMed

Browne, Sarah K; Roesser, James R; Zhu, Sheng Zu; Ginder, Gordon D

2006-12-15

IFNs regulate most MHC class I genes by stimulating transcription initiation. As shown previously, IFN-gamma controls HLA-A expression primarily at the posttranscriptional level. We have defined two 8-base sequences in a 39-nucleotide region in the 3'-transcribed region of the HLA-A gene that are required for the posttranscriptional response to IFN-gamma. Stimulation of HLA-A expression by IFN-gamma requires nuclear export of HLA-A mRNA by chromosome maintenance region 1 (CRM-1). Treatment of cells with leptomycin B, a specific inhibitor of CRM-1, completely inhibited IFN-gamma induction of HLA-A. Expression of a truncated, dominant-negative form of the nucleoporin NUP214/CAN, DeltaCAN, that specifically interacts with CRM-1, also prevented IFN-gamma stimulation of HLA-A, providing confirmation of the role of CRM-1. Increased expression of HLA-A induced by IFN-gamma also requires protein methylation, as shown by the fact that treatment of SK-N-MC cells or HeLa cells with the PRMT1 inhibitor 5'-methyl-5'-thioadenosine abolished the cellular response to IFN-gamma. In contrast with HLA-A, IFN-gamma-induced expression of the HLA class Ib gene, HLA-E, was not affected by either 5'-methyl-5'-thioadenosine or leptomycin B. These results provide proof of principle that it is possible to differentially modulate the IFN-gamma-induced expression of the HLA-E and HLA-A genes, whose products often mediate opposing effects on cellular immunity to tumor cells, pathogens, and autoantigens.

Ferroportin-mediated iron transport: expression and regulation

PubMed Central

Ward, Diane; Kaplan, Jerry

2013-01-01

The distinguishing feature between iron homeostasis in single versus multicellular organisms is the need for multicellular organisms to transfer iron from sites of absorption to sites of utilization and storage. Ferroportin is the only known iron exporter and ferroportin plays an essential role in the export of iron from cells to blood. Ferroportin can be regulated at many different levels including transcriptionally, post-transcriptionally, through mRNA stability and post-translationally, through protein turnover. Additionally, ferroportin may be regulated in both cell-dependent and cell-autonomous fashions. Regulation of ferroportin is critical for iron homeostasis as alterations in ferroportin may result in either iron deficiency or iron overload. PMID:22440327

Pumping Milk Without Ever Feeding at the Breast in the Moms2Moms Study.

PubMed

Keim, Sarah A; Boone, Kelly M; Oza-Frank, Reena; Geraghty, Sheela R

2017-09-01

More than 85% of contemporary lactating women in the United States express their milk at least sometimes. Some produce milk exclusively through pumping. We characterized women who pumped but never fed at the breast and compared their infant feeding practices with those of women who fed at the breast with or without pumping. Study participants were those delivered at Ohio State University Wexner Medical Center in 2011 and completed a questionnaire at 12 months postpartum (n = 478). We used bivariate and multivariate approaches (survival analysis) to compare women who pumped but never fed at the breast with women who fed at the breast with or without pumping. Women (n = 33, 6.9%) who pumped but never fed at the breast comprised a diverse group but were more likely to have delivered preterm and were of lower socioeconomic status on average. They initiated pumping and formula feeding earlier (median = day 1 after delivery) and were more likely to report difficulty making enough milk compared with women who fed at the breast with or without pumping. They had much shorter total duration of milk production (adjusted hazard ratio = 3.3, 95% confidence interval: 2.1, 5.2) after controlling for clinical and sociodemographic confounders. Pumping without feeding at the breast is associated with shorter milk feeding duration and earlier introduction of formula compared with feeding at the breast with or without pumping. Establishing feeding at the breast, rather than exclusive pumping, may be important for achieving human milk feeding goals.

Influenza A Virus NS1 Protein Promotes Efficient Nuclear Export of Unspliced Viral M1 mRNA.

PubMed

Pereira, Carina F; Read, Eliot K C; Wise, Helen M; Amorim, Maria J; Digard, Paul

2017-08-01

Influenza A virus mRNAs are transcribed by the viral RNA-dependent RNA polymerase in the cell nucleus before being exported to the cytoplasm for translation. Segment 7 produces two major transcripts: an unspliced mRNA that encodes the M1 matrix protein and a spliced transcript that encodes the M2 ion channel. Export of both mRNAs is dependent on the cellular NXF1/TAP pathway, but it is unclear how they are recruited to the export machinery or how the intron-containing but unspliced M1 mRNA bypasses the normal quality-control checkpoints. Using fluorescent in situ hybridization to monitor segment 7 mRNA localization, we found that cytoplasmic accumulation of unspliced M1 mRNA was inefficient in the absence of NS1, both in the context of segment 7 RNPs reconstituted by plasmid transfection and in mutant virus-infected cells. This effect was independent of any major effect on steady-state levels of segment 7 mRNA or splicing but corresponded to a ∼5-fold reduction in the accumulation of M1. A similar defect in intronless hemagglutinin (HA) mRNA nuclear export was seen with an NS1 mutant virus. Efficient export of M1 mRNA required both an intact NS1 RNA-binding domain and effector domain. Furthermore, while wild-type NS1 interacted with cellular NXF1 and also increased the interaction of segment 7 mRNA with NXF1, mutant NS1 polypeptides unable to promote mRNA export did neither. Thus, we propose that NS1 facilitates late viral gene expression by acting as an adaptor between viral mRNAs and the cellular nuclear export machinery to promote their nuclear export. IMPORTANCE Influenza A virus is a major pathogen of a wide variety of mammalian and avian species that threatens public health and food security. A fuller understanding of the virus life cycle is important to aid control strategies. The virus has a small genome that encodes relatively few proteins that are often multifunctional. Here, we characterize a new function for the NS1 protein, showing that, as well as previously identified roles in antagonizing the innate immune defenses of the cell and directly upregulating translation of viral mRNAs, it also promotes the nuclear export of the viral late gene mRNAs by acting as an adaptor between the viral mRNAs and the cellular mRNA nuclear export machinery. Copyright © 2017 Pereira et al.

Glucose-dependent growth arrest of leukemia cells by MCT1 inhibition: Feeding Warburg's sweet tooth and blocking acid export as an anticancer strategy.

PubMed

Pivovarova, Aleksandra I; MacGregor, Gordon G

2018-02-01

This study aims to investigate the utilization of The Warburg Effect, cancer's "sweet tooth" and natural greed for glucose to enhance the effect of monocarboxylate transporter inhibition on cellular acidification. By simulating hyperglycemia with high glucose we may increase the effectiveness of inhibition of lactate and proton export on the dysregulation of cell pH homeostasis causing cell death or disruption of growth in cancer cells. MCT1 and MCT4 expression was determined in MCF7 and K562 cell lines using RT-PCR. Cell viability, growth, intracellular pH and cell cycle analysis was measured in the cell lines grown in 5 mM and 25 mM glucose containing media in the presence and absence of the MCT1 inhibitor AR-C155858 (1 μM) and the NHE1 inhibitor cariporide (10 μM). The MCT1 inhibitor, AR-C155858 had minimal effect on the viability, growth and intracellular pH of MCT4 expressing MCF7 cells. AR-C155858 had no effect on the viability of the MCT1 expressing K562 cells, but decreased intracellular pH and cell proliferation, by a glucose-dependent mechanism. Inhibition of NHE1 on its own had a no effect on cell growth, but together with AR-C155858 showed an additive effect on inhibition of cell growth. In cancer cells that only express MCT1, increased glucose concentrations in the presence of an MCT1 inhibitor decreased intracellular pH and reduced cell growth by G1 phase cell-cycle arrest. Thus we propose a transient hyperglycemic-clamp in combination with proton export inhibitors be evaluated as an adjunct to cancer treatment in clinical studies. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Accounting System for Water Use by Vegetation in the Lower Colorado River Valley

USGS Publications Warehouse

Owen-Joyce, Sandra J.

1992-01-01

The Colorado River is the principal source of water in the valley of the Colorado River between Hoover Dam and the international boundary with Mexico (fig. 1). Agricultural, domestic, municipal, industrial, hydroelectric-power genera-tion, and recreation are the primary uses of river water in the valley. Most of the consumptive use of water from the river occurs downstream from Davis Dam, where water is diverted to irrigate crops along the river or is exported to interior regions of California and Arizona. Most of the agricultural areas are on the alluvium of the flood plain; in a few areas, land on the alluvial terraces has been cultivated. River water is consumed mainly by vegetation (crops and phreatophytes) on the flood plain. Crops were grown on 70.3 percent of the vegetated area classified by using 1984 digital image satellite data. Phreatophytes, natural vege-tation that obtain water from the alluvial aquifer, covered the remaining vegetated areas on the uncultivated flood plain. Most of the water used for irrigation is diverted or pumped from the river. In some areas, water is pumped from wells completed in the alluvial aquifer, which is hydraulically connected to the river.

The brain-specific double-stranded RNA-binding protein Staufen2: nucleolar accumulation and isoform-specific exportin-5-dependent export.

PubMed

Macchi, Paolo; Brownawell, Amy M; Grunewald, Barbara; DesGroseillers, Luc; Macara, Ian G; Kiebler, Michael A

2004-07-23

The mammalian double-stranded RNA-binding proteins Staufen (Stau1 and Stau2) are involved in RNA localization in polarized neurons. In contrast to the more ubiquitously expressed Stau1, Stau2 is mainly expressed in the nervous system. In Drosophila, the third double-stranded RNA-binding domain (RBD3) of Staufen is essential for RNA interaction. When conserved amino acids within the RBD3 of Stau2 were mutated to render Stau2 defective for RNA binding, the mutant Stau2 proteins accumulate predominantly in the nucleolus. This is in contrast to wild type Stau2 that mostly localizes in the cytosol. The nuclear import is dependent on a nuclear localization signal in close proximity to the RBD3. The nuclear export of Stau2 is not dependent on CRM1 but rather on Exportin-5. We show that Exportin-5 interacts with the RBD3 of wild type Stau2 in an RNA-dependent manner in vitro but not with mutant Stau2. When Exportin-5 is down-regulated by RNA interference, only the largest isoform of Stau2 (Stau2(62)) preferentially accumulates in the nucleolus. It is tempting to speculate that Stau2(62) binds RNA in the nucleus and assembles into ribonucleoparticles, which are then exported via the Exportin-5 pathway to their final destination.

[Effects of exogenous spermidine on lipid peroxidation and membrane proton pump activity of cucumber seedling leaves under high temperature stress].

PubMed

Tian, Jing; Guo, Shi-Rong; Sun, Jin; Wang, Li-Ping; Yang, Yan-Juan; Li, Bin

2011-12-01

Taking a relatively heat-resistant cucumber (Cucumis sativus) cultivar 'Jinchun No. 4' as test material, a sand culture experiment was conducted in growth chamber to investigate the effects of foliar spraying spermidine (Spd) on the lipid peroxidation, membrane proton pump activity, and corresponding gene expression of cucumber seedling leaves under high temperature stress. Compared with the control, foliar spraying Spd increased the plant height, stem diameter, dry and fresh mass, and leaf area significantly, and inhibited the increase of leaf relative conductivity, malondialdehyde (MDA) content, and lipoxygenase (LOX) activity effectively. Foliar spraying Spd also helped to the increase of leaf plasma membrane- and tonoplast H(+)-ATPase activity, but no significant difference was observed in the gene expression levels. These results suggested that exogenous Spd could significantly decrease the leaf lipid peroxidation and increase the proton pump activity, and thus, stabilize the leaf membrane structure and function, alleviate the damage induced by high temperature stress, and enhance the heat tolerance of cucumber seedlings.

Novobiocin binding to NalD induces the expression of the MexAB-OprM pump in Pseudomonas aeruginosa.

PubMed

Chen, Weizhong; Wang, Dan; Zhou, Wenquan; Sang, Hong; Liu, Xichun; Ge, Zhiyun; Zhang, Jin; Lan, Lefu; Yang, Cai-Guang; Chen, Hao

2016-06-01

NalD was reported to be the secondary repressor of the MexAB-OprM multidrug efflux pump, the major system contributing to intrinsic multidrug resistance in Pseudomonas aeruginosa. Here, we show that novobiocin binds directly to NalD, which leads NalD to dissociate from the DNA promoter, and thus de-represses the expression of the MexAB-OprM pump. In addition, we have solved the crystal structure of NalD at a resolution of 2.90 Å. The structural alignment of NalD to its homologue TtgR reveals that the residues N129 and H167 in NalD are involved in its novobiocin-binding ability. We have confirmed the function of these two amino acids by EMSA and plate assay. The results presented here highlight the importance and diversity of regulatory mechanism in bacterial antibiotic resistance, and provide further insight for novel antimicrobial development. © 2016 John Wiley & Sons Ltd.

Differential effects of histone deacetylase inhibitors on cellular drug transporters and their implications for using epigenetic modifiers in combination chemotherapy.

PubMed

Valdez, Benigno C; Li, Yang; Murray, David; Brammer, Jonathan E; Liu, Yan; Hosing, Chitra; Nieto, Yago; Champlin, Richard E; Andersson, Borje S

2016-09-27

HDAC inhibitors, DNA alkylators and nucleoside analogs are effective components of combination chemotherapy. To determine a possible mechanism of their synergism, we analyzed the effects of HDAC inhibitors on the expression of drug transporters which export DNA alkylators. Exposure of PEER lymphoma T-cells to 15 nM romidepsin (Rom) resulted in 40%-50% reduction in mRNA for the drug transporter MRP1 and up to ~500-fold increase in the MDR1 mRNA within 32-48 hrs. MRP1 protein levels concomitantly decreased while MDR1 increased. Other HDAC inhibitors - panobinostat, belinostat and suberoylanilide hydroxamic acid (SAHA) - had similar effects on these transporters. The protein level of MRP1 correlated with cellular resistance to busulfan and chlorambucil, and Rom exposure sensitized cells to these DNA alkylators. The decrease in MRP1 correlated with decreased cellular drug export activity, and increased level of MDR1 correlated with increased export of daunorubicin. A similar decrease in the level of MRP1 protein, and increase in MDR1, were observed when mononuclear cells derived from patients with T-cell malignancies were exposed to Rom. Decreased MRP1 and increased MDR1 expressions were also observed in blood mononuclear cells from lymphoma patients who received SAHA-containing chemotherapy in a clinical trial. This inhibitory effect of HDAC inhibitors on the expression of MRP1 suggests that their synergism with DNA alkylating agents is partly due to decreased efflux of these alkylators. Our results further imply the possibility of antagonistic effects when HDAC inhibitors are combined with anthracyclines and other MDR1 drug ligands in chemotherapy.

Human protein Staufen-2 promotes HIV-1 proliferation by positively regulating RNA export activity of viral protein Rev.

PubMed

Banerjee, Atoshi; Benjamin, Ronald; Balakrishnan, Kannan; Ghosh, Payel; Banerjee, Sharmistha

2014-02-13

The export of intron containing viral RNAs from the nucleus to the cytoplasm is an essential step in the life cycle of Human Immunodeficiency Virus-1 (HIV-1). As the eukaryotic system does not permit the transport of intron containing RNA out of the nucleus, HIV-1 makes a regulatory protein, Rev, that mediates the transportation of unspliced and partially spliced viral mRNA from the nucleus to the cytoplasm, thereby playing a decisive role in the generation of new infectious virus particles. Therefore, the host factors modulating the RNA export activity of Rev can be major determinants of virus production in an infected cell. In this study, human Staufen-2 (hStau-2) was identified as a host factor interacting with HIV-1 Rev through affinity chromatography followed by MALDI analyses. Our experiments involving transient expressions, siRNA mediated knockdowns and infection assays conclusively established that hStau-2 is a positive regulator of HIV-1 pathogenesis. We demonstrated that Rev-hStau-2 interactions positively regulated the RNA export activity of Rev and promoted progeny virus synthesis. The Rev-hStau-2 interaction was independent of RNA despite both being RNA binding proteins. hStau-2 mutant, with mutations at Q314R-A318F-K319E, deficient of binding Rev, failed to promote hStau-2 dependent Rev activity and viral production, validating the essentiality of this protein-protein interaction. The expression of this positive regulator was elevated upon HIV-1 infection in both human T-lymphocyte and astrocyte cell lines. With this study, we establish that human Staufen-2, a host factor which is up-regulated upon HIV-1 infection, interacts with HIV-1 Rev, thereby promoting its RNA export activity and progeny virus formation. Altogether, our study provides new insights into the emerging role of the Staufen family of mRNA transporters in host-pathogen interaction and supports the notion that obliterating interactions between viral and host proteins that positively regulate HIV-1 proliferation can significantly contribute to anti-retroviral treatments.

Identification of Bicarbonate as a Trigger and Genes Involved with Extracellular DNA Export in Mycobacterial Biofilms.

PubMed

Rose, Sasha J; Bermudez, Luiz E

2016-12-06

Extracellular DNA (eDNA) is an integral biofilm matrix component of numerous pathogens, including nontuberculous mycobacteria (NTM). Cell lysis is the source of eDNA in certain bacteria, but the source of eDNA remains unidentified for NTM, as well as for other eDNA-containing bacterial species. In this study, conditions affecting eDNA export were examined, and genes involved with the eDNA export mechanism were identified. After a method for monitoring eDNA in real time in undisturbed biofilms was established, different conditions affecting eDNA were investigated. Bicarbonate positively influenced eDNA export in a pH-independent manner in Mycobacterium avium, M. abscessus, and M. chelonae The surface-exposed proteome of M. avium in eDNA-containing biofilms revealed abundant carbonic anhydrases. Chemical inhibition of carbonic anhydrases with ethoxzolamide significantly reduced eDNA export. An unbiased transposon mutant library screen for eDNA export in M. avium identified many severely eDNA-attenuated mutants, including one not expressing a unique FtsK/SpoIIIE-like DNA-transporting pore, two with inactivation of carbonic anhydrases, and nine with inactivation of genes belonging to a unique genomic region, as well as numerous mutants involved in metabolism and energy production. Complementation of nine mutants that included the FtsK/SpoIIIE and carbonic anhydrase significantly restored eDNA export. Interestingly, several attenuated eDNA mutants have mutations in genes encoding proteins that were found with the surface proteomics, and many more mutations are localized in operons potentially encoding surface proteins. Collectively, our data strengthen the evidence of eDNA export being an active mechanism that is activated by the bacterium responding to bicarbonate. Many bacteria contain extracellular DNA (eDNA) in their biofilm matrix, as it has various biological and physical functions. We recently reported that nontuberculous mycobacteria (NTM) can contain significant quantities of eDNA in their biofilms. In some bacteria, eDNA is derived from dead cells, but that does not appear to be the case for all eDNA-containing organisms, including NTM. In this study, we found that eDNA export in NTM is conditionally dependent on the molecules to which the bacteria are exposed and that bicarbonate positively influences eDNA export. We also identified genes and proteins important for eDNA export, which begins to piece together a description of a mechanism for eDNA. Better understanding of eDNA export can give new targets for the development of antivirulence drugs, which are attractive because resistance to classical antibiotics is currently a significant problem. Copyright © 2016 Rose and Bermudez.

Research on energy conversion mechanism of rotodynamic pump and design of non-overload centrifugal pump

NASA Astrophysics Data System (ADS)

Zhang, X. L.; Hu, S. B.; Shen, Z. Z.; Wu, S. P.; Li, K.

2016-05-01

In this paper, an attempt has been made for the calculation of an expression for the intrinsic law of input power which has not yet been given by current theory of Rotodynamic pump. By adequate recognition of the characteristics of non-inertial system within the rotating impeller, it is concluded that the input power consists of two power components, the first power component, whose magnitude increases with the increase of the flow rate, corresponds to radial velocity component, and the second power component, whose magnitude decreases with the increase of the flow rate, corresponds to tangential velocity component, therefore, the law of rise, basic levelness and drop of input power curves of centrifugal pump, mixed-flow pump and axial-flow pump can be explained reasonably. Through further analysis, the main ways for realizing non-overload of centrifugal pump are obtained, and its equivalent design factor is found out, the factor correlates with the outlet angle of leading face and back face of the blade, wrap angle, number of blades, outlet width, area ratio, and the ratio of operating flow rate to specified flow rate and so on. These are verified with actual example.

Ram locus is a key regulator to trigger multidrug resistance in Enterobacter aerogenes.

PubMed

Molitor, Alexander; James, Chloë E; Fanning, Séamus; Pagès, Jean-Marie; Davin-Regli, Anne

2018-02-01

Several genetic regulators belonging to AraC family are involved in the emergence of MDR isolates of E. aerogenes due to alterations in membrane permeability. Compared with the genetic regulator Mar, RamA may be more relevant towards the emergence of antibiotic resistance. Focusing on the global regulators, Mar and Ram, we compared the amino acid sequences of the Ram repressor in 59 clinical isolates and laboratory strains of E. aerogenes. Sequence types were associated with their corresponding multi-drug resistance phenotypes and membrane protein expression profiles using MIC and immunoblot assays. Quantitative gene expression analysis of the different regulators and their targets (porins and efflux pump components) were performed. In the majority of the MDR isolates tested, ramR and a region upstream of ramA were mutated but marR or marA were unchanged. Expression and cloning experiments highlighted the involvement of the ram locus in the modification of membrane permeability. Overexpression of RamA lead to decreased porin production and increased expression of efflux pump components, whereas overexpression of RamR had the opposite effects. Mutations or deletions in ramR, leading to the overexpression of RamA predominated in clinical MDR E. aerogenes isolates and were associated with a higher-level of expression of efflux pump components. It was hypothesised that mutations in ramR, and the self-regulating region proximal to ramA, probably altered the binding properties of the RamR repressor; thereby producing the MDR phenotype. Consequently, mutability of RamR may play a key role in predisposing E. aerogenes towards the emergence of a MDR phenotype.

Drug transporters, the blood–testis barrier, and spermatogenesis

PubMed Central

Su, Linlin; Mruk, Dolores D; Cheng, C Yan

2015-01-01

The blood–testis barrier (BTB), which is created by adjacent Sertoli cells near the basement membrane, serves as a ‘gatekeeper’ to prohibit harmful substances from reaching developing germ cells, most notably postmeiotic spermatids. The BTB also divides the seminiferous epithelium into the basal and adluminal (apical) compartment so that postmeiotic spermatid development, namely spermiogenesis, can take place in a specialized microenvironment in the apical compartment behind the BTB. The BTB also contributes, at least in part, to the immune privilege status of the testis, so that anti-sperm antibodies are not developed against antigens that are expressed transiently during spermatogenesis. Recent studies have shown that numerous drug transporters are expressed by Sertoli cells. However, many of these same drug transporters are also expressed by spermatogonia, spermatocytes, round spermatids, elongating spermatids, and elongated spermatids, suggesting that the developing germ cells are also able to selectively pump drugs ‘in’ and/or ‘out’ via influx or efflux pumps. We review herein the latest developments regarding the role of drug transporters in spermatogenesis. We also propose a model utilized by the testis to protect germ cell development from ‘harmful’ environmental toxicants and xenobiotics and/or from ‘therapeutic’ substances (e.g. anticancer drugs). We also discuss how drug transporters that are supposed to protect spermatogenesis can work against the testis in some instances. For example, when drugs (e.g. male contraceptives) that can perturb germ cell adhesion and/or maturation are actively pumped out of the testis or are prevented from entering the apical compartment, such as by efflux pumps. PMID:21134990

27 CFR 28.43 - Evidence of exportation and lading for use on vessels and aircraft: beer.

Code of Federal Regulations, 2012 CFR

2012-04-01

.... L. 85-859, 72 Stat. 1334, as amended, 1335, as amended (26 U.S.C. 5053, 5055)) [T.D. ATF-224, 51 FR...) or Form 1689 (5130.12) under § 28.275; or (3) A bill of lading (§ 28.250), a railway express receipt (§ 28.251), or an air express or air freight bill of lading (§ 28.252), when such bills of lading or...

27 CFR 28.43 - Evidence of exportation and lading for use on vessels and aircraft: beer.

Code of Federal Regulations, 2013 CFR

2013-04-01

.... L. 85-859, 72 Stat. 1334, as amended, 1335, as amended (26 U.S.C. 5053, 5055)) [T.D. ATF-224, 51 FR...) or Form 1689 (5130.12) under § 28.275; or (3) A bill of lading (§ 28.250), a railway express receipt (§ 28.251), or an air express or air freight bill of lading (§ 28.252), when such bills of lading or...

27 CFR 28.43 - Evidence of exportation and lading for use on vessels and aircraft: beer.

Code of Federal Regulations, 2014 CFR

2014-04-01

.... L. 85-859, 72 Stat. 1334, as amended, 1335, as amended (26 U.S.C. 5053, 5055)) [T.D. ATF-224, 51 FR...) or Form 1689 (5130.12) under § 28.275; or (3) A bill of lading (§ 28.250), a railway express receipt (§ 28.251), or an air express or air freight bill of lading (§ 28.252), when such bills of lading or...

Functional expression of a human GDP-L-fucose transporter in Escherichia coli.

PubMed

Förster-Fromme, Karin; Schneider, Sarah; Sprenger, Georg A; Albermann, Christoph

2017-02-01

To investigate the translocation of nucleotide-activated sugars from the cytosol across a membrane into the endoplasmatic reticulum or the Golgi apparatus which is an important step in the synthesis of glycoproteins and glycolipids in eukaryotes. The heterologous expression of the recombinant and codon-adapted human GDP-L-fucose antiporter gene SLC35C1 (encoding an N-terminal OmpA-signal sequence) led to a functional transporter protein located in the cytoplasmic membrane of Escherichia coli. The in vitro transport was investigated using inverted membrane vesicles. SLC35C1 is an antiporter specific for GDP-L-fucose and depending on the concomitant reverse transport of GMP. The recombinant transporter FucT1 exhibited an activity for the transport of 3 H-GDP-L-fucose with a V max of 8 pmol/min mg with a K m of 4 µM. The functional expression of SLC35C1 in GDP-L-fucose overproducing E. coli led to the export of GDP-L-fucose to the culture supernatant. The export of GDP-L-fucose by E. coli provides the opportunity for the engineering of a periplasmatic fucosylation reaction in recombinant bacterial cells.

Erythropoietin and carbamylated erythropoietin promote histone deacetylase 5 phosphorylation and nuclear export in rat hippocampal neurons

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

Jo, Hye-Ryeong; Kim, Yong-Seok; Department of Biochemistry and Molecular Biology, College of Medicine, Hanyang University, 17 Haengdang-dong, Sungdong-gu, Seoul 133-791

2016-01-29

Erythropoietin (EPO) produces neurotrophic effects in animal model of neurodegeneration. However, clinical use of EPO is limited due to thrombotic risk. Carbamylated EPO (cEPO), devoid of thrombotic risk, has been proposed as a novel neuroprotective and neurotrophic agent although the molecular mechanisms of cEPO remain incomplete. Here, we show a previously unidentified role of histone deacetylase 5 (HDAC5) in the actions of EPO and cEPO. EPO and cEPO regulate the HDAC5 phosphorylation at two critical sites, Ser259 and Ser498 through a protein kinase D (PKD) dependent pathway. In addition, EPO and cEPO rapidly stimulates nuclear export of HDAC5 in ratmore » hippocampal neurons which expressing HDAC5-GFP. Consequently, EPO and cEPO enhanced the myocyte enhancer factor-2 (MEF2) target gene expression. Taken together, our results reveal that EPO and cEPO mediate MEF2 target gene expression via the regulation of HDAC5 phosphorylation at Ser259/498, and suggest that HDAC5 could be a potential mechanism contributing to the therapeutic actions of EPO and cEPO.« less

Table-top two-color soft X-ray laser by means of Ni-like plasmas

NASA Astrophysics Data System (ADS)

Masoudnia, Leili; Ruiz-Lopez, Mabel; Bleiner, Davide

2016-04-01

Laser-produced Ni-like plasmas are known as active media for extreme ultraviolet lasing, with the flexibility to two-color lasing. Two-color laser generation is very complex at accelerator facilities. In this work, plasma lasing at the 3d94d1(J = 0) → 3d94p1(J = 1) (collisional-pumping process) and the 3d94f1(J = 1) → 3d94d1(J = 1) (photo-pumping process) transitions is studied experimentally and computationally. Several key characteristics of collisional- and photo-pumping laser, such as divergence, pointing stability, and intensity have been investigated. The measurements showed different pulse characteristics for the two lasing processes affected by plasma inhomogeneity in temperature and density. Analytical expressions of these characteristics for both collisional- and photo-pumping are derived. It is found that the plasma that maximizes the photo-pumping lasing is 20% hotter and 70% denser than the plasma that optimizes the collisional-pumping lasing. The gain of collisional pumping is ≈4 times higher than the gain for the photo-pumping. The gain lifetime is a factor of ≈5.2 larger for the monopole-pumping. Similarly, the gain thickness is a factor of ≈1.8 larger. It is also found that the gain build-up time for collisional- and photo-pumping is 0.7 ps and 0.9 ps, respectively, whereas the build-up length-scale is 11.5 μm and 6.3 μm, respectively.

How NaCl raises blood pressure: a new paradigm for the pathogenesis of salt-dependent hypertension

PubMed Central

Leenen, Frans H. H.; Chen, Ling; Golovina, Vera A.; Hamlyn, John M.; Pallone, Thomas L.; Van Huysse, James W.; Zhang, Jin; Wier, W. Gil

2012-01-01

Excess dietary salt is a major cause of hypertension. Nevertheless, the specific mechanisms by which salt increases arterial constriction and peripheral vascular resistance, and thereby raises blood pressure (BP), are poorly understood. Here we summarize recent evidence that defines specific molecular links between Na+ and the elevated vascular resistance that directly produces high BP. In this new paradigm, high dietary salt raises cerebrospinal fluid [Na+]. This leads, via the Na+-sensing circumventricular organs of the brain, to increased sympathetic nerve activity (SNA), a major trigger of vasoconstriction. Plasma levels of endogenous ouabain (EO), the Na+ pump ligand, also become elevated. Remarkably, high cerebrospinal fluid [Na+]-evoked, locally secreted (hypothalamic) EO participates in a pathway that mediates the sustained increase in SNA. This hypothalamic signaling chain includes aldosterone, epithelial Na+ channels, EO, ouabain-sensitive α2 Na+ pumps, and angiotensin II (ANG II). The EO increases (e.g.) hypothalamic ANG-II type-1 receptor and NADPH oxidase and decreases neuronal nitric oxide synthase protein expression. The aldosterone-epithelial Na+ channel-EO-α2 Na+ pump-ANG-II pathway modulates the activity of brain cardiovascular control centers that regulate the BP set point and induce sustained changes in SNA. In the periphery, the EO secreted by the adrenal cortex directly enhances vasoconstriction via an EO-α2 Na+ pump-Na+/Ca2+ exchanger-Ca2+ signaling pathway. Circulating EO also activates an EO-α2 Na+ pump-Src kinase signaling cascade. This increases the expression of the Na+/Ca2+ exchanger-transient receptor potential cation channel Ca2+ signaling pathway in arterial smooth muscle but decreases the expression of endothelial vasodilator mechanisms. Additionally, EO is a growth factor and may directly participate in the arterial structural remodeling and lumen narrowing that is frequently observed in established hypertension. These several central and peripheral mechanisms are coordinated, in part by EO, to effect and maintain the salt-induced elevation of BP. PMID:22058154

Migrant biomass and respiratory carbon flux by zooplankton and micronekton in the subtropical northeast Atlantic Ocean (Canary Islands)

NASA Astrophysics Data System (ADS)

Ariza, A.; Garijo, J. C.; Landeira, J. M.; Bordes, F.; Hernández-León, S.

2015-05-01

Diel Vertical Migration (DVM) in marine ecosystems is performed by zooplankton and micronekton, promoting a poorly accounted export of carbon to the deep ocean. Major efforts have been made to estimate carbon export due to gravitational flux and to a lesser extent, to migrant zooplankton. However, migratory flux by micronekton has been largely neglected in this context, due to its time-consuming and difficult sampling. In this paper, we evaluated gravitational and migratory flux due to the respiration of zooplankton and micronekton in the northeast subtropical Atlantic Ocean (Canary Islands). Migratory flux was addressed by calculating the biomass of migrating components and measuring the electron transfer system (ETS) activity in zooplankton and dominant species representing micronekton (Euphausia gibboides, Sergia splendens and Lobianchia dofleini). Our results showed similar biomass in both components. The main taxa contributing to DVM within zooplankton were juvenile euphausiids, whereas micronekton were mainly dominated by fish, followed by adult euphausiids and decapods. The contribution to respiratory flux of zooplankton (3.4 ± 1.9 mg C m-2 d-1) was similar to that of micronekton (2.9 ± 1.0 mg C m-2 d-1). In summary, respiratory flux accounted for 53% (range 23-71) of the gravitational flux measured at 150 m depth (11.9 ± 5.8 mg C m-2 d-1). However, based on larger migratory ranges and gut clearance rates, micronekton are expected to be the dominant component that contributes to carbon export in deeper waters. Micronekton estimates in this paper as well as those in existing literature, although variable due to regional differences and difficulties in calculating their biomass, suggest that carbon fluxes driven by this community are important for future models of the biological carbon pump.

A lagged variable model for characterizing temporally dynamic export of legacy anthropogenic nitrogen from watersheds to rivers.

PubMed

Chen, Dingjiang; Guo, Yi; Hu, Minpeng; Dahlgren, Randy A

2015-08-01

Legacy nitrogen (N) sources originating from anthropogenic N inputs (NANI) may be a major cause of increasing riverine N exports in many regions, despite a significant decline in NANI. However, little quantitative knowledge exists concerning the lag effect of NANI on riverine N export. As a result, the N leaching lag effect is not well represented in most current watershed models. This study developed a lagged variable model (LVM) to address temporally dynamic export of watershed NANI to rivers. Employing a Koyck transformation approach used in economic analyses, the LVM expresses the indefinite number of lag terms from previous years' NANI with a lag term that incorporates the previous year's riverine N flux, enabling us to inversely calibrate model parameters from measurable variables using Bayesian statistics. Applying the LVM to the upper Jiaojiang watershed in eastern China for 1980-2010 indicated that ~97% of riverine export of annual NANI occurred in the current year and succeeding 10 years (~11 years lag time) and ~72% of annual riverine N flux was derived from previous years' NANI. Existing NANI over the 1993-2010 period would have required a 22% reduction to attain the target TN level (1.0 mg N L(-1)), guiding watershed N source controls considering the lag effect. The LVM was developed with parsimony of model structure and parameters (only four parameters in this study); thus, it is easy to develop and apply in other watersheds. The LVM provides a simple and effective tool for quantifying the lag effect of anthropogenic N input on riverine export in support of efficient development and evaluation of watershed N control strategies.

Suppressor Analysis Reveals a Role for SecY in the SecA2-Dependent Protein Export Pathway of Mycobacteria

PubMed Central

Ligon, Lauren S.; Rigel, Nathan W.; Romanchuk, Artur; Jones, Corbin D.

2013-01-01

All bacteria use the conserved Sec pathway to transport proteins across the cytoplasmic membrane, with the SecA ATPase playing a central role in the process. Mycobacteria are part of a small group of bacteria that have two SecA proteins: the canonical SecA (SecA1) and a second, specialized SecA (SecA2). The SecA2-dependent pathway exports a small subset of proteins and is required for Mycobacterium tuberculosis virulence. The mechanism by which SecA2 drives export of proteins across the cytoplasmic membrane remains poorly understood. Here we performed suppressor analysis on a dominant negative secA2 mutant (secA2 K129R) of the model mycobacterium Mycobacterium smegmatis to better understand the pathway used by SecA2 to export proteins. Two extragenic suppressor mutations were identified as mapping to the promoter region of secY, which encodes the central component of the canonical Sec export channel. These suppressor mutations increased secY expression, and this effect was sufficient to alleviate the secA2 K129R phenotype. We also discovered that the level of SecY protein was greatly diminished in the secA2 K129R mutant, but at least partially restored in the suppressors. Furthermore, the level of SecY in a suppressor strongly correlated with the degree of suppression. Our findings reveal a detrimental effect of SecA2 K129R on SecY, arguing for an integrated system in which SecA2 works with SecY and the canonical Sec translocase to export proteins. PMID:23913320

Examining the intersection between splicing, nuclear export and small RNA pathways.

PubMed

Nabih, Amena; Sobotka, Julia A; Wu, Monica Z; Wedeles, Christopher J; Claycomb, Julie M

2017-11-01

Nuclear Argonaute/small RNA pathways in a variety of eukaryotic species are generally known to regulate gene expression via chromatin modulation and transcription attenuation in a process known as transcriptional gene silencing (TGS). However, recent data, including genetic screens, phylogenetic profiling, and molecular mechanistic studies, also point to a novel and emerging intersection between the splicing and nuclear export machinery with nuclear Argonaute/small RNA pathways in many organisms. In this review, we summarize the field's current understanding regarding the relationship between splicing, export and small RNA pathways, and consider the biological implications for coordinated regulation of transcripts by these pathways. We also address the importance and available approaches for understanding the RNA regulatory logic generated by the intersection of these particular pathways in the context of synthetic biology. The interactions between various eukaryotic RNA regulatory pathways, particularly splicing, nuclear export and small RNA pathways provide a type of combinatorial code that informs the identity ("self" versus "non-self") and dictates the fate of each transcript in a cell. Although the molecular mechanisms for how splicing and nuclear export impact small RNA pathways are not entirely clear at this early stage, the links between these pathways are widespread across eukaryotic phyla. The link between splicing, nuclear export, and small RNA pathways is emerging and establishes a new frontier for understanding the combinatorial logic of gene regulation across species that could someday be harnessed for therapeutic, biotechnology and agricultural applications. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

Featured Article: Nuclear export of opioid growth factor receptor is CRM1 dependent.

PubMed

Kren, Nancy P; Zagon, Ian S; McLaughlin, Patricia J

2016-02-01

Opioid growth factor receptor (OGFr) facilitates growth inhibition in the presence of its specific ligand opioid growth factor (OGF), chemically termed [Met(5)]-enkephalin. The function of the OGF-OGFr axis requires the receptor to translocate to the nucleus. However, the mechanism of nuclear export of OGFr is unknown. In this study, endogenous OGFr, as well as exogenously expressed OGFr-EGFP, demonstrated significant nuclear accumulation in response to leptomycin B (LMB), an inhibitor of CRM1-dependent nuclear export, suggesting that OGFr is exported in a CRM1-dependent manner. One consensus sequence for a nuclear export signal (NES) was identified. Mutation of the associated leucines, L217 L220 L223 and L225, to alanine resulted in decreased nuclear accumulation. NES-EGFP responded to LMB, indicating that this sequence is capable of functioning as an export signal in isolation. To determine why the sequence functions differently in isolation than as a full length protein, the localization of subNES was evaluated in the presence and absence of MG132, a potent inhibitor of proteosomal degradation. MG132 had no effect of subNES localization. The role of tandem repeats located at the C-terminus of OGFr was examined for their role in nuclear trafficking. Six of seven tandem repeats were removed to form deltaTR. DeltaTR localized exclusively to the nucleus indicating that the tandem repeats may contribute to the localization of the receptor. Similar to the loss of cellular proliferation activity (i.e. inhibition) recorded with subNES, deltaTR also demonstrated a significant loss of inhibitory activity indicating that the repeats may be integral to receptor function. These experiments reveal that OGFr contains one functional NES, L217 L220 L223 and L225 and can be exported from the nucleus in a CRM1-dependent manner. © 2015 by the Society for Experimental Biology and Medicine.

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

PubMed Central

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

2016-01-01

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

CD4-CCR5 interaction in intracellular compartments contributes to receptor expression at the cell surface

PubMed Central

Achour, Lamia; Scott, Mark G.H.; Shirvani, Hamasseh; Thuret, Alain; Bismuth, Georges; Labbé-Jullié, Catherine; Marullo, Stefano

2009-01-01

The association of CD4, a glycoprotein involved in T cell development and antigen recognition, and CCR5, a chemotactic G protein-coupled receptor, which regulates trafficking and effector functions of immune cells, forms the main receptor for the human immunodeficiency virus HIV. We observed that the vast majority of CCR5 is maintained within the intracellular compartments of primary T lymphocytes and in a monocytic cell line, contrasting with its relative low density at the cell surface. The CCR5-CD4 association, which occurs in the endoplasmic reticulum, enhanced CCR5 export to the plasma membrane in a concentration–dependent manner, whereas inhibition of endogenous CD4 with small interfering RNAs decreased cell surface expression of endogenous CCR5. This effect was specific for CCR5, as CD4 did not affect cell distribution of CXCR4, the other HIV co-receptor. These results reveal a previously unappreciated role of CD4, which contributes to regulate CCR5 export to the plasma membrane. PMID:19064722

Energy Factor Analysis for Gas Heat Pump Water Heaters

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

Gluesenkamp, Kyle R

2016-01-01

Gas heat pump water heaters (HPWHs) can improve water heating efficiency with zero GWP and zero ODP working fluids. The energy factor (EF) of a gas HPWH is sensitive to several factors. In this work, expressions are derived for EF of gas HPWHs, as a function of heat pump cycle COP, tank heat losses, burner efficiency, electrical draw, and effectiveness of supplemental heat exchangers. The expressions are used to investigate the sensitivity of EF to each parameter. EF is evaluated on a site energy basis (as used by the US DOE for rating water heater EF), and a primary energy-basismore » energy factor (PEF) is also defined and included. Typical ranges of values for the six parameters are given. For gas HPWHs, using typical ranges for component performance, EF will be 59 80% of the heat pump cycle thermal COP (for example, a COP of 1.60 may result in an EF of 0.94 1.28). Most of the reduction in COP is due to burner efficiency and tank heat losses. Gas-fired HPWHs are theoretically be capable of an EF of up to 1.7 (PEF of 1.6); while an EF of 1.1 1.3 (PEF of 1.0 1.1) is expected from an early market entry.« less

Crystal structure of Escherichia coli-expressed Haloarcula marismortui bacteriorhodopsin I in the trimeric form.

PubMed

Shevchenko, Vitaly; Gushchin, Ivan; Polovinkin, Vitaly; Round, Ekaterina; Borshchevskiy, Valentin; Utrobin, Petr; Popov, Alexander; Balandin, Taras; Büldt, Georg; Gordeliy, Valentin

2014-01-01

Bacteriorhodopsins are a large family of seven-helical transmembrane proteins that function as light-driven proton pumps. Here, we present the crystal structure of a new member of the family, Haloarcula marismortui bacteriorhodopsin I (HmBRI) D94N mutant, at the resolution of 2.5 Å. While the HmBRI retinal-binding pocket and proton donor site are similar to those of other archaeal proton pumps, its proton release region is extended and contains additional water molecules. The protein's fold is reinforced by three novel inter-helical hydrogen bonds, two of which result from double substitutions relative to Halobacterium salinarum bacteriorhodopsin and other similar proteins. Despite the expression in Escherichia coli and consequent absence of native lipids, the protein assembles as a trimer in crystals. The unique extended loop between the helices D and E of HmBRI makes contacts with the adjacent protomer and appears to stabilize the interface. Many lipidic hydrophobic tail groups are discernible in the membrane region, and their positions are similar to those of archaeal isoprenoid lipids in the crystals of other proton pumps, isolated from native or native-like sources. All these features might explain the HmBRI properties and establish the protein as a novel model for the microbial rhodopsin proton pumping studies.

Coccolithovirus facilitation of carbon export in the North Atlantic.

PubMed

Laber, Christien P; Hunter, Jonathan E; Carvalho, Filipa; Collins, James R; Hunter, Elias J; Schieler, Brittany M; Boss, Emmanuel; More, Kuldeep; Frada, Miguel; Thamatrakoln, Kimberlee; Brown, Christopher M; Haramaty, Liti; Ossolinski, Justin; Fredricks, Helen; Nissimov, Jozef I; Vandzura, Rebecca; Sheyn, Uri; Lehahn, Yoav; Chant, Robert J; Martins, Ana M; Coolen, Marco J L; Vardi, Assaf; DiTullio, Giacomo R; Van Mooy, Benjamin A S; Bidle, Kay D

2018-05-01

Marine phytoplankton account for approximately half of global primary productivity 1 , making their fate an important driver of the marine carbon cycle. Viruses are thought to recycle more than one-quarter of oceanic photosynthetically fixed organic carbon 2 , which can stimulate nutrient regeneration, primary production and upper ocean respiration 2 via lytic infection and the 'virus shunt'. Ultimately, this limits the trophic transfer of carbon and energy to both higher food webs and the deep ocean 2 . Using imagery taken by the Moderate Resolution Imaging Spectroradiometer (MODIS) onboard the Aqua satellite, along with a suite of diagnostic lipid- and gene-based molecular biomarkers, in situ optical sensors and sediment traps, we show that Coccolithovirus infections of mesoscale (~100 km) Emiliania huxleyi blooms in the North Atlantic are coupled with particle aggregation, high zooplankton grazing and greater downward vertical fluxes of both particulate organic and particulate inorganic carbon from the upper mixed layer. Our analyses captured blooms in different phases of infection (early, late and post) and revealed the highest export flux in 'early-infected blooms' with sinking particles being disproportionately enriched with infected cells and subsequently remineralized at depth in the mesopelagic. Our findings reveal viral infection as a previously unrecognized ecosystem process enhancing biological pump efficiency.

Enhanced Particulate Organic Carbon Export at Eddy Edges in the Oligotrophic Western North Pacific Ocean

PubMed Central

Shih, Yung-Yen; Hung, Chin-Chang; Gong, Gwo-Ching; Chung, Wan-Chen; Wang, Yu-Huai; Lee, I-Huan; Chen, Kuo-Shu; Ho, Chuang-Yi

2015-01-01

Mesoscale eddies in the subtropical oligotrophic ocean are ubiquitous and play an important role in nutrient supply and oceanic primary production. However, it is still unclear whether these mesoscale eddies can efficiently transfer CO2 from the atmosphere to deep waters via biological pump because of the sampling difficulty due to their transient nature. In 2007, particulate organic carbon (POC) fluxes, measured below the euphotic zone at the edge of warm eddy were 136–194 mg-C m−2 d−1 which was greatly elevated over that (POC flux = 26–35 mg-C m−2 d−1) determined in the nutrient-depleted oligotrophic waters in the Western North Pacific (WNP). In 2010, higher POC fluxes (83–115 mg-C m−2 d−1) were also observed at the boundary of mesoscale eddies in the WNP. The enhanced POC flux at the edge of eddies was mainly attributed to both large denuded diatom frustules and zooplankton fecal pellets based on scanning electron microscopy (SEM) examination. The result suggests that mesoscale eddies in the oligotrophic waters in the subtropical WNP can efficiently increase the oceanic carbon export flux and the eddy edge is a crucial conduit in carbon sequestration to deep waters. PMID:26171611

Pertussis toxin export genes are regulated by the ptx promoter and may be required for efficient translation of ptx mRNA in Bordetella pertussis.

PubMed Central

Baker, S M; Masi, A; Liu, D F; Novitsky, B K; Deich, R A

1995-01-01

The gene products from an 8-kb region adjacent to the 3' end of the ptx operon are required by Bordetella pertussis for the export of pertussis holotoxin. At least one of these gene products (PtlC) is specifically required for the export of assembled holotoxin from the periplasmic space. ptlC mutants exhibit a 20-fold reduction in the amount of holotoxin present in the culture supernatant but have no effect upon the assembly or steady-state level of holotoxin present in the periplasmic space. Impaired export of holotoxin from the ptlC strain blocks expression of toxin at a posttranscriptional level, and wild-type levels of ptx mRNA are detected in the mutant strain. The transcription of ptl is subject to modulation by MgSO4 in the same manner as ptx is; however, in B. pertussis strains containing an E. coli tac promoter in place of the native ptx promoter, wild-type levels of ptx mRNA are present and holotoxin is synthesized and exported even in the presence of MgSO4. Promoter mapping of the region extending from the ptxS3 coding region to the ptlC coding region failed to detect the ptl transcription initiation site. Additional RNase protection experiments with ptx promoter deletion and substitution strains indicate that the ptl operon is transcribed from the ptx promoter as part of a > 11-kb mRNA. PMID:7558300

PAUSED Encodes the Arabidopsis Exportin-t Ortholog1

PubMed Central

Hunter, Christine A.; Aukerman, Milo J.; Sun, Hui; Fokina, Maria; Poethig, R. Scott

2003-01-01

Los1p/exportin-t (XPOT) mediates the nuclear export of tRNAs in yeast and mammals. The requirements for this transport pathway are unclear, however, because los1 mutations do not affect yeast growth, and the phenotype of XPOT mutations in mammals is unknown. Here, we show that PAUSED (PSD) is the Arabidopsis ortholog of LOS1/XPOT and is capable of rescuing the tRNA export defect of los1 in Brewer's yeast (Saccharomyces cerevisiae), suggesting that its function has been conserved. Putative null alleles of PSD disrupt the initiation of the shoot apical meristem and delay leaf initiation after germination, the emergence of the radicle and lateral roots, and the transition to flowering. Plants doubly mutant for psd and hasty, the Arabidopsis ortholog of exportin 5, are viable but have a more severe phenotype than either single mutant. These results suggest that PSD plays a role in tRNA export in Arabidopsis, but that at least one—and perhaps several—additional tRNA export pathways also exist. The PSD transcript is broadly expressed during development and is alternatively spliced in the 3′-untranslated region. No temporal or spatial difference in the abundance of different splice forms was observed. We propose that the mutant phenotype of psd reflects defects in developmental events and cell/tissue types that require elevated levels of protein synthesis and are therefore acutely sensitive to a reduction in tRNA export. PMID:12913168

RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs

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

Huang, Feng; Zhang, Junsong; Zhang, Yijun

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-boxmore » of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. - Highlights: • MOV10 can function as a co-factor of HIV-1 Rev. • MOV10 facilitates Rev/RRE-dependent transport of viral mRNAs. • MOV10 interacts with Rev in an RNA-independent manner. • The DEAG-box of MOV10 is required for the enhancement of Rev/RRE-dependent export.« less

Determinants of the Intention to Pump Breast Milk on a University Campus.

PubMed

Bai, Yeon K; Dinour, Lauren M; Pope, Gina A

2016-09-01

The number of young mothers in the workforce and in schools of higher education has steadily increased. In order to maintain a breastfeeding relationship with their children, these mothers need to pump or express breast milk multiple times a day while at work or school. This study examines the factors associated with the intention to pump breast milk at one university campus. Between January and February 2015, an online survey invitation was sent out to all female employees and students at one university. The survey, based on the Theory of Planned Behavior, assessed intentions to pump breast milk on campus. The intention to pump breast milk was examined between employees and students separately. Within these 2 groups, behavioral performers (women who pump or have pumped breast milk while on campus) were compared to nonperformers. Using multiple regression analysis, the most influential predictors of the intention to pump (ie, attitudes, subjective norm, perceived behavioral control, and underlying beliefs) were identified. A total of 218 women participated in the study (62 employees and 156 students, a 71.7% survey completion rate). Among university employees, the most influential factor that predicted pumping intention among performers was attitude toward pumping (β = 0.36, P = .03). Among student performers, the most influential factor to predict pumping intention was the subjective norm (β = 0.31, P = .02). For student nonperformers, perceived behavioral control (β = 0.54, P < .001) was the most influential factor. Important determinants of the intention to pump on campus included relieving discomfort from engorgement, availability of milk storage, experiencing other people's approval of pumping breast milk, and the inconvenience of carrying pump equipment. Continued efforts are needed to create a supportive culture for breastfeeding in the campus community as well as to provide pump loan and milk storage options for both employee and student mothers. © 2016 by the American College of Nurse-Midwives.

The effect of pumping noise on the characteristics of a single-stage parametric amplifier

NASA Astrophysics Data System (ADS)

Medvedev, S. Iu.; Muzychuk, O. V.

1983-10-01

An analysis is made of the operation of a single-stage parametric amplifier based on a varactor with a sharp transition. Analytical expressions are obtained for the statistical moments of the output signal, the signal-noise ratio, and other characteristics in the case when the output signal and the pump are a mixture of harmonic oscillation and Gaussian noise. It is shown that, when a noise component is present in the pump, an increase of its harmonic component to values close to the threshold leads to a sharp decrease in the signal-noise ratio at the amplifier output.

The plasma membrane calcium pumps: focus on the role in (neuro)pathology.

PubMed

Brini, Marisa; Carafoli, Ernesto; Calì, Tito

2017-02-19

The plasma membrane Ca 2+ ATPase (PMCA pump) is a member of the superfamily of P-type pumps. It is organized in the plasma membrane with ten transmembrane helices and two main cytosolic loops, one of which contains the catalytic center. It also contains a long C-terminal tail that houses the binding site for calmodulin, the main regulator of the activity of the pump. The pump also contains a number of other regulators, among them acidic phospholipids, kinases, and numerous protein interactors. Separate genes code for 4 basic pump isoforms in mammals, additional isoform complexity being generated by the alternative splicing of primary transcripts. Pumps 1 and 4 are expressed ubiquitously, pumps 2 and 3 are tissue restricted, with preference for the nervous system. In essentially all cells, the pump coexists with much more powerful systems that clear Ca 2+ from the cytosol, e.g. the SERCA pump and the Na + /Ca 2+ exchanger. Its role in the global regulation of cellular Ca 2+ homeostasis is thus quantitatively marginal: its main function is the regulation of Ca 2+ signaling in selected sub-plasma membrane microdomains where Ca 2+ modulated interactors also reside. Malfunctions of the pump linked to genetic mutations are now described with increasing frequency, the disease phenotypes being especially severe in the nervous system where isoforms 2 and 3 predominate. The analysis of the pump defects suggests that the disease phenotypes are likely to be related to the imperfect modulation of Ca 2+ signaling in selected sub-plasma membrane microdomains, leading to the defective control of the activity of important Ca 2+ dependent interactors. Copyright © 2016 Elsevier Inc. All rights reserved.

The anthraquinone emodin inhibits the non-exported FIKK kinase from Plasmodium falciparum.

PubMed

Lin, Benjamin C; Harris, Darcy R; Kirkman, Lucy M D; Perez, Astrid M; Qian, Yiwen; Schermerhorn, Janse T; Hong, Min Y; Winston, Dennis S; Xu, Lingyin; Lieber, Alexander M; Hamilton, Matthew; Brandt, Gabriel S

2017-12-01

The FIKK family of kinases is unique to parasites of the Apicomplexan order, which includes all malaria parasites. Plasmodium falciparum, the most virulent form of human malaria, has a family of 19 FIKK kinases, most of which are exported into the host red blood cell during malaria infection. Here, we confirm that FIKK 8 is a non-exported member of the FIKK kinase family. Through expression and purification of the recombinant kinase domain, we establish that emodin is a relatively high-affinity (IC 50 =2μM) inhibitor of PfFk8. Closely related anthraquinones do not inhibit PfFk8, suggesting that the particular substitution pattern of emodin is critical to the inhibitory pharmacophore. This first report of a P. falciparum FIKK kinase inhibitor lays the groundwork for developing specific inhibitors of the various members of the FIKK kinase family in order to probe their physiological function. Copyright © 2017 Elsevier Inc. All rights reserved.

Cytoplasmic proliferating cell nuclear antigen connects glycolysis and cell survival in acute myeloid leukemia.

PubMed

Ohayon, Delphine; De Chiara, Alessia; Chapuis, Nicolas; Candalh, Céline; Mocek, Julie; Ribeil, Jean-Antoine; Haddaoui, Lamya; Ifrah, Norbert; Hermine, Olivier; Bouillaud, Frédéric; Frachet, Philippe; Bouscary, Didier; Witko-Sarsat, Véronique

2016-10-19

Cytosolic proliferating cell nuclear antigen (PCNA), a scaffolding protein involved in DNA replication, has been described as a key element in survival of mature neutrophil granulocytes, which are non-proliferating cells. Herein, we demonstrated an active export of PCNA involved in cell survival and chemotherapy resistance. Notably, daunorubicin-resistant HL-60 cells (HL-60R) have a prominent cytosolic PCNA localization due to increased nuclear export compared to daunorubicin-sensitive HL-60 cells (HL-60S). By interacting with nicotinamide phosphoribosyltransferase (NAMPT), a protein involved in NAD biosynthesis, PCNA coordinates glycolysis and survival, especially in HL-60R cells. These cells showed a dramatic increase in intracellular NAD+ concentration as well as glycolysis including increased expression and activity of hexokinase 1 and increased lactate production. Furthermore, this functional activity of cytoplasmic PCNA was also demonstrated in patients with acute myeloid leukemia (AML). Our data uncover a novel pathway of nuclear export of PCNA that drives cell survival by increasing metabolism flux.

Cytoplasmic proliferating cell nuclear antigen connects glycolysis and cell survival in acute myeloid leukemia

PubMed Central

Ohayon, Delphine; De Chiara, Alessia; Chapuis, Nicolas; Candalh, Céline; Mocek, Julie; Ribeil, Jean-Antoine; Haddaoui, Lamya; Ifrah, Norbert; Hermine, Olivier; Bouillaud, Frédéric; Frachet, Philippe; Bouscary, Didier; Witko-Sarsat, Véronique

2016-01-01

Cytosolic proliferating cell nuclear antigen (PCNA), a scaffolding protein involved in DNA replication, has been described as a key element in survival of mature neutrophil granulocytes, which are non-proliferating cells. Herein, we demonstrated an active export of PCNA involved in cell survival and chemotherapy resistance. Notably, daunorubicin-resistant HL-60 cells (HL-60R) have a prominent cytosolic PCNA localization due to increased nuclear export compared to daunorubicin-sensitive HL-60 cells (HL-60S). By interacting with nicotinamide phosphoribosyltransferase (NAMPT), a protein involved in NAD biosynthesis, PCNA coordinates glycolysis and survival, especially in HL-60R cells. These cells showed a dramatic increase in intracellular NAD+ concentration as well as glycolysis including increased expression and activity of hexokinase 1 and increased lactate production. Furthermore, this functional activity of cytoplasmic PCNA was also demonstrated in patients with acute myeloid leukemia (AML). Our data uncover a novel pathway of nuclear export of PCNA that drives cell survival by increasing metabolism flux. PMID:27759041

How log-normal is your country? An analysis of the statistical distribution of the exported volumes of products

NASA Astrophysics Data System (ADS)

Annunziata, Mario Alberto; Petri, Alberto; Pontuale, Giorgio; Zaccaria, Andrea

2016-10-01

We have considered the statistical distributions of the volumes of 1131 products exported by 148 countries. We have found that the form of these distributions is not unique but heavily depends on the level of development of the nation, as expressed by macroeconomic indicators like GDP, GDP per capita, total export and a recently introduced measure for countries' economic complexity called fitness. We have identified three major classes: a) an incomplete log-normal shape, truncated on the left side, for the less developed countries, b) a complete log-normal, with a wider range of volumes, for nations characterized by intermediate economy, and c) a strongly asymmetric shape for countries with a high degree of development. Finally, the log-normality hypothesis has been checked for the distributions of all the 148 countries through different tests, Kolmogorov-Smirnov and Cramér-Von Mises, confirming that it cannot be rejected only for the countries of intermediate economy.

A light-driven proton pump from Haloterrigena turkmenica: Functional expression in Escherichia coli membrane and coupling with a H{sup +} co-transporter

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

Kamo, Naoki; Hashiba, Tsuyoshi; Kikukawa, Takashi

2006-03-10

A gene encoding putative retinal protein was cloned from Haloterrigena turkmenica (JCM9743). The deduced amino acid sequence was most closely related to that of deltarhodopsin, which functions as a light-driven H{sup +} pump and was identified in a novel strain Haloterrigena sp. arg-4 (K. Ihara, T. Uemura, I. Katagiri, T. Kitajima-Ihara, Y. Sugiyama, Y. Kimura, Y. Mukohata, Evolution of the archaeal rhodopsins: Evolution rate changes by gene duplication and functional differentiation, J. Mol. Biol. 285 (1999) 163-174. GenBank Accession No. AB009620). Thus, we called the present protein H. turkmenica deltarhodopsin (HtdR) in this report. Differing from the Halobacterium salinarum bacteriorhodopsinmore » (bR), functional expression of HtdR was achieved in Escherichia coli membrane with a high yield of 10-15mg protein/L culture. The photocycle of purified HtdR was similar to that of bR. The photo-induced electrogenic proton pumping activity of HtdR was verified. We co-expressed both HtdR and EmrE, a proton-coupled multi-drug efflux transporter in E. coli, and the cells successfully extruded ethidium, a substrate of EmrE, on illumination.« less

Increased Usage of Antiseptics Is Associated with Reduced Susceptibility in Clinical Isolates of Staphylococcus aureus.

PubMed

Hardy, Katherine; Sunnucks, Katie; Gil, Hannah; Shabir, Sahida; Trampari, Eleftheria; Hawkey, Peter; Webber, Mark

2018-05-29

Hospital-acquired infection is a major cause of morbidity and mortality, and regimes to prevent infection are crucial in infection control. These include the decolonization of vulnerable patients with methicillin-resistant Staphylococcus aureus (MRSA) carriage using antiseptics, including chlorhexidine and octenidine. Concern has been raised, however, regarding the possible development of biocide resistance. In this study, we assembled a panel of S. aureus isolates, including isolates collected before the development of chlorhexidine and octenidine and isolates, from a major hospital trust in the United Kingdom during a period when the decolonization regimes were altered. We observed significant increases in the MIC and minimum bactericidal concentration (MBC) of chlorhexidine in isolates from periods of high usage of chlorhexidine. Isolates with increased MICs and MBCs of octenidine rapidly emerged after octenidine was introduced in the trust. There was no apparent cross-resistance between the two biocidal agents. A combination of variable-number tandem repeat (VNTR) analysis, PCR for qac genes, and whole-genome sequencing was used to type isolates and examine possible mechanisms of resistance. There was no expansion of a single strain associated with decreased biocide tolerance, and biocide susceptibility did not correlate with carriage of qac efflux pump genes. Mutations within the NorA or NorB efflux pumps, previously associated with chlorhexidine export, were identified, however, suggesting that this may be an important mechanism of biocide tolerance. We present evidence that isolates are evolving in the face of biocide challenge in patients and that changes in decolonization regimes are reflected in changes in susceptibility of isolates. IMPORTANCE Infection in hospitals remains a major cause of death and disease. One way in which we combat this is by decolonizing at-risk patients from carriage of bacteria which can cause disease such as MRSA. This is done with antiseptics, including chlorhexidine and octenidine. There is concern, however, that bacteria may be able to become resistant to these antiseptics. In this study, we looked at isolates of MRSA and found that there was a correlation between the use of antiseptics and increased resistance in the isolates. We also suggest that the mechanism by which these more tolerant isolates may become resistant to antiseptics is that of changing a transport pump that exports these agents. This information suggests that we need to study the impact of antiseptics on clinically important bacteria more closely. Copyright © 2018 Hardy et al.

HIV-1 viral protein R (Vpr) induces fatty liver in mice via LXRα and PPARα dysregulation: implications for HIV-specific pathogenesis of NAFLD.

PubMed

Agarwal, Neeti; Iyer, Dinakar; Gabbi, Chiara; Saha, Pradip; Patel, Sanjeet G; Mo, Qianxing; Chang, Benny; Goswami, Biman; Schubert, Ulrich; Kopp, Jeffrey B; Lewis, Dorothy E; Balasubramanyam, Ashok

2017-10-17

HIV patients develop hepatic steatosis. We investigated hepatic steatosis in transgenic mice expressing the HIV-1 accessory protein Vpr (Vpr-Tg) in liver and adipose tissues, and WT mice infused with synthetic Vpr. Vpr-Tg mice developed increased liver triglyceride content and elevated ALT, bilirubin and alkaline phosphatase due to three hepatic defects: 1.6-fold accelerated de novo lipogenesis (DNL), 45% slower fatty acid ß-oxidation, and 40% decreased VLDL-triglyceride export. Accelerated hepatic DNL was due to coactivation by Vpr of liver X receptor-α (LXRα) with increased expression of its lipogenic targets Srebp1c, Chrebp, Lpk, Dgat, Fasn and Scd1, and intranuclear SREBP1c and ChREBP. Vpr enhanced association of LXRα with Lxrα and Srebp1c promoters, increased LXRE-LXRα binding, and broadly altered hepatic expression of LXRα-regulated lipid metabolic genes. Diminished hepatic fatty acid ß-oxidation was associated with decreased mRNA expression of Pparα and its targets Cpt1, Aox, Lcad, Ehhadh, Hsd10 and Acaa2, and blunted VLDL export with decreased expression of Mttp and its product microsomal triglyceride transfer protein. With our previous findings that Vpr circulates in HIV patients (including those with undetectable plasma HIV-1 RNA), co-regulates the glucocorticoid receptor and PPARγ and transduces hepatocytes, these data indicate a potential role for Vpr in HIV-associated fatty liver disease.

Comparative analysis of seven viral nuclear export signals (NESs) reveals the crucial role of nuclear export mediated by the third NES consensus sequence of nucleoprotein (NP) in influenza A virus replication.

PubMed

Chutiwitoonchai, Nopporn; Kakisaka, Michinori; Yamada, Kazunori; Aida, Yoko

2014-01-01

The assembly of influenza virus progeny virions requires machinery that exports viral genomic ribonucleoproteins from the cell nucleus. Currently, seven nuclear export signal (NES) consensus sequences have been identified in different viral proteins, including NS1, NS2, M1, and NP. The present study examined the roles of viral NES consensus sequences and their significance in terms of viral replication and nuclear export. Mutation of the NP-NES3 consensus sequence resulted in a failure to rescue viruses using a reverse genetics approach, whereas mutation of the NS2-NES1 and NS2-NES2 sequences led to a strong reduction in viral replication kinetics compared with the wild-type sequence. While the viral replication kinetics for other NES mutant viruses were also lower than those of the wild-type, the difference was not so marked. Immunofluorescence analysis after transient expression of NP-NES3, NS2-NES1, or NS2-NES2 proteins in host cells showed that they accumulated in the cell nucleus. These results suggest that the NP-NES3 consensus sequence is mostly required for viral replication. Therefore, each of the hydrophobic (Φ) residues within this NES consensus sequence (Φ1, Φ2, Φ3, or Φ4) was mutated, and its viral replication and nuclear export function were analyzed. No viruses harboring NP-NES3 Φ2 or Φ3 mutants could be rescued. Consistent with this, the NP-NES3 Φ2 and Φ3 mutants showed reduced binding affinity with CRM1 in a pull-down assay, and both accumulated in the cell nucleus. Indeed, a nuclear export assay revealed that these mutant proteins showed lower nuclear export activity than the wild-type protein. Moreover, the Φ2 and Φ3 residues (along with other Φ residues) within the NP-NES3 consensus were highly conserved among different influenza A viruses, including human, avian, and swine. Taken together, these results suggest that the Φ2 and Φ3 residues within the NP-NES3 protein are important for its nuclear export function during viral replication.

Na+/K+ ATPase regulates the expression and localization of acetylcholine receptors in a pump activity-independent manner

PubMed Central

Doi, Motomichi; Iwasaki, Kouichi

2008-01-01

Na+/K+ ATPase is a plasma membrane-localized sodium pump that maintains the ion gradients between the extracellular and intracellular environments, which in turn controls the cellular resting membrane potential. Recent evidence suggests that the pump is also localized at synapses and regulates synaptic efficacy. However, its precise function at the synapse is unknown. Here we show that two mutations in the α subunit of the eat-6 Na+/K+ ATPase in Caenorhabditis elegans dramatically increase the sensitivity to acetylcholine (Ach) agonists and alter the localization of nicotinic Ach receptors at the neuromuscular junction (NMJ). These defects can be rescued by mutated EAT-6 proteins which lack its pump activity, suggesting the presence of a novel function for Ach signaling. The Na+/K+ ATPase accumulates at postsynaptic sites and appears to surround Ach receptors to maintain rigid clusters at the NMJ. Our findings suggest a critical pump activity-independent, allele –specific role for Na+/K+ ATPase on postsynaptic organization and synaptic efficacy. PMID:18599311

Matter wave coupling of spatially separated and unequally pumped polariton condensates

NASA Astrophysics Data System (ADS)

Kalinin, Kirill P.; Lagoudakis, Pavlos G.; Berloff, Natalia G.

2018-03-01

Spatial quantum coherence between two separated driven-dissipative polariton condensates created nonresonantly and with a different occupation is studied. We identify the regions where the condensates remain coherent with the phase difference continuously changing with the pumping imbalance and the regions where each condensate acquires its own chemical potential with phase differences exhibiting time-dependent oscillations. We show that in the mutual coherence limit the coupling consists of two competing contributions: a symmetric Heisenberg exchange and the Dzyloshinskii-Moriya asymmetric interactions that enable a continuous tuning of the phase relation across the dyad and derive analytic expressions for these types of interactions. The introduction of nonequal pumping increases the complexity of the type of problems that can be solved by polariton condensates arranged in a graph configuration. If equally pumped polaritons condensates arrange their phases to solve the constrained quadratic minimisation problem with a real symmetric matrix, the nonequally pumped condensates solve that problem for a general Hermitian matrix.

Fluorescent Trimethoprim Conjugate Probes To Assess Drug Accumulation in Wild Type and Mutant Escherichia coli

PubMed Central

2016-01-01

Reduced susceptibility to antimicrobials in Gram-negative bacteria may result from multiple resistance mechanisms, including increased efflux pump activity or reduced porin protein expression. Up-regulation of the efflux pump system is closely associated with multidrug resistance (MDR). To help investigate the role of efflux pumps on compound accumulation, a fluorescence-based assay was developed using fluorescent derivatives of trimethoprim (TMP), a broad-spectrum synthetic antibiotic that inhibits an intracellular target, dihydrofolate reductase (DHFR). Novel fluorescent TMP probes inhibited eDHFR activity with comparable potency to TMP, but did not kill or inhibit growth of wild type Escherichia coli. However, bactericidal activity was observed against an efflux pump deficient E. coli mutant strain (ΔtolC). A simple and quick fluorescence assay was developed to measure cellular accumulation of the TMP probe using either fluorescence spectroscopy or flow cytometry, with validation by LC-MS/MS. This fluorescence assay may provide a simple method to assess efflux pump activity with standard laboratory equipment. PMID:27737551

Measuring Dependence on Imported Oil

EIA Publications

1995-01-01

U.S. dependence on imported oil can be measured in at least two ways. The differences hinge largely on whether oil imports are defined as net imports (total imports minus exports) or as total imports. EIA introduces a revised table that expresses dependence on imports in terms of both measures.

BRACEROS, MEXICO, AND FOREIGN TRADE.

ERIC Educational Resources Information Center

1966

CRITICISM OF GOVERNMENTAL POLICY RESTRICTING USE OF FOREIGN CONTRACT WORKERS ON AMERICAN FARMS IN 1965 EXPRESSES THREE POSSIBILITIES -- (1) A SEVERE DISRUPTION OF DOMESTIC FRUIT AND VEGETABLE PRODUCTION, (2) AN INCREASE OF AMERICAN INVESTMENT IN FOREIGN, PARTICULARLY MEXICAN, AGRICULTURAL PRODUCTION, AND (3) A LOSS OF UNITED STATES EXPORT MARKET…

Generation of high-order Hermite-Gaussian modes in end-pumped solid-state lasers for square vortex array laser beam generation.

PubMed

Chu, Shu-Chun; Chen, Yun-Ting; Tsai, Ko-Fan; Otsuka, Kenju

2012-03-26

This study reports the first systematic approach to the excitation of all high-order Hermite-Gaussian modes (HGMs) in end-pumped solid-state lasers. This study uses a metal-wire-inserted laser resonator accompanied with the "off axis pumping" approach. This study presents numerical analysis of the excitation of HGMs in end-pumped solid-state lasers and experimentally generated HGM patterns. This study also experimentally demonstrates the generation of an square vortex array laser beams by passing specific high-order HGMs (HGn,n + 1 or HGn + 1,n modes) through a Dove prism-embedded unbalanced Mach-Zehnder interferometer [Optics Express 16, 19934-19949]. The resulting square vortex array laser beams with embedded vortexes aligned in a square array can be applied to multi-spot dark optical traps in the future.

Persistent and High-Level Expression of Human Liver Prolidase in Vivo in Mice Using Adenovirus

DTIC Science & Technology

2013-01-01

types of nerve agents and pesticide compounds, is mostly exported into the circulation [11]. Similarly, human paraoxonase1, a promising enzyme in the...of human butyrylcholinesetrase results in persistent high-level transgene expression in vivo, Chem. Biol. Interact. 175 (2008) 327– 331. [11] K...paraoxonase1 gene transfer to provide protection against the toxicity of the organophosphorus pesticide toxicant diazoxon, Gene Ther. 18 (2011) 250–257. [14

The role of nutricline depth in regulating the ocean carbon cycle.

PubMed

Cermeño, Pedro; Dutkiewicz, Stephanie; Harris, Roger P; Follows, Mick; Schofield, Oscar; Falkowski, Paul G

2008-12-23

Carbon uptake by marine phytoplankton, and its export as organic matter to the ocean interior (i.e., the "biological pump"), lowers the partial pressure of carbon dioxide (pCO(2)) in the upper ocean and facilitates the diffusive drawdown of atmospheric CO(2). Conversely, precipitation of calcium carbonate by marine planktonic calcifiers such as coccolithophorids increases pCO(2) and promotes its outgassing (i.e., the "alkalinity pump"). Over the past approximately 100 million years, these two carbon fluxes have been modulated by the relative abundance of diatoms and coccolithophores, resulting in biological feedback on atmospheric CO(2) and Earth's climate; yet, the processes determining the relative distribution of these two phytoplankton taxa remain poorly understood. We analyzed phytoplankton community composition in the Atlantic Ocean and show that the distribution of diatoms and coccolithophorids is correlated with the nutricline depth, a proxy of nutrient supply to the upper mixed layer of the ocean. Using this analysis in conjunction with a coupled atmosphere-ocean intermediate complexity model, we predict a dramatic reduction in the nutrient supply to the euphotic layer in the coming century as a result of increased thermal stratification. Our findings indicate that, by altering phytoplankton community composition, this causal relationship may lead to a decreased efficiency of the biological pump in sequestering atmospheric CO(2), implying a positive feedback in the climate system. These results provide a mechanistic basis for understanding the connection between upper ocean dynamics, the calcium carbonate-to-organic C production ratio and atmospheric pCO(2) variations on time scales ranging from seasonal cycles to geological transitions.

Evaluating Southern Ocean Carbon Eddy-Pump From Biogeochemical-Argo Floats

NASA Astrophysics Data System (ADS)

Llort, Joan; Langlais, C.; Matear, R.; Moreau, S.; Lenton, A.; Strutton, Peter G.

2018-02-01

The vertical transport of surface water and carbon into ocean's interior, known as subduction, is one of the main mechanisms through which the ocean influences Earth's climate. New instrumental approaches have shown the occurrence of localized and intermittent subduction episodes associated with small-scale ocean circulation features. These studies also revealed the importance of such events for the export of organic matter, the so-called eddy-pump. However, the transient and localized nature of episodic subduction hindered its large-scale evaluation to date. In this work, we present an approach to detect subduction events at the scale of the Southern Ocean using measurements collected by biogeochemical autonomous floats (BGCArgo). We show how subduction events can be automatically identified as anomalies of spiciness and Apparent Oxygen Utilization (AOU) below the mixed layer. Using this methodology over more than 4,000 profiles, we detected 40 subduction events unevenly distributed across the Sothern Ocean. Events were more likely found in hot spots of eddy kinetic energy (EKE), downstream major bathymetric features. Moreover, the bio-optical measurements provided by BGCArgo allowed measuring the amount of Particulate Organic Carbon (POC) being subducted and assessing the contribution of these events to the total downward carbon flux at 100 m (EP100). We estimated that the eddy-pump represents less than 19% to the EP100 in the Southern Ocean, although we observed particularly strong events able to locally duplicate the EP100. This approach provides a novel perspective on where episodic subduction occurs that will be naturally improved as BGCArgo observations continue to increase.

Oceanic acidification affects marine carbon pump and triggers extended marine oxygen holes.

PubMed

Hofmann, Matthias; Schellnhuber, Hans-Joachim

2009-03-03

Rising atmospheric CO(2) levels will not only drive future global mean temperatures toward values unprecedented during the whole Quaternary but will also lead to massive acidification of sea water. This constitutes by itself an anthropogenic planetary-scale perturbation that could significantly modify oceanic biogeochemical fluxes and severely damage marine biota. As a step toward the quantification of such potential impacts, we present here a simulation-model-based assessment of the respective consequences of a business-as-usual fossil-fuel-burning scenario where a total of 4,075 Petagrams of carbon is released into the atmosphere during the current millennium. In our scenario, the atmospheric pCO(2) level peaks at approximately 1,750 microatm in the year 2200 while the sea-surface pH value drops by >0.7 units on global average, inhibiting the growth of marine calcifying organisms. The study focuses on quantifying 3 major concomitant effects. The first one is a significant (climate-stabilizing) negative feedback on rising pCO(2) levels as caused by the attenuation of biogenic calcification. The second one is related to the biological carbon pump. Because mineral ballast, notably CaCO(3), is found to play a dominant role in carrying organic matter through the water column, a reduction of its export fluxes weakens the strength of the biological carbon pump. There is, however, a third effect with severe consequences: Because organic matter is oxidized in shallow waters when mineral-ballast fluxes weaken, oxygen holes (hypoxic zones) start to expand considerably in the oceans in our model world--with potentially harmful impacts on a variety of marine ecosystems.

Discovery of a Siderophore Export System Essential for Virulence of Mycobacterium tuberculosis

PubMed Central

Wells, Ryan M.; Jones, Christopher M.; Xi, Zhaoyong; Speer, Alexander; Danilchanka, Olga; Doornbos, Kathryn S.; Sun, Peibei; Wu, Fangming; Tian, Changlin; Niederweis, Michael

2013-01-01

Iron is an essential nutrient for most bacterial pathogens, but is restricted by the host immune system. Mycobacterium tuberculosis (Mtb) utilizes two classes of small molecules, mycobactins and carboxymycobactins, to capture iron from the human host. Here, we show that an Mtb mutant lacking the mmpS4 and mmpS5 genes did not grow under low iron conditions. A cytoplasmic iron reporter indicated that the double mutant experienced iron starvation even under high-iron conditions. Loss of mmpS4 and mmpS5 did not change uptake of carboxymycobactin by Mtb. Thin layer chromatography showed that the ΔmmpS4/S5 mutant was strongly impaired in biosynthesis and secretion of siderophores. Pull-down experiments with purified proteins demonstrated that MmpS4 binds to a periplasmic loop of the associated transporter protein MmpL4. This interaction was corroborated by genetic experiments. While MmpS5 interacted only with MmpL5, MmpS4 interacted with both MmpL4 and MmpL5. These results identified MmpS4/MmpL4 and MmpS5/MmpL5 as siderophore export systems in Mtb and revealed that the MmpL proteins transport small molecules other than lipids. MmpS4 and MmpS5 resemble periplasmic adapter proteins of tripartite efflux pumps of Gram-negative bacteria, however, they are not only required for export but also for efficient siderophore synthesis. Membrane association of MbtG suggests a link between siderophore synthesis and transport. The structure of the soluble domain of MmpS4 (residues 52–140) was solved by NMR and indicates that mycobacterial MmpS proteins constitute a novel class of transport accessory proteins. The bacterial burden of the mmpS4/S5 deletion mutant in mouse lungs was lower by 10,000-fold and none of the infected mice died within 180 days compared to wild-type Mtb. This is the strongest attenuation observed so far for Mtb mutants lacking genes involved in iron utilization. In conclusion, this study identified the first components of novel siderophore export systems which are essential for virulence of Mtb. PMID:23431276

Climate Change Response of Ocean Net Primary Production (NPP) and Export Production (EP) Regulated by Stratification Increases in The CMIP5 models

NASA Astrophysics Data System (ADS)

Fu, W.; Randerson, J. T.; Moore, J. K.

2014-12-01

Ocean warming due to rising atmospheric CO2 has increasing impacts on ocean ecosystems by modifying the ecophysiology and distribution of marine organisms, and by altering ocean circulation and stratification. We explore ocean NPP and EP changes at the global scale with simulations performed in the framework of the fifth Coupled Model Inter-comparison Project (CMIP5). Global NPP and EP are reduced considerably by the end of the century for the representative concentration pathway (RCP) 8.5 scenario, although models differ in their significantly in their direct temperature impacts on production and remineralization. The Earth system models used here project similar NPP trends albeit the magnitudes vary substantially. In general, projected changes in the 2090s for NPP range between -2.3 to -16.2% while export production reach -7 to -18% relative to 1990s. This is accompanied by increased stratification by 17-30%. Results indicate that globally reduced NPP is closely related to increased ocean stratification (R2=0.78). This is especially the case for global export production, that seems to be mostly controlled by the increased stratification (R2=0.95). We also identify phytoplankton community impacts on these patterns, that vary across the models. The negative response of NPP to climate change may be through bottom-up control, leading to a reduced capacity of oceans to regulate climate through the biological carbon pump. There are large disagreements among the CMIP5 models in terms of simulated nutrient and oxygen concentrations for the 1990s, and their trends over time with climate change. In addition, potentially important marine biogeochemical feedbacks on the climate system were not well represented in the CMIP5 models, including important feedbacks with aerosol deposition and the marine iron cycle, and feedbacks involving the oxygen minimum zones and the marine nitrogen cycle. Thus, these substantial reductions in primary productivity and export production over the 21st century simulated under the RCP 8.5 scenario were likely conservative estimates, and may need to be revised as marine biogeochemistry in Earth System Models (ESMs) continues to be developed.

Biophysical characterization of the outer membrane polysaccharide export protein and the polysaccharide co-polymerase protein from Xanthomonas campestris.

PubMed

Bianco, M I; Jacobs, M; Salinas, S R; Salvay, A G; Ielmini, M V; Ielpi, L

2014-09-01

This study investigated the structural and biophysical characteristics of GumB and GumC, two Xanthomonas campestris membrane proteins that are involved in xanthan biosynthesis. Xanthan is an exopolysaccharide that is thought to be a virulence factor that contributes to bacterial in planta growth. It also is one of the most important industrial biopolymers. The first steps of xanthan biosynthesis are well understood, but the polymerization and export mechanisms remain unclear. For this reason, the key proteins must be characterized to better understand these processes. Here we characterized, by biochemical and biophysical techniques, GumB, the outer membrane polysaccharide export protein, and GumC, the polysaccharide co-polymerase protein of the xanthan biosynthesis system. Our results suggested that recombinant GumB is a tetrameric protein in solution. On the other hand, we observed that both native and recombinant GumC present oligomeric conformation consistent with dimers and higher-order oligomers. The transmembrane segments of GumC are required for GumC expression and/or stability. These initial results provide a starting point for additional studies that will clarify the roles of GumB and GumC in the xanthan polymerization and export processes and further elucidate their functions and mechanisms of action. Copyright © 2014 Elsevier Inc. All rights reserved.

Dental Items of Significance, Number 43

DTIC Science & Technology

1994-08-15

The dental ofice versions, on the other hand, commonly include a resin cement and dentin bonding agent to faciitate the clinical aspect of...resin bonding procedures. The recall was ordered because of incidents In which the atchant gel was inadvertently expressed out of the barrel of the...iterally "gum up the wash water recirculating pump and cause the unit to drain slowly. Wash agent pumps are a convenience feature but also serve the

A Separate Pool of Cardiac Phospholemman That Does Not Regulate or Associate with the Sodium Pump

PubMed Central

Wypijewski, Krzysztof J.; Howie, Jacqueline; Reilly, Louise; Tulloch, Lindsay B.; Aughton, Karen L.; McLatchie, Linda M.; Shattock, Michael J.; Calaghan, Sarah C.; Fuller, William

2013-01-01

Phospholemman (PLM), the principal quantitative sarcolemmal substrate for protein kinases A and C in the heart, regulates the cardiac sodium pump. Much like phospholamban, which regulates the related ATPase SERCA, PLM is reported to oligomerize. We investigated subpopulations of PLM in adult rat ventricular myocytes based on phosphorylation status. Co-immunoprecipitation identified two pools of PLM: one not associated with the sodium pump phosphorylated at Ser63 and one associated with the pump, both phosphorylated at Ser68 and unphosphorylated. Phosphorylation of PLM at Ser63 following activation of PKC did not abrogate association of PLM with the pump, so its failure to associate with the pump was not due to phosphorylation at this site. All pools of PLM co-localized to cell surface caveolin-enriched microdomains with sodium pump α subunits, despite the lack of caveolin-binding motif in PLM. Mass spectrometry analysis of phosphospecific immunoprecipitation reactions revealed no unique protein interactions for Ser63-phosphorylated PLM, and cross-linking reagents also failed to identify any partner proteins for this pool. In lysates from hearts of heterozygous transgenic animals expressing wild type and unphosphorylatable PLM, Ser63-phosphorylated PLM co-immunoprecipitated unphosphorylatable PLM, confirming the existence of PLM multimers. Dephosphorylation of the PLM multimer does not change sodium pump activity. Hence like phospholamban, PLM exists as a pump-inhibiting monomer and an unassociated oligomer. The distribution of different PLM phosphorylation states to different pools may be explained by their differential proximity to protein phosphatases rather than a direct effect of phosphorylation on PLM association with the pump. PMID:23532852

Late summer particulate organic carbon export and twilight zone remineralisation in the Atlantic sector of the Southern Ocean

NASA Astrophysics Data System (ADS)

Planchon, F.; Cavagna, A.-J.; Cardinal, D.; André, L.; Dehairs, F.

2012-03-01

During the Bonus-GoodHope (BGH) expedition (Jan-Mar 2008) we studied the water column distribution of total 234Th and biogenic particulate Ba (Baxs) in the Atlantic sector of the Southern Ocean. The objective was to assess the export flux of particulate organic carbon (POC) from the surface to the mesopelagic twilight zone along a section between the Cape Basin and Weddell Gyre. Export production of POC was estimated from steady state and non steady state export fluxes of 234Th which were converted into POC fluxes, using the POC/234Th ratio of large (>53 μm) suspended particles, collected via in-situ pumps. Deficits in 234Th activities were observed at all stations from the surface to the bottom of the mixed-layer. 234Th export fluxes from the upper 100 m ranged from 496 ± 57 dpm m-2 d-1 to 1195 ± 120 dpm m-2 d-1 for the steady state model and from 149 ± 18 dpm m-2 d-1 to 1217 ± 146 dpm m-2 d-1 for the non steady state model calculated for a time window of 15 to 22 days preceding the timing of the present cruise. The POC/234Thp ratio of large, potentially sinking particles (>53 μm), was observed to increase with latitude, from 1.9 ± 0.2 μmol dpm-1 and 1.7 ± 0.3 μmol dpm-1 in the Subtropical Zone (STZ) and Subantarctic Zone (SAZ), respectively, to 3.0 ± 0.2 μmol dpm-1 in the Polar Front Zone (PFZ), 4.8 ± 1.9 μmol dpm-1 at the Southern Antarctic Circumpolar Current Front (SACCF) to 4.1 ± 1.7 μmol dpm-1 in the northern Weddell Gyre, in line with an increasing contribution of larger cell diatoms. Steady state and non steady state POC export from the upper 100 m ranged from 0.9 ± 0.2 mmolC m-2 d-1 to 5.1 ± 2.1 mmolC m-2 d-1 and from 0.3 ± 0.0 mmolC m-2 d-1 to 4.9 ± 3.2 mmolC m-2 d-1, respectively. From the SAZ to the SACCF, non steady state POC export production represented only 15 to 54 % of the steady state POC flux, suggesting that the intensity of export had decreased over time partly due to the fact that regenerated-production based communities of small-sized phytoplankton became predominant. In contrast, for the HNLC area south of the SACCF, we found an excellent agreement between the two modeling approaches indicating that surface POC export remained rather constant there. Estimated POC export represented between 6 to 54 % of the potential export as represented by new production, indicating that export efficiency was particularly low throughout the studied area, except close to the SACCF. Below the export layer, in the mesopelagic zone, 234Th activities generally reached equilibrium with 238U, but sometimes were in large excess relative to 238U (234Th/238U ratio>1.1), reflecting intense remineralisation/disaggregation of 234Th-bearing particles. The accumulation of excess 234Th in the 100-600 m depth interval ranged from 458 ± 55 dpm m-2 d-1 to 3068 ± 368 dpm m-2 d-1. Using POC to 234Th ratio of sinking particles, we converted this 234Th flux into a POC remineralisation flux, which ranged between 0.9 to 9.2 mmolC m-2 d-1. Mesopelagic remineralisation was also evidenced by Baxs inventories which are related to bacterial degradation of sinking material and offer a means to quantify the flux of respired C. Highest biogenic particulate Ba (Baxs) contents were generally observed in the 200-400 m depth interval with values reaching up to >1000 pM in the northern PFZ. Depth weighted average mesopelagic Baxs (meso-Baxs) was high in the PFZ and low in the northernmost (STZ-SAZ) and the southernmost (SACCF-AZ-WG) parts of the BGH section; conversion into respired C flux yielded a range of -0.23 to 6.4 mmolC m-2 d-1. Excluding two outliers, we found a significant positive correlation for mesopelagic waters between POC remineralisation estimated from meso-Baxs and from 234Th excess (R2 = 0.73). Our results indicate that POC export production in this area of the Southern Ocean was strongly attenuated in the mesopelagic waters due to remineralisation, a process which thus appears to strongly impact on longer term bathypelagic zone sequestration of POC.

Phosphorylation of rat kidney Na-K pump at Ser938 is required for rapid angiotensin II-dependent stimulation of activity and trafficking in proximal tubule cells

PubMed Central

Massey, Katherine J.; Li, Quanwen; Rossi, Noreen F.; Keezer, Susan M.; Mattingly, Raymond R.

2015-01-01

How angiotensin (ANG) II acutely stimulates the Na-K pump in proximal tubules is only partially understood, limiting insight into how ANG II increases blood pressure. First, we tested whether ANG II increases the number of pumps in plasma membranes of native rat proximal tubules under conditions of rapid activation. We found that exposure to 100 pM ANG II for 2 min, which was previously shown to increase affinity of the Na-K pump for Na and stimulate activity threefold, increased the amount of the Na-K pump in plasma membranes of native tubules by 33%. Second, we tested whether previously observed increases in phosphorylation of the Na-K pump at Ser938 were part of the stimulatory mechanism. These experiments were carried out in opossum kidney cells, cultured proximal tubules stably coexpressing the ANG type 1 (AT1) receptor, and either wild-type or a S938A mutant of rat kidney Na-K pump under conditions found by others to stimulate activity. We found that 10 min of incubation in 10 pM ANG II stimulated activity of wild-type pumps from 2.3 to 3.5 nmol K·mg protein−1·min−1 and increased the amount of the pump in the plasma membrane by 80% but had no effect on cells expressing the S938A mutant. We conclude that acute stimulation of Na-K pump activity in native rat proximal tubules includes increased trafficking to the plasma membrane and that phosphorylation at Ser938 is part of the mechanism by which ANG II directly stimulates activity and trafficking of the rat kidney Na-K pump in opossum kidney cells. PMID:26582472

Suppression of the endoplasmic reticulum calcium pump during zebrafish gastrulation affects left-right asymmetry of the heart and brain.

PubMed

Kreiling, Jill A; Balantac, Zaneta L; Crawford, Andrew R; Ren, Yuexin; Toure, Jamal; Zchut, Sigalit; Kochilas, Lazaros; Creton, Robbert

2008-01-01

Vertebrate embryos generate striking Ca(2+) patterns, which are unique regulators of dynamic developmental events. In the present study, we used zebrafish embryos as a model system to examine the developmental roles of Ca(2+) during gastrulation. We found that gastrula stage embryos maintain a distinct pattern of cytosolic Ca(2+) along the dorsal-ventral axis, with higher Ca(2+) concentrations in the ventral margin and lower Ca(2+) concentrations in the dorsal margin and dorsal forerunner cells. Suppression of the endoplasmic reticulum Ca(2+) pump with 0.5 microM thapsigargin elevates cytosolic Ca(2+) in all embryonic regions and induces a randomization of laterality in the heart and brain. Affected hearts, visualized in living embryos by a subtractive imaging technique, displayed either a reversal or loss of left-right asymmetry. Brain defects include a left-right reversal of pitx2 expression in the dorsal diencephalon and a left-right reversal of the prominent habenular nucleus in the brain. Embryos are sensitive to inhibition of the endoplasmic reticulum Ca(2+) pump during early and mid gastrulation and lose their sensitivity during late gastrulation and early segmentation. Suppression of the endoplasmic reticulum Ca(2+) pump during gastrulation inhibits expression of no tail (ntl) and left-right dynein related (lrdr) in the dorsal forerunner cells and affects development of Kupffer's vesicle, a ciliated organ that generates a counter-clockwise flow of fluid. Previous studies have shown that Ca(2+) plays a role in Kupffer's vesicle function, influencing ciliary motility and translating the vesicle's counter-clockwise flow into asymmetric patterns of gene expression. The present results suggest that Ca(2+) plays an additional role in the formation of Kupffer's vesicle.

HvHMA2, a P1B-ATPase from Barley, Is Highly Conserved among Cereals and Functions in Zn and Cd Transport

PubMed Central

Mills, Rebecca F.; Peaston, Kerry A.; Runions, John; Williams, Lorraine E.

2012-01-01

Manipulation of crops to improve their nutritional value (biofortification) and optimisation of plants for removal of toxic metals from contaminated soils (phytoremediation) are major goals. Identification of membrane transporters with roles in zinc and cadmium transport would be useful for both aspects. The P1B-ATPases play important roles in heavy metal allocation and detoxification in Arabidopsis and it is now important to elucidate their roles in monocots. We identified nine P1B-ATPases in barley and this study focuses on the functional characterization of HvHMA2, providing evidence for its role in heavy metal transport. HvHMA2 was cloned using information from EST analysis and 5′ RACE. It possesses the conserved aspartate that is phosphorylated during the reaction cycle of P-type pumps and has motifs and key residues characteristic of P1B-ATPases, falling into the P1B-2 subclass. Homologous sequences occur in three major sub-families of the Poaceae (Gramineae). Heterologous expression in Saccharomyces cerevisiae demonstrates that HvHMA2 functions as a Zn and Cd pump. Mutagenesis studies show that proposed cation coordination sites of the P1B-2 pumps are crucial for the metal responses conferred by HvHMA2 in yeast. HvHMA2 expression suppresses the Zn-deficient phenotype of the Arabidopsis hma2hma4 mutant indicating that HvHMA2 functions as a Zn pump in planta and could play a role in root to shoot Zn transport. When expressed in Arabidopsis, HvHMA2 localises predominantly to the plasma membrane. PMID:22880063

Can the HIV-1 splicing machinery be targeted for drug discovery?

PubMed Central

Dlamini, Zodwa; Hull, Rodney

2017-01-01

HIV-1 is able to express multiple protein types and isoforms from a single 9 kb mRNA transcript. These proteins are also expressed at particular stages of viral development, and this is achieved through the control of alternative splicing and the export of these transcripts from the nucleus. The nuclear export is controlled by the HIV protein Rev being required to transport incompletely spliced and partially spliced mRNA from the nucleus where they are normally retained. This implies a close relationship between the control of alternate splicing and the nuclear export of mRNA in the control of HIV-1 viral proliferation. This review discusses both the processes. The specificity and regulation of splicing in HIV-1 is controlled by the use of specific splice sites as well as exonic splicing enhancer and exonic splicing silencer sequences. The use of these silencer and enhancer sequences is dependent on the serine arginine family of proteins as well as the heterogeneous nuclear ribonucleoprotein family of proteins that bind to these sequences and increase or decrease splicing. Since alternative splicing is such a critical factor in viral development, it presents itself as a promising drug target. This review aims to discuss the inhibition of splicing, which would stall viral development, as an anti-HIV therapeutic strategy. In this review, the most recent knowledge of splicing in human immunodeficiency viral development and the latest therapeutic strategies targeting human immunodeficiency viral splicing are discussed. PMID:28331370

A Simple Method for Assessment of MDR Bacteria for Over-Expressed Efflux Pumps

PubMed Central

Martins, Marta; McCusker, Matthew P; Viveiros, Miguel; Couto, Isabel; Fanning, Séamus; Pagès, Jean-Marie; Amaral, Leonard

2013-01-01

It is known that bacteria showing a multi-drug resistance phenotype use several mechanisms to overcome the action of antibiotics. As a result, this phenotype can be a result of several mechanisms or a combination of thereof. The main mechanisms of antibiotic resistance are: mutations in target genes (such as DNA gyrase and topoisomerase IV); over-expression of efflux pumps; changes in the cell envelope; down regulation of membrane porins, and modified lipopolysaccharide component of the outer cell membrane (in the case of Gram-negative bacteria). In addition, adaptation to the environment, such as quorum sensing and biofilm formation can also contribute to bacterial persistence. Due to the rapid emergence and spread of bacterial isolates showing resistance to several classes of antibiotics, methods that can rapidly and efficiently identify isolates whose resistance is due to active efflux have been developed. However, there is still a need for faster and more accurate methodologies. Conventional methods that evaluate bacterial efflux pump activity in liquid systems are available. However, these methods usually use common efflux pump substrates, such as ethidium bromide or radioactive antibiotics and therefore, require specialized instrumentation, which is not available in all laboratories. In this review, we will report the results obtained with the Ethidium Bromide-agar Cartwheel method. This is an easy, instrument-free, agar based method that has been modified to afford the simultaneous evaluation of as many as twelve bacterial strains. Due to its simplicity it can be applied to large collections of bacteria to rapidly screen for multi-drug resistant isolates that show an over-expression of their efflux systems. The principle of the method is simple and relies on the ability of the bacteria to expel a fluorescent molecule that is substrate for most efflux pumps, ethidium bromide. In this approach, the higher the concentration of ethidium bromide required to produce fluorescence of the bacterial mass, the greater the efflux capacity of the bacterial cells. We have tested and applied this method to a large number of Gram-positive and Gram-negative bacteria to detect efflux activity among these multi-drug resistant isolates. The presumptive efflux activity detected by the Ethidium Bromide-agar Cartwheel method was subsequently confirmed by the determination of the minimum inhibitory concentration for several antibiotics in the presence and absence of known efflux pump inhibitors. PMID:23589748

2,2'-Dihydroxychalcone, a glutathione transferase inhibitor, sensitises human colon adenocarcinoma cells to chlorambucil and melphalan, but not to actinomycin D.

PubMed

Goh, Kenneth; Chen, Yufan; Zheng, Lin; Ong, Laichun; Jin, Yi; Chow, Pierce; Zhang, Kai

2008-01-01

2,2'-Dihydroxychalcone (2,2'DHC) is a potent inhibitor of glutathione S-transferases (GSTs). Pre-treatment of human colon cancer cells by a non-toxic concentration of this GST inhibitor significantly sensitised cancer cells to chlorambucil and melphalan, which are substrates of glutathione (GSH) conjugation. However, sensitisation to actinomycin D, which has not been shown to be detoxified by GSH-related mechanisms, was not observed. These results further confirm the contribution of GSH-related mechanisms to drug resistance by increased detoxification of drugs. 2,2'DHC inhibited GST activity and the transport of GSH conjugates by cancer cells. Its combined effects on GST and glutathione conjugate export (GS-X) pump may provide more potent sensitisation of cancer cells to chemotherapeutic drugs.

Cesium alignment produced by pumping with unpolarized light★

NASA Astrophysics Data System (ADS)

Shi, Yongqi; Weis, Antoine

2018-04-01

We demonstrate optical pumping on the four hyperfine components of the Cs D 1 transition by unpolarized (UPL) resonant laser light. The evidence is based on the reduction of the absorption coefficients κ 0 with increasing light power P in an uncoated Cs vapor cell with isotropic spin relaxation. For comparison we perform the same quantitative κ 0( P) measurements with linearly-polarized light (LPL) and circularly-polarized light (CPL). We find that our previously published algebraic expressions give an excellent description of all experimentally recorded induced transparency signals. Based on this we can make reliable absolute predictions for the power dependence of the spin orientation and alignment produced by pumping with LPL, CPL and UPL.

An N-Terminal ER Export Signal Facilitates the Plasma Membrane Targeting of HCN1 Channels in Photoreceptors.

PubMed

Pan, Yuan; Laird, Joseph G; Yamaguchi, David M; Baker, Sheila A

2015-06-01

Hyperpolarization-activated cyclic nucleotide-gated 1 (HCN1) channels are widely expressed in the retina. In photoreceptors, the hyperpolarization-activated current (Ih) carried by HCN1 is important for shaping the light response. It has been shown in multiple systems that trafficking HCN1 channels to specific compartments is key to their function. The localization of HCN1 in photoreceptors is concentrated in the plasma membrane of the inner segment (IS). The mechanisms controlling this localization are not understood. We previously identified a di-arginine endoplasmic reticulum (ER) retention motif that negatively regulates the surface targeting of HCN1. In this study, we sought to identify a forward trafficking signal that could counter the function of the ER retention signal. We studied trafficking of HCN1 and several mutants by imaging their subcellular localization in transgenic X. laevis photoreceptors. Velocity sedimentation was used to assay the assembly state of HCN1 channels. We found the HCN1 N-terminus can redirect a membrane reporter from outer segments (OS) to the plasma membrane of the IS. The sequence necessary for this behavior was mapped to a 20 amino acid region containing a leucine-based ER export motif. The ER export signal is necessary for forward trafficking but not channel oligomerization. Moreover, this ER export signal alone counteracted the di-arginine ER retention signal. We identified an ER export signal in HCN1 that functions with the ER retention signal to maintain equilibrium of HCN1 between the endomembrane system and the plasma membrane.

An N-Terminal ER Export Signal Facilitates the Plasma Membrane Targeting of HCN1 Channels in Photoreceptors

PubMed Central

Pan, Yuan; Laird, Joseph G.; Yamaguchi, David M.; Baker, Sheila A.

2015-01-01

Purpose. Hyperpolarization-activated cyclic nucleotide-gated 1 (HCN1) channels are widely expressed in the retina. In photoreceptors, the hyperpolarization-activated current (Ih) carried by HCN1 is important for shaping the light response. It has been shown in multiple systems that trafficking HCN1 channels to specific compartments is key to their function. The localization of HCN1 in photoreceptors is concentrated in the plasma membrane of the inner segment (IS). The mechanisms controlling this localization are not understood. We previously identified a di-arginine endoplasmic reticulum (ER) retention motif that negatively regulates the surface targeting of HCN1. In this study, we sought to identify a forward trafficking signal that could counter the function of the ER retention signal. Methods. We studied trafficking of HCN1 and several mutants by imaging their subcellular localization in transgenic X. laevis photoreceptors. Velocity sedimentation was used to assay the assembly state of HCN1 channels. Results. We found the HCN1 N-terminus can redirect a membrane reporter from outer segments (OS) to the plasma membrane of the IS. The sequence necessary for this behavior was mapped to a 20 amino acid region containing a leucine-based ER export motif. The ER export signal is necessary for forward trafficking but not channel oligomerization. Moreover, this ER export signal alone counteracted the di-arginine ER retention signal. Conclusions. We identified an ER export signal in HCN1 that functions with the ER retention signal to maintain equilibrium of HCN1 between the endomembrane system and the plasma membrane. PMID:26030105

Adiabatic pumping solutions in global AdS

NASA Astrophysics Data System (ADS)

Carracedo, Pablo; Mas, Javier; Musso, Daniele; Serantes, Alexandre

2017-05-01

We construct a family of very simple stationary solutions to gravity coupled to a massless scalar field in global AdS. They involve a constantly rising source for the scalar field at the boundary and thereby we name them pumping solutions. We construct them numerically in D = 4. They are regular and, generically, have negative mass. We perform a study of linear and nonlinear stability and find both stable and unstable branches. In the latter case, solutions belonging to different sub-branches can either decay to black holes or to limiting cycles. This observation motivates the search for non-stationary exactly timeperiodic solutions which we actually construct. We clarify the role of pumping solutions in the context of quasistatic adiabatic quenches. In D = 3 the pumping solutions can be related to other previously known solutions, like magnetic or translationally-breaking backgrounds. From this we derive an analytic expression.

Pumping of magnons in a Dzyaloshinskii-Moriya ferromagnet

NASA Astrophysics Data System (ADS)

Kovalev, Alexey A.; Zyuzin, Vladimir A.; Li, Bo

2017-04-01

We formulate a microscopic linear response theory of magnon pumping applicable to multiple-magnonic-band uniform ferromagnets with Dzyaloshinskii-Moriya interactions. From the linear response theory, we identify the extrinsic and intrinsic contributions where the latter is expressed via the Berry curvature of magnonic bands. We observe that in the presence of a time-dependent magnetization Dzyaloshinskii-Moriya interactions can act as fictitious electric fields acting on magnons. We study various current responses to this fictitious field and analyze the role of Berry curvature. In particular, we obtain an analog of the Hall-like response in systems with nontrivial Berry curvature of magnon bands. After identifying the magnon-mediated contribution to the equilibrium Dzyaloshinskii-Moriya interaction, we also establish the Onsager reciprocity between the magnon mediated thermal torques and heat pumping. We apply our theory to the magnonic heat pumping and torque responses in honeycomb and kagome lattice ferromagnets.

The H,K-ATPase beta-subunit can act as a surrogate for the beta-subunit of Na,K-pumps.

PubMed

Horisberger, J D; Jaunin, P; Reuben, M A; Lasater, L S; Chow, D C; Forte, J G; Sachs, G; Rossier, B C; Geering, K

1991-10-15

Na,K-ATPase and H,K-ATPase are the only members of the P-type ATPases in which a glycosylated beta-subunit is part of the purified active enzyme. In this study, we have followed the synthesis and the posttranslational processing of the beta-subunit of H,K-ATPase (beta HK) in Xenopus oocytes injected with beta HK cRNA and have tested whether it can act as a surrogate for the beta-subunit of Na,K-ATPase (beta NaK) to support the functional expression of Na,K-pumps. In Xenopus oocytes, beta HK is processed from an Endo H-sensitive 51-kDa coreglycosylated form to an Endo H-resistant 71-kDa fully glycosylated form. Similar to beta NaK, beta HK can stabilize and increase the trypsin resistance of alpha-subunits of Na,K-ATPase (alpha NaK). Finally, expression of beta HK together with alpha NaK leads to an increased number of ouabain binding sites at the plasma membrane accompanied by an increased Rb+ uptake and Na,K-pump current. Our data suggest that beta HK, similar to beta NaK, can assemble to alpha NaK, support the structural maturation and the intracellular transport of catalytic alpha NaK, and ultimately form active alpha NaK-beta HK complexes with Na,K-pump transport properties.

Non-equilibrium thermodynamical description of rhythmic motion patterns of active systems: a canonical-dissipative approach.

PubMed

Dotov, D G; Kim, S; Frank, T D

2015-02-01

We derive explicit expressions for the non-equilibrium thermodynamical variables of a canonical-dissipative limit cycle oscillator describing rhythmic motion patterns of active systems. These variables are statistical entropy, non-equilibrium internal energy, and non-equilibrium free energy. In particular, the expression for the non-equilibrium free energy is derived as a function of a suitable control parameter. The control parameter determines the Hopf bifurcation point of the deterministic active system and describes the effective pumping of the oscillator. In analogy to the equilibrium free energy of the Landau theory, it is shown that the non-equilibrium free energy decays as a function of the control parameter. In doing so, a similarity between certain equilibrium and non-equilibrium phase transitions is pointed out. Data from an experiment on human rhythmic movements is presented. Estimates for pumping intensity as well as the thermodynamical variables are reported. It is shown that in the experiment the non-equilibrium free energy decayed when pumping intensity was increased, which is consistent with the theory. Moreover, pumping intensities close to zero could be observed at relatively slow intended rhythmic movements. In view of the Hopf bifurcation underlying the limit cycle oscillator model, this observation suggests that the intended limit cycle movements were actually more similar to trajectories of a randomly perturbed stable focus. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Towards a human oral vaccine for anthrax: the utility of a Salmonella Typhi Ty21a-based prime boost immunization strategy

PubMed Central

Baillie, Leslie W.J.; Rodriguez, Ana L.; Moore, Stephen; Atkins, Helen S.; Feng, Chiguang; Nataro, James P.; Pasetti, Marcela F.

2008-01-01

We previously demonstrated the ability of an orally administered attenuated Salmonella enterica serovar Typhimurium strain expressing the protective antigen (PA) of Bacillus anthracis to confer protection against lethal anthrax aerosol spore challenge [1]. To extend the utility of this approach to humans we constructed variants of S. enterica serovar Typhi Ty21a, an attenuated typhoid vaccine strain licensed for human use, which expressed and exported PA via two distinct plasmid-based transport systems: the Escherichia coli HlyA haemolysin and the S. Typhi ClyA export apparatus. Murine immunogenicity studies confirmed the ability of these constructs, especially Ty21a expressing the ClyA-PA fusion protein, to stimulate strong PA-specific immune responses following intranasal immunization. These responses were further enhanced by a subsequent boost with either parenterally delivered recombinant PA or the licensed US human alum-adsorbed anthrax vaccine (AVA). Anthrax toxin neutralizing antibody responses using this prime-boost regimen were rapid, vigorous and broad in nature. The results of this study demonstrate the feasibility of employing a mucosal prime with a licensed Salmonella Typhi vaccine strain followed by a parenteral protein boost to stimulate rapid protective immunity against anthrax. PMID:18805452

Myosin Vb mediates Cu+ export in polarized hepatocytes

PubMed Central

Gupta, Arnab; Schell, Michael J.; Bhattacharjee, Ashima; Lutsenko, Svetlana; Hubbard, Ann L.

2016-01-01

ABSTRACT The cellular machinery responsible for Cu+-stimulated delivery of the Wilson-disease-associated protein ATP7B to the apical domain of hepatocytes is poorly understood. We demonstrate that myosin Vb regulates the Cu+-stimulated delivery of ATP7B to the apical domain of polarized hepatic cells, and that disruption of the ATP7B-myosin Vb interaction reduces the apical surface expression of ATP7B. Overexpression of the myosin Vb tail, which competes for binding of subapical cargos to myosin Vb bound to subapical actin, disrupted the surface expression of ATP7B, leading to reduced cellular Cu+ export. The myosin-Vb-dependent targeting step occurred in parallel with hepatocyte-like polarity. If the myosin Vb tail was expressed acutely in cells just prior to the establishment of polarity, it appeared as part of an intracellular apical compartment, centered on γ-tubulin. ATP7B became selectively arrested in this compartment at high [Cu+] in the presence of myosin Vb tail, suggesting that these compartments are precursors of donor–acceptor transfer stations for apically targeted cargos of myosin Vb. Our data suggest that reduced hepatic Cu+ clearance in idiopathic non-Wilsonian types of disease might be associated with the loss of function of myosin Vb. PMID:26823605

A Critical Role for CRM1 in Regulating HOXA Gene Transcription in CALM-AF10 Leukemias

PubMed Central

Conway, Amanda E.; Haldeman, Jonathan M.; Wechsler, Daniel S.; Lavau, Catherine P.

2014-01-01

The leukemogenic CALM-AF10 fusion protein is found in patients with immature acute myeloid and T-lymphoid malignancies. CALM-AF10 leukemias display abnormal H3K79 methylation and increased HOXA cluster gene transcription. Elevated expression of HOXA genes is critical for leukemia maintenance and progression; however, the precise mechanism by which CALM-AF10 alters HOXA gene expression is unclear. We previously determined that CALM contains a CRM1-dependent nuclear export signal (NES), which is both necessary and sufficient for CALM-AF10-mediated leukemogenesis. Here, we find that interaction of CALM-AF10 with the nuclear export receptor CRM1 is necessary for activating HOXA gene expression. We show that CRM1 localizes to HOXA loci where it recruits CALM-AF10, leading to transcriptional and epigenetic activation of HOXA genes. Genetic and pharmacological inhibition of the CALM-CRM1 interaction prevents CALM-AF10 enrichment at HOXA chromatin, resulting in immediate loss of transcription. These results provide a comprehensive mechanism by which the CALM-AF10 translocation activates the critical HOXA cluster genes. Furthermore, this report identifies a novel function of CRM1: the ability to bind chromatin and recruit the NES-containing CALM-AF10 transcription factor. PMID:25027513

Analysis of the subcellular localization of the human histone methyltransferase SETDB1

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

Tachibana, Keisuke, E-mail: nya@phs.osaka-u.ac.jp; Gotoh, Eiko; Kawamata, Natsuko

2015-10-02

SET domain, bifurcated 1 (SETDB1) is a histone methyltransferase that methylates lysine 9 on histone H3. Although it is important to know the localization of proteins to elucidate their physiological function, little is known of the subcellular localization of human SETDB1. In the present study, to investigate the subcellular localization of hSETDB1, we established a human cell line constitutively expressing enhanced green fluorescent protein fused to hSETDB1. We then generated a monoclonal antibody against the hSETDB1 protein. Expression of both exogenous and endogenous hSETDB1 was observed mainly in the cytoplasm of various human cell lines. Combined treatment with the nuclearmore » export inhibitor leptomycin B and the proteasome inhibitor MG132 led to the accumulation of hSETDB1 in the nucleus. These findings suggest that hSETDB1, localized in the nucleus, might undergo degradation by the proteasome and be exported to the cytosol, resulting in its detection mainly in the cytosol. - Highlights: • Endogenous human SETDB1 was localized mainly in the cytoplasm. • Combined treatment with LMB and MG132 led to accumulation of human SETDB1 in the nucleus. • HeLa cells expressing EFGP-hSETDB1 are useful for subcellular localization analyses.« less

Association of CD147 and Calcium Exporter PMCA4 Uncouples IL-2 Expression from Early TCR Signaling.

PubMed

Supper, Verena; Schiller, Herbert B; Paster, Wolfgang; Forster, Florian; Boulègue, Cyril; Mitulovic, Goran; Leksa, Vladimir; Ohradanova-Repic, Anna; Machacek, Christian; Schatzlmaier, Philipp; Zlabinger, Gerhard J; Stockinger, Hannes

2016-02-01

The Ig superfamily member CD147 is upregulated following T cell activation and was shown to serve as a negative regulator of T cell proliferation. Thus, Abs targeting CD147 are being tested as new treatment strategies for cancer and autoimmune diseases. How CD147 mediates immunosuppression and whether association with other coreceptor complexes is needed have remained unknown. In the current study, we show that silencing of CD147 in human T cells increases IL-2 production without affecting the TCR proximal signaling components. We mapped the immunosuppressive moieties of CD147 to its transmembrane domain and Ig-like domain II. Using affinity purification combined with mass spectrometry, we determined the domain specificity of CD147 interaction partners and identified the calcium exporter plasma membrane calcium ATPase isoform 4 (PMCA4) as the interaction partner of the immunosuppressive moieties of CD147. CD147 does not control the proper membrane localization of PMCA4, but PMCA4 is essential for the CD147-dependent inhibition of IL-2 expression via a calcium-independent mechanism. In summary, our data show that CD147 interacts via its immunomodulatory domains with PMCA4 to bypass TCR proximal signaling and inhibit IL-2 expression. Copyright © 2016 by The American Association of Immunologists, Inc.

Regulation of the aceI multidrug efflux pump gene in Acinetobacter baumannii.

PubMed

Liu, Qi; Hassan, Karl A; Ashwood, Heather E; Gamage, Hasinika K A H; Li, Liping; Mabbutt, Bridget C; Paulsen, Ian T

2018-06-01

To investigate the function of AceR, a putative transcriptional regulator of the chlorhexidine efflux pump gene aceI in Acinetobacter baumannii. Chlorhexidine susceptibility and chlorhexidine induction of aceI gene expression were determined by MIC and quantitative real-time PCR, respectively, in A. baumannii WT and ΔaceR mutant strains. Recombinant AceR was prepared as both a full-length protein and as a truncated protein, AceR (86-299), i.e. AceRt, which has the DNA-binding domain deleted. The binding interaction of the purified AceR protein and its putative operator region was investigated by electrophoretic mobility shift assays and DNase I footprinting assays. The binding of AceRt with its putative ligand chlorhexidine was examined using surface plasmon resonance and tryptophan fluorescence quenching assays. MIC determination assays indicated that the ΔaceI and ΔaceR mutant strains both showed lower resistance to chlorhexidine than the parental strain. Chlorhexidine-induced expression of aceI was abolished in a ΔaceR background. Electrophoretic mobility shift assays and DNase I footprinting assays demonstrated chlorhexidine-stimulated binding of AceR with two sites upstream of the putative aceI promoter. Surface plasmon resonance and tryptophan fluorescence quenching assays suggested that the purified ligand-binding domain of the AceR protein was able to bind with chlorhexidine with high affinity. This study provides strong evidence that AceR is an activator of aceI gene expression when challenged with chlorhexidine. This study is the first characterization, to our knowledge, of a regulator controlling expression of a PACE family multidrug efflux pump.

Hepatitis C virus core protein triggers abnormal porphyrin metabolism in human hepatocellular carcinoma cells.

PubMed

Nakano, Takafumi; Moriya, Kyoji; Koike, Kazuhiko; Horie, Toshiharu

2018-01-01

Porphyria cutanea tarda (PCT), the most common of the human porphyrias, arises from a deficiency of uroporphyrinogen decarboxylase. Studies have shown a high prevalence of hepatitis C virus (HCV) infection in patients with PCT. While these observations implicate HCV infection as a risk factor for PCT pathogenesis, the mechanism of interaction between the virus and porphyrin metabolism is unknown. This study aimed to assess the effect of HCV core protein on intracellular porphyrin metabolism to elucidate the link between HCV infection and PCT. The accumulation and excretion of porphyrins after treatment with 5-aminolevulinic acid, a porphyrin precursor, were compared between cells stably expressing HCV core protein and controls. Cells expressing HCV core protein had lower amounts of intracellular protoporphyrin IX and heme and had higher amounts of excreted coproporphyrin III, the oxidized form of coproporphyrinogen III, compared with controls. These observations suggest that HCV core protein affects porphyrin metabolism and facilitates the export of excess coproporphyrinogen III and/or coproporphyrin III, possibly via porphyrin transporters. Real-time PCR analysis revealed that the presence of HCV core protein increased the mRNA expression of porphyrin exporters ABCG2 and FLVCR1. Western blot analysis showed a higher expression level of FLVCR1, but not ABCG2, as well as a higher expression level of mature ALAS1, which is the rate-limiting enzyme in the heme synthesis pathway, in HCV core protein-expressing cells compared with controls. The data indicate that HCV core protein induced abnormal intracellular porphyrin metabolism, with an over-excretion of coproporphyrin III. These findings may partially account for the susceptibility of HCV-infected individuals to PCT development.

Depletion of mRNA export regulator DBP5/DDX19, GLE1 or IPPK that is a key enzyme for the production of IP6, resulting in differentially altered cytoplasmic mRNA expression and specific cell defect

PubMed Central

Okamura, Masumi; Yamanaka, Yasutaka; Shigemoto, Maki; Kitadani, Yuya; Kobayashi, Yuhko; Kambe, Taiho; Nagao, Masaya; Kobayashi, Issei; Okumura, Katsuzumi

2018-01-01

DBP5, also known as DDX19, GLE1 and inositol hexakisphosphate (IP6) function in messenger RNA (mRNA) export at the cytoplasmic surface of the nuclear pore complex in eukaryotic cells. DBP5 is a DEAD-box RNA helicase, and its activity is stimulated by interactions with GLE1 and IP6. In addition, these three factors also have unique role(s). To investigate how these factors influenced the cytoplasmic mRNA expression and cell phenotype change, we performed RNA microarray analysis to detect the effect and function of DBP5, GLE1 and IP6 on the cytoplasmic mRNA expression. The expression of some cytoplasmic mRNA subsets (e.g. cell cycle, DNA replication) was commonly suppressed by the knock-down of DBP5, GLE1 and IPPK (IP6 synthetic enzyme). The GLE1 knock-down selectively reduced the cytoplasmic mRNA expression required for mitotic progression, results in an abnormal spindle phenotype and caused the delay of mitotic process. Meanwhile, G1/S cell cycle arrest was observed in DBP5 and IPPK knock-down cells. Several factors that function in immune response were also down-regulated in DBP5 or IPPK knock-down cells. Thereby, IFNβ-1 mRNA transcription evoked by poly(I:C) treatment was suppressed. These results imply that DBP5, GLE1 and IP6 have a conserved and individual function in the cytoplasmic mRNA expression. Variations in phenotype are due to the difference in each function of DBP5, GLE1 and IPPK in intracellular mRNA metabolism. PMID:29746542

Differential expression of THOC1 and ALY mRNP biogenesis/export factors in human cancers.

PubMed

Domínguez-Sánchez, María S; Sáez, Carmen; Japón, Miguel A; Aguilera, Andrés; Luna, Rosa

2011-02-17

One key step in gene expression is the biogenesis of mRNA ribonucleoparticle complexes (mRNPs). Formation of the mRNP requires the participation of a number of conserved factors such as the THO complex. THO interacts physically and functionally with the Sub2/UAP56 RNA-dependent ATPase, and the Yra1/REF1/ALY RNA-binding protein linking transcription, mRNA export and genome integrity. Given the link between genome instability and cancer, we have performed a comparative analysis of the expression patterns of THOC1, a THO complex subunit, and ALY in tumor samples. The mRNA levels were measured by quantitative real-time PCR and hybridization of a tumor tissue cDNA array; and the protein levels and distribution by immunostaining of a custom tissue array containing a set of paraffin-embedded samples of different tumor and normal tissues followed by statistical analysis. We show that the expression of two mRNP factors, THOC1 and ALY are altered in several tumor tissues. THOC1 mRNA and protein levels are up-regulated in ovarian and lung tumors and down-regulated in those of testis and skin, whereas ALY is altered in a wide variety of tumors. In contrast to THOC1, ALY protein is highly detected in normal proliferative cells, but poorly in high-grade cancers. These results suggest a differential connection between tumorogenesis and the expression levels of human THO and ALY. This study opens the possibility of defining mRNP biogenesis factors as putative players in cell proliferation that could contribute to tumor development.

The putative drug efflux systems of the Bacillus cereus group

PubMed Central

Elbourne, Liam D. H.; Vörös, Aniko; Kroeger, Jasmin K.; Simm, Roger; Tourasse, Nicolas J.; Finke, Sarah; Henderson, Peter J. F.; Økstad, Ole Andreas; Paulsen, Ian T.; Kolstø, Anne-Brit

2017-01-01

The Bacillus cereus group of bacteria includes seven closely related species, three of which, B. anthracis, B. cereus and B. thuringiensis, are pathogens of humans, animals and/or insects. Preliminary investigations into the transport capabilities of different bacterial lineages suggested that genes encoding putative efflux systems were unusually abundant in the B. cereus group compared to other bacteria. To explore the drug efflux potential of the B. cereus group all putative efflux systems were identified in the genomes of prototypical strains of B. cereus, B. anthracis and B. thuringiensis using our Transporter Automated Annotation Pipeline. More than 90 putative drug efflux systems were found within each of these strains, accounting for up to 2.7% of their protein coding potential. Comparative analyses demonstrated that the efflux systems are highly conserved between these species; 70–80% of the putative efflux pumps were shared between all three strains studied. Furthermore, 82% of the putative efflux system proteins encoded by the prototypical B. cereus strain ATCC 14579 (type strain) were found to be conserved in at least 80% of 169 B. cereus group strains that have high quality genome sequences available. However, only a handful of these efflux pumps have been functionally characterized. Deletion of individual efflux pump genes from B. cereus typically had little impact to drug resistance phenotypes or the general fitness of the strains, possibly because of the large numbers of alternative efflux systems that may have overlapping substrate specificities. Therefore, to gain insight into the possible transport functions of efflux systems in B. cereus, we undertook large-scale qRT-PCR analyses of efflux pump gene expression following drug shocks and other stress treatments. Clustering of gene expression changes identified several groups of similarly regulated systems that may have overlapping drug resistance functions. In this article we review current knowledge of the small molecule efflux pumps encoded by the B. cereus group and suggest the likely functions of numerous uncharacterised pumps. PMID:28472044

Table-top two-color soft X-ray laser by means of Ni-like plasmas

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

Masoudnia, Leili; Ruiz-Lopez, Mabel; Bleiner, Davide, E-mail: davide.bleiner@empa.ch

2016-04-15

Laser-produced Ni-like plasmas are known as active media for extreme ultraviolet lasing, with the flexibility to two-color lasing. Two-color laser generation is very complex at accelerator facilities. In this work, plasma lasing at the 3d{sup 9}4d{sup 1}(J = 0) → 3d{sup 9}4p{sup 1}(J = 1) (collisional-pumping process) and the 3d{sup 9}4f{sup 1}(J = 1) → 3d{sup 9}4d{sup 1}(J = 1) (photo-pumping process) transitions is studied experimentally and computationally. Several key characteristics of collisional- and photo-pumping laser, such as divergence, pointing stability, and intensity have been investigated. The measurements showed different pulse characteristics for the two lasing processes affected by plasma inhomogeneity in temperature and density. Analytical expressions of these characteristicsmore » for both collisional- and photo-pumping are derived. It is found that the plasma that maximizes the photo-pumping lasing is 20% hotter and 70% denser than the plasma that optimizes the collisional-pumping lasing. The gain of collisional pumping is ≈4 times higher than the gain for the photo-pumping. The gain lifetime is a factor of ≈5.2 larger for the monopole-pumping. Similarly, the gain thickness is a factor of ≈1.8 larger. It is also found that the gain build-up time for collisional- and photo-pumping is 0.7 ps and 0.9 ps, respectively, whereas the build-up length-scale is 11.5 μm and 6.3 μm, respectively.« less

Employee perception of breastfeeding-friendly support and benefits of breastfeeding as a predictor of intention to use breast-pumping breaks after returning to work among employed mothers.

PubMed

Tsai, Su-Ying

2014-01-01

Although increasing numbers of large companies are complying with demands for a breastfeeding-friendly workplace by providing lactation rooms and breast-pumping breaks, the effectiveness for intention to use breast-pumping breaks to express breast milk among employed mothers is uncertain. To explore the impact of employees' perceived breastfeeding support from the workplace and the benefits of breastfeeding on a woman's intention to use breast-pumping breaks after returning to work, we conducted a survey at a female labor-intensive electronics manufacturer in Taiwan. A structured questionnaire survey was administered to 715 working mothers employed in an electronics manufacturing plant in Tainan Science Park in Southern Taiwan. Questionnaire content included female employee demographics, employment characteristics, and breastfeeding behavior after returning to work, as well as employees' perception of breastfeeding-friendly support and awareness of the benefits of breastfeeding when raising their most recently born child. Higher education (odds ratio [OR] 2.33), non-clean room worksite (OR 1.51), awareness of breast-pumping breaks (OR 4.70), encouragement by colleagues to use breast-pumping breaks (OR 1.76), and greater awareness of the benefits of breastfeeding (OR 1.08) were significant predictors of the use of breast-pumping breaks after returning to work, whereas the perception of inefficiency when using breast-pumping breaks reduced an employed mother's intention to use breast-pumping breaks (OR 0.55). This study finds an association between an appreciation of the benefits provided by the employer and the likelihood of increased usage of breastfeeding breaks. Workplaces and employers can help employed mothers to understand the benefits of breastfeeding, which may increase the intention of the mother to take breast-pumping breaks after returning to work.

'2TM proteins': an antigenically diverse superfamily with variable functions and export pathways.

PubMed

Kaur, Jasweer; Hora, Rachna

2018-01-01

Malaria is a disease that affects millions of people annually. An intracellular habitat and lack of protein synthesizing machinery in erythrocytes pose numerous difficulties for survival of the human pathogen Plasmodium falciparum . The parasite refurbishes the infected red blood cell (iRBC) by synthesis and export of several proteins in an attempt to suffice its metabolic needs and evade the host immune response. Immune evasion is largely mediated by surface display of highly polymorphic protein families known as variable surface antigens. These include the two trans-membrane (2TM) superfamily constituted by multicopy repetitive interspersed family (RIFINs), subtelomeric variable open reading frame (STEVORs) and Plasmodium falciparum Maurer's cleft two trans-membrane proteins present only in P. falciparum and some simian infecting Plasmodium species. Their hypervariable region flanked by 2TM domains exposed on the iRBC surface is believed to generate antigenic diversity. Though historically named "2TM superfamily," several A-type RIFINs and some STEVORs assume one trans-membrane topology. RIFINs and STEVORs share varied functions in different parasite life cycle stages like rosetting, alteration of iRBC rigidity and immune evasion. Additionally, a member of the STEVOR family has been implicated in merozoite invasion. Differential expression of these families in laboratory strains and clinical isolates propose them to be important for host cell survival and defense. The role of RIFINs in modulation of host immune response and presence of protective antibodies against these surface exposed molecules in patient sera highlights them as attractive targets of antimalarial therapies and vaccines. 2TM proteins are Plasmodium export elements positive, and several of these are exported to the infected erythrocyte surface after exiting through the classical secretory pathway within parasites. Cleaved and modified proteins are trafficked after packaging in vesicles to reach Maurer's clefts, while information regarding delivery to the iRBC surface is sparse. Expression and export timing of the RIFIN and Plasmodium falciparum erythrocyte membrane protein1 families correspond to each other. Here, we have compiled and comprehended detailed information regarding orthologues, domain architecture, surface topology, functions and trafficking of members of the "2TM superfamily." Considering the large repertoire of proteins included in the 2TM superfamily and recent advances defining their function in malaria biology, a surge in research carried out on this important protein superfamily is likely.

Calmodulin activation of an endoplasmic reticulum-located calcium pump involves an interaction with the N-terminal autoinhibitory domain

NASA Technical Reports Server (NTRS)

Hwang, I.; Harper, J. F.; Liang, F.; Sze, H.

2000-01-01

To investigate how calmodulin regulates a unique subfamily of Ca(2+) pumps found in plants, we examined the kinetic properties of isoform ACA2 identified in Arabidopsis. A recombinant ACA2 was expressed in a yeast K616 mutant deficient in two endogenous Ca(2+) pumps. Orthovanadate-sensitive (45)Ca(2+) transport into vesicles isolated from transformants demonstrated that ACA2 is a Ca(2+) pump. Ca(2+) pumping by the full-length protein (ACA2-1) was 4- to 10-fold lower than that of the N-terminal truncated ACA2-2 (Delta2-80), indicating that the N-terminal domain normally acts to inhibit the pump. An inhibitory sequence (IC(50) = 4 microM) was localized to a region within valine-20 to leucine-44, because a peptide corresponding to this sequence lowered the V(max) and increased the K(m) for Ca(2+) of the constitutively active ACA2-2 to values comparable to the full-length pump. The peptide also blocked the activity (IC(50) = 7 microM) of a Ca(2+) pump (AtECA1) belonging to a second family of Ca(2+) pumps. This inhibitory sequence appears to overlap with a calmodulin-binding site in ACA2, previously mapped between aspartate-19 and arginine-36 (J.F. Harper, B. Hong, I. Hwang, H.Q. Guo, R. Stoddard, J.F. Huang, M.G. Palmgren, H. Sze inverted question mark1998 J Biol Chem 273: 1099-1106). These results support a model in which the pump is kept "unactivated" by an intramolecular interaction between an autoinhibitory sequence located between residues 20 and 44 and a site in the Ca(2+) pump core that is highly conserved between different Ca(2+) pump families. Results further support a model in which activation occurs as a result of Ca(2+)-induced binding of calmodulin to a site overlapping or immediately adjacent to the autoinhibitory sequence.

Expressing the sense of Congress that all direct and indirect subsidies that benefit the production or export of sugar by all major sugar producing and consuming countries should be eliminated.

THOMAS, 113th Congress

Rep. Yoho, Ted S. [R-FL-3

2013-06-14

House - 06/25/2013 Referred to the Subcommittee on General Farm Commodities and Risk Management. (All Actions) Tracker: This bill has the status IntroducedHere are the steps for Status of Legislation:

Broad Specificity Efflux pumps and Their Role in Multidrug Resistance of Gram Negative Bacteria

PubMed Central

Nikaido, Hiroshi; Pagès, Jean-Marie

2013-01-01

Antibiotic resistance mechanisms reported in Gram-negative bacteria are producing a worldwide health problem. The continuous dissemination of «multi-drug resistant» (MDR) bacteria drastically reduces the efficacy of our antibiotic “arsenal” and consequently increases the frequency of therapeutic failure. In MDR bacteria, the over-expression of efflux pumps that expel structurally-unrelated drugs contributes to the reduced susceptibility by decreasing the intracellular concentration of antibiotics. During the last decade, several clinical data indicate an increasing involvement of efflux pumps in the emergence and dissemination of resistant Gram-negative bacteria. It is necessary to clearly define the molecular, functional and genetic bases of the efflux pump in order to understand the translocation of antibiotic molecules through the efflux transporter. The recent investigation on the efflux pump AcrB at its structural and physiological level, including the identification of drug affinity sites and kinetic parameters for various antibiotics, may open the way to rationally develop an improved new generation of antibacterial agents as well as efflux inhibitors in order to efficiently combat efflux-based resistance mechanisms. PMID:21707670

Carbon export in the naturally iron-fertilized Kerguelen area of the Southern Ocean based on the 234Th approach

NASA Astrophysics Data System (ADS)

Planchon, F.; Ballas, D.; Cavagna, A.-J.; Bowie, A. R.; Davies, D.; Trull, T.; Laurenceau, E.; Van Der Merwe, P.; Dehairs, F.

2014-11-01

The Kerguelen Plateau region in the Indian sector of the Southern Ocean supports annually a large-scale phytoplankton bloom which is naturally fertilized with iron. As part of the second KErguelen Ocean and Plateau compared Study expedition (KEOPS2) in austral spring (October-November 2011), we examined upper-ocean Particulate Organic Carbon (POC) export using the 234Th approach. We aimed at characterizing the spatial and the temporal variability of POC export production at high productivity sites over and downstream the Kerguelen plateau. Export production is compared to a High Nutrient Low Chlorophyll area upstream of the plateau in order to assess the impact of iron-induced productivity on the vertical export of carbon. Deficits in 234Th activities relative to its parent nuclide 238U were observed at all stations in surface waters, indicating that scavenging by particles occurred during the early stages of the phytoplankton bloom. 234Th export was lowest at reference station R-2 (412 ± 134 dpm m-2 d-1) and highest inside a~permanent meander of the Polar Front (PF) at stations E (1995 ± 176 dpm m-2 d-1, second visit E-3) where a detailed time series was obtained as part of a~pseudo-lagrangian study. 234Th export over the central plateau was relatively limited at station A3 early (776 ± 171 dpm m-2 d-1, first visit A3-1) and late in the survey (993 ± 223 dpm m-2 d-1, second visit A3-2), but it was higher at high biomass stations TNS-8 (1372 ± 255 dpm m-2 d-1) and E-4W (1068 ± 208 dpm m-2 d-1) in waters which could be considered as derived from plateau. Limited 234Th export of 973 ± 207 dpm m-2 d-1 was also found in the northern branch of the Kerguelen bloom located downstream of the island, north of the PF (station F-L). The 234Th results support that Fe fertilization increased particle export in all iron fertilized waters. The impact was greatest in the recirculation feature (3-4 fold at 200 m depth), but more moderate over the central Kerguelen plateau and in the northern plume of the Kerguelen bloom (∼2-fold at 200 m depth). The C : Th ratio of large (> 53 μm) potentially sinking particles collected via sequential filtration using in situ pumping (ISP) systems were used to convert the 234Th flux into a POC export flux. The C : Th ratios of sinking particles were highly variable (range: 3.1 ± 0.1-10.5 ± 0.2 μmol dpm-1) with no clear site related trend, despite the variety of ecosystem responses in the fertilized regions. C : Th ratios showed a decreasing trend between 100 and 200 m depth suggesting preferential loss of carbon relative to 234Th possibly due to heterotrophic degradation and/or grazing activity. Comparison of the C : Th ratios within sinking particles obtained with the drifting sediment traps showed in most cases very good agreement to those collected via ISP deployments (> 53 μm particles). Carbon export production varied between 3.5 ± 0.9 mmol m-2 d-1 and 11.8 ± 1.3 mmol m-2 d-1 from the upper 100 m and between 1.8 ± 0.9 mmol m-2 d-1 and 8.2 ± 0.9 mmol m-2 d-1 from the upper 200 m. Highest export production was found inside the PF meander with a range of 5.4 ± 0.7 mmol m-2 d-1 to 11.8 ± 1.1 mmol m-2 d-1 at 100 m depth decreasing to 5.3 ± 1.0 mmol m-2 d-1 to 8.2 ± 0.8 mmol m-2 d-1 at 200 m depth over the 19 day survey period. The impact of Fe fertilization is highest inside the PF meander with 2.9- up to 4.5-fold higher carbon flux at 200 m depth in comparison to the HNLC control station. The impact of Fe fertilization was significantly less over the central plateau (stations A3 and E-4W) and in the northern branch of the bloom (station F-L) with 1.6- up to 2.0-fold higher carbon flux compared to the reference station R. Export efficiencies (ratio of export to primary production) were particularly variable with relatively high values in the recirculation feature (6-27%) and low values (1-5%) over the central plateau (station A3) and north of the PF (station F-L) indicating spring biomass accumulation. Comparison with KEOPS1 results indicated that carbon export production is much lower during the onset of the bloom in austral spring in comparison to the peak and declining phase in late summer.

Ketamine produces antidepressant-like effects through phosphorylation-dependent nuclear export of histone deacetylase 5 (HDAC5) in rats

PubMed Central

Choi, Miyeon; Lee, Seung Hoon; Wang, Sung Eun; Ko, Seung Yeon; Song, Mihee; Choi, June-Seek; Duman, Ronald S.; Son, Hyeon

2015-01-01

Ketamine produces rapid antidepressant-like effects in animal assays for depression, although the molecular mechanisms underlying these behavioral actions remain incomplete. Here, we demonstrate that ketamine rapidly stimulates histone deacetylase 5 (HDAC5) phosphorylation and nuclear export in rat hippocampal neurons through calcium/calmodulin kinase II- and protein kinase D-dependent pathways. Consequently, ketamine enhanced the transcriptional activity of myocyte enhancer factor 2 (MEF2), which leads to regulation of MEF2 target genes. Transfection of a HDAC5 phosphorylation-defective mutant (Ser259/Ser498 replaced by Ala259/Ala498, HDAC5-S/A), resulted in resistance to ketamine-induced nuclear export, suppression of ketamine-mediated MEF2 transcriptional activity, and decreased expression of MEF2 target genes. Behaviorally, viral-mediated hippocampal knockdown of HDAC5 blocked or occluded the antidepressant effects of ketamine both in unstressed and stressed animals. Taken together, our results reveal a novel role of HDAC5 in the actions of ketamine and suggest that HDAC5 could be a potential mechanism contributing to the therapeutic actions of ketamine. PMID:26647181

Quantitative Analysis Reveals that Actin and Src-Family Kinases Regulate Nuclear YAP1 and Its Export.

PubMed

Ege, Nil; Dowbaj, Anna M; Jiang, Ming; Howell, Michael; Hooper, Steven; Foster, Charles; Jenkins, Robert P; Sahai, Erik

2018-06-08

The transcriptional regulator YAP1 is critical for the pathological activation of fibroblasts. In normal fibroblasts, YAP1 is located in the cytoplasm, while in activated cancer-associated fibroblasts, it is nuclear and promotes the expression of genes required for pro-tumorigenic functions. Here, we investigate the dynamics of YAP1 shuttling in normal and activated fibroblasts, using EYFP-YAP1, quantitative photobleaching methods, and mathematical modeling. Imaging of migrating fibroblasts reveals the tight temporal coupling of cell shape change and altered YAP1 localization. Both 14-3-3 and TEAD binding modulate YAP1 shuttling, but neither affects nuclear import. Instead, we find that YAP1 nuclear accumulation in activated fibroblasts results from Src and actomyosin-dependent suppression of phosphorylated YAP1 export. Finally, we show that nuclear-constrained YAP1, upon XPO1 depletion, remains sensitive to blockade of actomyosin function. Together, these data place nuclear export at the center of YAP1 regulation and indicate that the cytoskeleton can regulate YAP1 within the nucleus. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Host cell subversion by Toxoplasma GRA16, an exported dense granule protein that targets the host cell nucleus and alters gene expression.

PubMed

Bougdour, Alexandre; Durandau, Eric; Brenier-Pinchart, Marie-Pierre; Ortet, Philippe; Barakat, Mohamed; Kieffer, Sylvie; Curt-Varesano, Aurélie; Curt-Bertini, Rose-Laurence; Bastien, Olivier; Coute, Yohann; Pelloux, Hervé; Hakimi, Mohamed-Ali

2013-04-17

After invading host cells, Toxoplasma gondii multiplies within a parasitophorous vacuole (PV) that is maintained by parasite proteins secreted from organelles called dense granules. Most dense granule proteins remain within the PV, and few are known to access the host cell cytosol. We identify GRA16 as a dense granule protein that is exported through the PV membrane and reaches the host cell nucleus, where it positively modulates genes involved in cell-cycle progression and the p53 tumor suppressor pathway. GRA16 binds two host enzymes, the deubiquitinase HAUSP and PP2A phosphatase, which exert several functions, including regulation of p53 and the cell cycle. GRA16 alters p53 levels in a HAUSP-dependent manner and induces nuclear translocation of the PP2A holoenzyme. Additionally, certain GRA16-deficient strains exhibit attenuated virulence, indicating the importance of these host alterations in pathogenesis. Therefore, GRA16 represents a potentially emerging subfamily of exported dense granule proteins that modulate host function. Copyright © 2013 Elsevier Inc. All rights reserved.

Constructing the wonders of the bacterial world: biosynthesis of complex enzymes.

PubMed

Sargent, Frank

2007-03-01

The prokaryotic cytoplasmic membrane not only maintains cell integrity and forms a barrier between the cell and its outside environment, but is also the location for essential biochemical processes. Microbial model systems provide excellent bases for the study of fundamental problems in membrane biology including signal transduction, chemotaxis, solute transport and, as will be the topic of this review, energy metabolism. Bacterial respiration requires a diverse array of complex, multi-subunit, cofactor-containing redox enzymes, many of which are embedded within, or located on the extracellular side of, the membrane. The biosynthesis of these enzymes therefore requires carefully controlled expression, assembly, targeting and transport processes. Here, focusing on the molybdenum-containing respiratory enzymes central to anaerobic respiration in Escherichia coli, recent descriptions of a chaperone-mediated 'proofreading' system involved in coordinating assembly and export of complex extracellular enzymes will be discussed. The paradigm proofreading chaperones are members of a large group of proteins known as the TorD family, and recent research in this area highlights common principles that underpin biosynthesis of both exported and non-exported respiratory enzymes.

Time Resolved Efficiency Degradation in Potassium Diode Pumped Alkali Laser

DTIC Science & Technology

2014-08-07

study of the performance of a Potassium OPAL operating in pulsed mode with pulses up to 5 msec Jong at different pulse energies and cell...temperatures. The experiments showed the OPAL efficiency degradation in time with a characteristic time in the range from 0.5 msec to 4.5 msec. The recorded...Lasers", Optics Express, 19(8), 7894-7902 (20 I I) 1. Introduction There has been extensive research into Diode Pumped Alkali Lasers ( OPALs ) during the

Multidrug Efflux Systems in Microaerobic and Anaerobic Bacteria

PubMed Central

Xu, Zeling; Yan, Aixin

2015-01-01

Active drug efflux constitutes an important mechanism of antibiotic and multidrug resistance in bacteria. Understanding the distribution, expression, and physiological functions of multidrug efflux pumps, especially under physiologically and clinically relevant conditions of the pathogens, is the key to combat drug resistance. In animal hosts, most wounded, infected and inflamed tissues display low oxygen tensions. In this article, we summarize research development on multidrug efflux pumps in the medicinally relevant microaerobic and anaerobic pathogens and their implications in the effort to combat drug-resistant infections. PMID:27025630

Late summer particulate organic carbon export and twilight zone remineralisation in the Atlantic sector of the Southern Ocean

NASA Astrophysics Data System (ADS)

Planchon, F.; Cavagna, A.-J.; Cardinal, D.; André, L.; Dehairs, F.

2013-02-01

As part of the GEOTRACES Bonus-GoodHope (BGH) expedition (January-March 2008) in the Atlantic sector of the Southern Ocean, particulate organic carbon (POC) export was examined from the surface to the mesopelagic twilight zone using water column distributions of total 234Th and biogenic particulate Ba (Baxs). Surface POC export production was estimated from steady state and non steady state modelling of 234Th fluxes, which were converted into POC fluxes, using the POC/234Th ratio of large, potentially sinking particles (> 53 μm) collected via in situ pumps. Deficits in 234Th activities were observed at all stations from the surface to the bottom of the mixed layer, yielding 234Th export fluxes from the upper 100 m of 496 ± 214 dpm m-2 d-1 to 1195 ± 158 dpm m-2 d-1 for the steady state model and of 149 ±517 dpm m-2 d-1 to 1217 ± 231 dpm m-2 d-1 for the non steady state model. Using the POC/234Thp ratio of sinking particles (ratios varied from 1.7 ± 0.2 μmol dpm-1 to 4.8 ± 1.9 μmol dpm-1) POC export production at 100 m was calculated to range between 0.9 ± 0.4 and 5.1 ± 2.1 mmol C m-2 d-1,assuming steady state and between 0.3 ± 0.9 m-2 d-1 and 4.9 ± 3.3 mmol C m-2 d-1, assuming non steady state. From the comparison of both approaches, it appears that during late summer export decreased by 56 to 16% for the area between the sub-Antarctic zone and the southern Antarctic Circumpolar Current Front (SACCF), whereas it remained rather constant over time in the HNLC area south of the SACCF. POC export represented only 6 to 54% of new production, indicating that export efficiency was, in general, low, except in the vicinity of the SACCF, where export represented 56% of new production. Attenuation of the POC sinking flux in the upper mesopelagic waters (100-600 m depth interval) was evidenced both, from excess 234Th activities and from particulate biogenic Ba (Baxs) accumulation. Excess 234Th activities, reflected by 234Th/238U ratios as large as 1.21 ± 0.05, are attributed to remineralisation/disaggregation of 234Th-bearing particles. The accumulation of excess 234Th in the 100-600 m depth interval ranged from 458 ± 633 dpm m-2 d-1 to 3068 ± 897 dpm m-2 d-1, assuming steady state. Using the POC/234Thp ratio of sinking particles (> 53 μm), this 234Th accumulation flux was converted into a POC remineralisation flux which ranged between 0.9 ± 1.2 mmol C m-2 d-1 and 9.2 ± 2.9 mmol C m-2 d-1. Mesopelagic particulate biogenic Ba has been reported to reflect bacterial degradation of organic matter and to be related to oxygen consumption and bacterial carbon respiration. We observed that the highest Baxs contents (reaching up to > 1000 pM), in general, occurred between 200 and 400 m. Depth-weighted average mesopelagic Baxs (meso-Baxs) values were converted into respired C fluxes, which ranged between 0.23 and 6.4 mmol C m-2 d-1, in good agreement with 234Th-based remineralisation fluxes. A major outcome from this study is the observed significant positive correlation between POC remineralisation as estimated from meso-Baxs contents and from 234Th excess (R2 = 0.73; excluding 2 outliers). Remineralisation of POC in the twilight zone was particularly efficient relative to POC export resulting in negligible bathypelagic (> 600 m) POC export fluxes in the sub-Antarctic zone, the Polar Front zone and the northern Weddell Gyre, while the subtropical zone as well as the vicinity of the SACCF had significant deep POC fluxes.

Differential inhibition of rat and human Na+-dependent taurocholate cotransporting polypeptide (NTCP/SLC10A1)by bosentan: a mechanism for species differences in hepatotoxicity.

PubMed

Leslie, Elaine M; Watkins, Paul B; Kim, Richard B; Brouwer, Kim L R

2007-06-01

Bile acid accumulation in hepatocytes due to inhibition of the canalicular bile salt export pump (BSEP/ABCB11) has been proposed as a mechanism for bosentan-induced hepatotoxicity. The observation that bosentan does not induce hepatotoxicity in rats, although bosentan has been reported to inhibit rat Bsep and cause elevated serum bile acids, challenges this mechanism. The lack of hepatotoxicity could be explained if bosentan inhibited hepatocyte uptake as well as canalicular efflux of bile acids. In the current study, bosentan was found to be a more potent inhibitor of Na(+)-dependent taurocholate uptake in rat (IC(50) 5.4 microM) than human (IC(50) 30 microM) suspended hepatocytes. In addition, bosentan was a more potent inhibitor of taurocholate uptake by rat Na(+)-dependent taurocholate co-transporting polypeptide (Ntcp/Slc10a1) (IC(50) 0.71 microM) than human NTCP (SLC10A1) (IC(50) 24 microM) expressed in HEK293 cells. Thus, bosentan is a more potent inhibitor of Ntcp than NTCP, and this should result in less intrahepatocyte accumulation of bile acids in rats during bosentan treatment. To begin characterization of this species difference, two chimeric molecules were generated and expressed in HEK293 cells; NTCP(1-140)/Ntcp(141-362) and Ntcp(1-140)/NTCP(141-349). The mode of bosentan inhibition was noncompetitive for Ntcp, and competitive for NTCP (K(i) 18 microM) and NTCP(1-140)/Ntcp(141-362) (K(i) 1.7 microM); bosentan affected both the K(m) and V(max) of Ntcp(1-140)/NTCP(141-349) (K(i) 7.0 microM). The carboxyl portions of NTCP and Ntcp were found to confer species differences in basal taurocholate transport V(max). In conclusion, differential inhibition of Ntcp and NTCP may represent a novel mechanism for species differences in bosentan-induced hepatotoxicity.

Silencing the Menkes Copper-Transporting ATPase (Atp7a) Gene in Rat Intestinal Epithelial (IEC-6) Cells Increases Iron Flux via Transcriptional Induction of Ferroportin 1 (Fpn1)123

PubMed Central

Gulec, Sukru; Collins, James F.

2014-01-01

The Menkes copper-transporting ATPase (Atp7a) gene is induced in rat duodenum during iron deficiency, consistent with copper accumulation in the intestinal mucosa and liver. To test the hypothesis that ATP7A influences intestinal iron metabolism, the Atp7a gene was silenced in rat intestinal epithelial (IEC-6) cells using short hairpin RNA (shRNA) technology. Perturbations in intracellular copper homeostasis were noted in knockdown cells, consistent with the dual roles of ATP7A in pumping copper into the trans-Golgi (for cuproenzyme synthesis) and exporting copper from cells. Intracellular iron concentrations were unaffected by Atp7a knockdown. Unexpectedly, however, vectorial iron (59Fe) transport increased (∼33%) in knockdown cells grown in bicameral inserts and increased further (∼70%) by iron deprivation (compared with negative control shRNA-transfected cells). Additional experiments were designed to elucidate the molecular mechanism of increased transepithelial iron flux. Enhanced iron uptake by knockdown cells was associated with increased expression of a ferrireductase (duodenal cytochrome b) and activity of a cell-surface ferrireductase. Increased iron efflux from knockdown cells was likely mediated via transcriptional activation of the ferroportin 1 gene (by an unknown mechanism). Moreover, Atp7a knockdown significantly attenuated expression of an iron oxidase [hephaestin (HEPH); by ∼80%] and membrane ferroxidase activity (by ∼50%). Cytosolic ferroxidase activity, however, was retained in knockdown cells (75% of control cells), perhaps compensating for diminished HEPH activity. This investigation has thus documented alterations in iron homeostasis associated with Atp7a knockdown in enterocyte-like cells. Alterations in copper transport, trafficking, or distribution may underlie the increase in transepithelial iron flux noted when ATP7A activity is diminished. PMID:24174620

Towards a human oral vaccine for anthrax: the utility of a Salmonella Typhi Ty21a-based prime-boost immunization strategy.

PubMed

Baillie, Leslie W J; Rodriguez, Ana L; Moore, Stephen; Atkins, Helen S; Feng, Chiguang; Nataro, James P; Pasetti, Marcela F

2008-11-11

We previously demonstrated the ability of an orally administered attenuated Salmonella enterica serovar Typhimurium strain expressing the protective antigen (PA) of Bacillus anthracis to confer protection against lethal anthrax aerosol spore challenge [Stokes MG, Titball RW, Neeson BN, et al. Oral administration of a Salmonella enterica-based vaccine expressing Bacillus anthracis protective antigen confers protection against aerosolized B. anthracis. Infect Immun 2007;75(April (4)):1827-34]. To extend the utility of this approach to humans we constructed variants of S. enterica serovar Typhi Ty21a, an attenuated typhoid vaccine strain licensed for human use, which expressed and exported PA via two distinct plasmid-based transport systems: the Escherichia coli HlyA haemolysin and the S. Typhi ClyA export apparatus. Murine immunogenicity studies confirmed the ability of these constructs, especially Ty21a expressing the ClyA-PA fusion protein, to stimulate strong PA-specific immune responses following intranasal immunization. These responses were further enhanced by a subsequent boost with either parenterally delivered recombinant PA or the licensed US human alum-adsorbed anthrax vaccine (AVA). Anthrax toxin neutralizing antibody responses using this prime-boost regimen were rapid, vigorous and broad in nature. The results of this study demonstrate the feasibility of employing a mucosal prime with a licensed Salmonella Typhi vaccine strain followed by a parenteral protein boost to stimulate rapid protective immunity against anthrax.

The p40 Subunit of Interleukin (IL)-12 Promotes Stabilization and Export of the p35 Subunit

PubMed Central

Jalah, Rashmi; Rosati, Margherita; Ganneru, Brunda; Pilkington, Guy R.; Valentin, Antonio; Kulkarni, Viraj; Bergamaschi, Cristina; Chowdhury, Bhabadeb; Zhang, Gen-Mu; Beach, Rachel Kelly; Alicea, Candido; Broderick, Kate E.; Sardesai, Niranjan Y.; Pavlakis, George N.; Felber, Barbara K.

2013-01-01

IL-12 is a 70-kDa heterodimeric cytokine composed of the p35 and p40 subunits. To maximize cytokine production from plasmid DNA, molecular steps controlling IL-12p70 biosynthesis at the posttranscriptional and posttranslational levels were investigated. We show that the combination of RNA/codon-optimized gene sequences and fine-tuning of the relative expression levels of the two subunits within a cell resulted in increased production of the IL-12p70 heterodimer. We found that the p40 subunit plays a critical role in enhancing the stability, intracellular trafficking, and export of the p35 subunit. This posttranslational regulation mediated by the p40 subunit is conserved in mammals. Based on these findings, dual gene expression vectors were generated, producing an optimal ratio of the two subunits, resulting in a ∼1 log increase in human, rhesus, and murine IL-12p70 production compared with vectors expressing the wild type sequences. Such optimized DNA plasmids also produced significantly higher levels of systemic bioactive IL-12 upon in vivo DNA delivery in mice compared with plasmids expressing the wild type sequences. A single therapeutic injection of an optimized murine IL-12 DNA plasmid showed significantly more potent control of tumor development in the B16 melanoma cancer model in mice. Therefore, the improved IL-12p70 DNA vectors have promising potential for in vivo use as molecular vaccine adjuvants and in cancer immunotherapy. PMID:23297419

Inhibition of CRM1-mediated nuclear export of transcription factors by leukemogenic NUP98 fusion proteins.

PubMed

Takeda, Akiko; Sarma, Nayan J; Abdul-Nabi, Anmaar M; Yaseen, Nabeel R

2010-05-21

NUP98 is a nucleoporin that plays complex roles in the nucleocytoplasmic trafficking of macromolecules. Rearrangements of the NUP98 gene in human leukemia result in the expression of numerous fusion oncoproteins whose effect on nucleocytoplasmic trafficking is poorly understood. The present study was undertaken to determine the effects of leukemogenic NUP98 fusion proteins on CRM1-mediated nuclear export. NUP98-HOXA9, a prototypic NUP98 fusion, inhibited the nuclear export of two known CRM1 substrates: mutated cytoplasmic nucleophosmin and HIV-1 Rev. In vitro binding assays revealed that NUP98-HOXA9 binds CRM1 through the FG repeat motif in a Ran-GTP-dependent manner similar to but stronger than the interaction between CRM1 and its export substrates. Two NUP98 fusions, NUP98-HOXA9 and NUP98-DDX10, whose fusion partners are structurally and functionally unrelated, interacted with endogenous CRM1 in myeloid cells as shown by co-immunoprecipitation. These leukemogenic NUP98 fusion proteins interacted with CRM1, Ran, and the nucleoporin NUP214 in a manner fundamentally different from that of wild-type NUP98. NUP98-HOXA9 and NUP98-DDX10 formed characteristic aggregates within the nuclei of a myeloid cell line and primary human CD34+ cells and caused aberrant localization of CRM1 to these aggregates. These NUP98 fusions caused nuclear accumulation of two transcription factors, NFAT and NFkappaB, that are regulated by CRM1-mediated export. The nuclear entrapment of NFAT and NFkappaB correlated with enhanced transcription from promoters responsive to these transcription factors. Taken together, the results suggest a new mechanism by which NUP98 fusions dysregulate transcription and cause leukemia, namely, inhibition of CRM1-mediated nuclear export with aberrant nuclear retention of transcriptional regulators.

Reporter Dyes Demonstrate Functional Expression of Multidrug Resistance Proteins in the Marine Flatworm Macrostomum lignano: The Sponge-Derived Dye Ageladine A Is Not a Substrate of These Transporters

PubMed Central

Tietje, Kristin; Rivera-Ingraham, Georgina; Petters, Charlotte; Abele, Doris; Dringen, Ralf; Bickmeyer, Ulf

2013-01-01

The marine plathyhelminth Macrostomum lignano was recently isolated from Adriatic shore sediments where it experiences a wide variety of environmental challenges, ranging from hypoxia and reoxygenation, feeding on toxic algae, to exposure to anthropogenic contaminants. As multidrug resistance transporters constitute the first line of defense against toxins and toxicants we have studied the presence of such transporters in M. lignano in living animals by applying optical methods and pharmacological inhibitors that had been developed for mammalian cells. Application of the MDR1 inhibitor Verapamil or of the MRP1 inhibitors MK571 or Probenecid increased the intracellular fluorescence of the reporter dyes Fura-2 am, Calcein am, Fluo-3 am in the worms, but did not affect their staining with the dyes Rhodamine B, CMFDA or Ageladine A. The marine sponge alkaloid Ageladine A remained intracellularly trapped for several days in the worms, suggesting that it does not serve as substrate of multidrug resistance exporters. In addition, Ageladine A did not affect multidrug resistance-associated protein (MRP)-mediated dye export from M. lignano or the MRP1-mediated glutathione (GSH) export from cultured rat brain astrocytes. The data obtained demonstrate that life-imaging is a useful tool to address physiological drug export from intact marine transparent flatworms by using multiphoton scanning microscopy. PMID:24135911

Ecological response to collapse of the biological pump following the mass extinction at the Cretaceous-Paleogene boundary

NASA Astrophysics Data System (ADS)

Vellekoop, Johan; Woelders, Lineke; Açikalin, Sanem; Smit, Jan; van de Schootbrugge, Bas; Yilmaz, Ismail Ö.; Brinkhuis, Henk; Speijer, Robert P.

2017-02-01

It is commonly accepted that the mass extinction associated with the Cretaceous-Paleogene (K-Pg) boundary (˜ 66 Ma) is related to the environmental effects of a large extraterrestrial impact. The biological and oceanographic consequences of the mass extinction are, however, still poorly understood. According to the Living Ocean model, the biological crisis at the K-Pg boundary resulted in a long-term reduction of export productivity in the early Paleocene. Here, we combine organic-walled dinoflagellate cyst (dinocyst) and benthic foraminiferal analyses to provide new insights into changes in the coupling of pelagic and benthic ecosystems. To this end, we perform dinocyst and benthic foraminiferal analyses on the recently discovered Tethyan K-Pg boundary section at Okçular, Turkey, and compare the results with other K-Pg boundary sites in the Tethys. The post-impact dominance of epibenthic morphotypes and an increase of inferred heterotrophic dinocysts in the early Paleocene at Okçular are consistent with published records from other western Tethyan sites. Together, these records indicate that during the early Paleocene more nutrients remained available for the Tethyan planktonic community, whereas benthic communities were deprived of food. Hence, in the post-impact phase the reduction of export productivity likely resulted in enhanced recycling of nutrients in the upper part of the water column, all along the western Tethyan margins.

Advanced Concepts in Josephson Junction Reflection Amplifiers

NASA Astrophysics Data System (ADS)

Lähteenmäki, Pasi; Vesterinen, Visa; Hassel, Juha; Paraoanu, G. S.; Seppä, Heikki; Hakonen, Pertti

2014-06-01

Low-noise amplification at microwave frequencies has become increasingly important for the research related to superconducting qubits and nanoelectromechanical systems. The fundamental limit of added noise by a phase-preserving amplifier is the standard quantum limit, often expressed as noise temperature . Towards the goal of the quantum limit, we have developed an amplifier based on intrinsic negative resistance of a selectively damped Josephson junction. Here we present measurement results on previously proposed wide-band microwave amplification and discuss the challenges for improvements on the existing designs. We have also studied flux-pumped metamaterial-based parametric amplifiers, whose operating frequency can be widely tuned by external DC-flux, and demonstrate operation at pumping, in contrast to the typical metamaterial amplifiers pumped via signal lines at.

Interaction of HTLV-1 Tax protein with calreticulin: implications for Tax nuclear export and secretion.

PubMed

Alefantis, Timothy; Flaig, Katherine E; Wigdahl, Brian; Jain, Pooja

2007-05-01

Human T cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The HTLV-1 transcriptional transactivator protein Tax plays an integral role in virus replication and disease progression. Traditionally, Tax is described as a nuclear protein where it performs its primary role as a transcriptional transactivator. However, recent studies have clearly shown that Tax can also be localized to the cytoplasm where it has been shown to interact with a number of host transcription factors most notably NF-kappaB, constitutive expression of which is directly related to the T cell transforming properties of Tax in ATL patients. The presence of a functional nuclear export signal (NES) within Tax and the secretion of full-length Tax have also been demonstrated previously. Additionally, release of Tax from HTLV-1-infected cells and the presence of cell-free Tax was demonstrated in the CSF of HAM/TSP patients suggesting that the progression to HAM/TSP might be mediated by the ability of Tax to function as an extracellular cytokine. Therefore, in both ATL and HAM/TSP Tax nuclear export and nucleocytoplasmic shuttling may play a critical role, the mechanism of which remains unknown. In this study, we have demonstrated that the calcium binding protein calreticulin interacts with Tax by co-immunoprecipitation. This interaction was found to localize to a region at or near the nuclear membrane. In addition, differential expression of calreticulin was demonstrated in various cell types that correlated with their ability to retain cytoplasmic Tax, particularly in astrocytes. Finally, a comparison of a number of HTLV-1-infected T cell lines to non-infected T cells revealed higher expression of calreticulin in infected cells implicating a direct role for this protein in HTLV-1 infection.

Potential depletion of surface water in the Colorado River and agricultural drains by groundwater pumping in the Parker-Palo Verde-Cibola area, Arizona and California

USGS Publications Warehouse

Leake, Stanley A.; Owen-Joyce, Sandra J.; Heilman, Julian A.

2013-01-01

Water use along the lower Colorado River is allocated as “consumptive use,” which is defined to be the amount of water diverted from the river minus the amount that returns to the river. Diversions of water from the river include surface water in canals and water removed from the river by pumping wells in the aquifer connected to the river. A complication in accounting for water pumped by wells occurs if the pumping depletes water in drains and reduces measured return flow in those drains. In that case, consumptive use of water pumped by the wells is accounted for in the reduction of measured return flow. A method is needed to understand where groundwater pumping will deplete water in the river and where it will deplete water in drains. To provide a basis for future accounting for pumped groundwater in the Parker-Palo Verde-Cibola area, a superposition model was constructed. The model consists of three layers of finite-difference cells that cover most of the aquifer in the study area. The model was run repeatedly with each run having a pumping well in a different model cell. The source of pumped water that is depletion of the river, expressed as a fraction of the pumping rate, was computed for all active cells in model layer 1, and maps were constructed to understand where groundwater pumping depletes the river and where it depletes drains. The model results indicate that if one or more drains exist between a pumping well location and the river, nearly all of the depletion will be from drains, and little or no depletion will come from the Colorado River. Results also show that if a well pumps on a side of the river with no drains in the immediate area, depletion will come from the Colorado River. Finally, if a well pumps between the river and drains that parallel the river, a fraction of the pumping will come from the river and the rest will come from the drains. Model results presented in this report may be considered in development or refinement of strategies for accounting for groundwater pumping in the river aquifer connected to the Colorado River in the study area.

cFLIP overexpression in T cells in thymoma-associated myasthenia gravis

PubMed Central

Belharazem, Djeda; Schalke, Berthold; Gold, Ralf; Nix, Wilfred; Vitacolonna, Mario; Hohenberger, Peter; Roessner, Eric; Schulze, Torsten J; Saruhan-Direskeneli, Güher; Yilmaz, Vuslat; Ott, German; Ströbel, Philipp; Marx, Alexander

2015-01-01

Objective The capacity of thymomas to generate mature CD4+ effector T cells from immature precursors inside the tumor and export them to the blood is associated with thymoma-associated myasthenia gravis (TAMG). Why TAMG(+) thymomas generate and export more mature CD4+ T cells than MG(−) thymomas is unknown. Methods Unfixed thymoma tissue, thymocytes derived thereof, peripheral blood mononuclear cells (PBMCs), T-cell subsets and B cells were analysed using qRT-PCR and western blotting. Survival of PBMCs was measured by MTT assay. FAS-mediated apoptosis in PBMCs was quantified by flow cytometry. NF-κB in PBMCs was inhibited by the NF-κB-Inhibitor, EF24 prior to FAS-Ligand (FASLG) treatment for apoptosis induction. Results Expression levels of the apoptosis inhibitor cellular FLICE-like inhibitory protein (c-FLIP) in blood T cells and intratumorous thymocytes were higher in TAMG(+) than in MG(−) thymomas and non-neoplastic thymic remnants. Thymocytes and PBMCs of TAMG patients showed nuclear NF-κB accumulation and apoptosis resistance to FASLG stimulation that was sensitive to NF-κB blockade. Thymoma removal reduced cFLIP expression in PBMCs. Interpretation We conclude that thymomas induce cFLIP overexpression in thymocytes and their progeny, blood T cells. We suggest that the stronger cFLIP overexpression in TAMG(+) compared to MG(−) thymomas allows for the more efficient generation of mature CD4+ T cells in TAMG(+) thymomas. cFLIP overexpression in thymocytes and exported CD4+ T cells of patients with TAMG might contribute to the pathogenesis of TAMG by impairing central and peripheral T-cell tolerance. PMID:26401511