bluuebunny/crossref_metadata_2025 huggingface dataset:
Human T-Cell Leukemia Virus Type 1 Tax Relieves Repression of Proliferating Cell Nuclear Antigen Gene Expression
Authors: Edwards Dustin C. , Marriott Susan J.
Published: September 2008

Human T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia. The transforming ability of Tax, the viral oncoprotein, is believed to depend on interactions with cell cycle regulators and on transactivation of genes that control cellular proliferation, including proliferating cell nuclear antigen (PCNA), a cofactor associated with DNA replication and repair. Tax associates with cellular transcription factors to alter their affinity for cognate DNA elements, leading to increased or decreased transcription from that promoter. Although it has been demonstrated that Tax transactivates the PCNA promoter, the mechanism of transcriptional activation is unknown. Here we report a cellular complex that binds specifically to a novel site within the minimal Tax-responsive element of the TATAA-less PCNA promoter. Mutation at this binding site or Tax expression inhibited complex formation and increased promoter activity, suggesting that the complex is a transcriptional repressor. The activation of PCNA gene expression by Tax and consequential decrease in nucleotide excision repair mediated by PCNA overexpression could contribute to the reduced DNA repair capacity and genomic instability observed in HTLV-1-infected cells.
Asymptomatic Human CD4+Cytotoxic T-Cell Epitopes Identified from Herpes Simplex Virus Glycoprotein B
Authors: Chentoufi Aziz Alami , Binder Nicholas R. , Berka Noureddine , Durand Guillaume , Nguyen Alex , Bettahi Ilham , Maillère Bernard , BenMohamed Lbachir
Published: September 2008

The identification of “asymptomatic” (i.e., protective) epitopes recognized by T cells from herpes simplex virus (HSV)-seropositive healthy individuals is a prerequisite for an effective vaccine. Using the PepScan epitope mapping strategy, a library of 179 potential peptide epitopes (15-mers overlapping by 10 amino acids) was identified from HSV type 1 (HSV-1) glycoprotein B (gB), an antigen that induces protective immunity in both animal models and humans. Eighteen groups (G1 to G18) of 10 adjacent peptides each were first screened for T-cell antigenicity in 38 HSV-1-seropositive but HSV-2-seronegative individuals. Individual peptides within the two immunodominant groups (i.e., G4 and G14) were further screened with T cells from HLA-DR-genotyped and clinically defined symptomatic (n= 10) and asymptomatic (n= 10) HSV-1-seropositive healthy individuals. Peptides gB161-175and gB166-180within G4 and gB661-675within G14 recalled the strongest HLA-DR-dependent CD4+T-cell proliferation and gamma interferon production. gB166-180, gB661-675, and gB666-680elicited ex vivo CD4+cytotoxic T cells (CTLs) that lysed autologous HSV-1- and vaccinia virus (expressing gB)-infected lymphoblastoid cell lines. Interestingly, gB166-180and gB666-680peptide epitopes were strongly recognized by CD4+T cells from 10 of 10 asymptomatic patients but not by CD4+T cells from 10 of 10 symptomatic patients (P< 0.0001; analysis of variance posttest). Inversely, CD4+T cells from symptomatic patients preferentially recognized gB661-675(P< 0.0001). Thus, we identified three previously unrecognized CD4+CTL peptide epitopes in HSV-1 gB. Among these, gB166-180and gB666-680appear to be “asymptomatic” peptide epitopes and therefore should be considered in the design of future herpes vaccines.
Crystal Structure of the Avian Reovirus Inner Capsid Protein σA
Authors: Guardado-Calvo Pablo , Vazquez-Iglesias Lorena , Martinez-Costas José , Llamas-Saiz Antonio L. , Schoehn Guy , Fox Gavin C. , Hermo-Parrado X. Lois , Benavente Javier , van Raaij Mark J.
Published: September 2008

Avian reovirus, an important avian pathogen, expresses eight structural and four nonstructural proteins. The structural σA protein is a major component of the inner capsid, clamping together λA building blocks. σA has also been implicated in the resistance of avian reovirus to the antiviral action of interferon by strongly binding double-stranded RNA in the host cell cytoplasm and thus inhibiting activation of the double-stranded RNA-dependent protein kinase. We have solved the structure of bacterially expressed σA by molecular replacement and refined it using data to 2.3-Å resolution. Twelve σA molecules are present in the P1 unit cell, arranged as two short double helical hexamers. A positively charged patch is apparent on the surface of σA on the inside of this helix and mutation of either of two key arginine residues (Arg155 and Arg273) within this patch abolishes double-stranded RNA binding. The structural data, together with gel shift assay, electron microscopy, and sedimentation velocity centrifugation results, provide evidence for cooperative binding of σA to double-stranded RNA. The minimal length of double-stranded RNA required for σA binding was observed to be 14 to 18 bp.
Nuclear Localization of Cytoplasmic Poly(A)-Binding Protein upon Rotavirus Infection Involves the Interaction of NSP3 with eIF4G and RoXaN
Authors: Harb Maya , Becker Michelle M. , Vitour Damien , Baron Carolina H. , Vende Patrice , Brown Spencer C. , Bolte Susanne , Arold Stefan T. , Poncet Didier
Published: September 2008

Rotavirus nonstructural protein NSP3 interacts specifically with the 3′ end of viral mRNAs, with the eukaryotic translation initiation factor eIF4G, and with RoXaN, a cellular protein of yet-unknown function. By evicting cytoplasmic poly(A) binding protein (PABP-C1) from translation initiation complexes, NSP3 shuts off the translation of cellular polyadenylated mRNAs. We show here that PABP-C1 evicted from eIF4G by NSP3 accumulates in the nucleus of rotavirus-infected cells. Through modeling of the NSP3-RoXaN complex, we have identified mutations in NSP3 predicted to interrupt its interaction with RoXaN without disturbing the NSP3 interaction with eIF4G. Using these NSP3 mutants and a deletion mutant unable to associate with eIF4G, we show that the nuclear localization of PABP-C1 not only is dependent on the capacity of NSP3 to interact with eIF4G but also requires the interaction of NSP3 with a specific region in RoXaN, the leucine- and aspartic acid-rich (LD) domain. Furthermore, we show that the RoXaN LD domain functions as a nuclear export signal and that RoXaN tethers PABP-C1 with RNA. This work identifies RoXaN as a cellular partner of NSP3 involved in the nucleocytoplasmic localization of PABP-C1.
Retroviruses Human Immunodeficiency Virus and Murine Leukemia Virus Are Enriched in Phosphoinositides
Authors: Chan Robin , Uchil Pradeep D. , Jin Jing , Shui Guanghou , Ott David E. , Mothes Walther , Wenk Markus R.
Published: September 2008

Retroviruses acquire a lipid envelope during budding from the membrane of their hosts. Therefore, the composition of this envelope can provide important information about the budding process and its location. Here, we present mass spectrometry analysis of the lipid content of human immunodeficiency virus type 1 (HIV-1) and murine leukemia virus (MLV). The results of this comprehensive survey found that the overall lipid content of these viruses mostly matched that of the plasma membrane, which was considerably different from the total lipid content of the cells. However, several lipids are enriched in comparison to the composition of the plasma membrane: (i) cholesterol, ceramide, and GM3; and (ii) phosphoinositides, phosphorylated derivatives of phosphatidylinositol. Interestingly, microvesicles, which are similar in size to viruses and are also released from the cell periphery, lack phosphoinositides, suggesting a different budding mechanism/location for these particles than for retroviruses. One phosphoinositide, phosphatidylinositol 4,5-bisphosphate [PI(4,5)P
2
], has been implicated in membrane binding by HIV Gag. Consistent with this observation, we found that PI(4,5)P
2
was enriched in HIV-1 and that depleting this molecule in cells reduced HIV-1 budding. Analysis of mutant virions mapped the enrichment of PI(4,5)P
2
to the matrix domain of HIV Gag. Overall, these results suggest that HIV-1 and other retroviruses bud from cholesterol-rich regions of the plasma membrane and exploit matrix/PI(4,5)P
2
interactions for particle release from cells.
Coxsackievirus B4 Uses Autophagy for Replication after Calpain Activation in Rat Primary Neurons
Authors: Yoon Seung Yong , Ha Young Eun , Choi Jung Eun , Ahn Jeonghyun , Lee Heuiran , Kweon Hee-Seok , Lee Jee-Young , Kim Dong Hou
Published: September 2008

Coxsackievirus is the most important cause of meningitis and encephalitis in infants; an infection is sometimes fatal or may lead to neurodevelopmental defects. Here, we show that coxsackievirus B4 (CVB4) induces an autophagy pathway for replication in rat primary neurons. Notably, calpain inhibitors reduce autophagosome formation. Conversely, the inhibition of the autophagy pathway with 3-methyladenine inhibits calpain activation. This work reveals, for the first time, that calpain is essential for the autophagy pathway and viral replication in CVB4-infected neurons.
Induction of Cellular Stress Overcomes the Requirement of Herpes Simplex Virus Type 1 for Immediate-Early Protein ICP0 and Reactivates Expression from Quiescent Viral Genomes
Authors: Preston Chris M. , Nicholl Mary Jane
Published: September 2008

Herpes simplex virus type 1 (HSV-1) mutants impaired in the activities of the structural protein VP16 and the immediate-early (IE) proteins ICP0 and ICP4 establish a quiescent infection in human fibroblasts, with most cells retaining an inactive, repressed viral genome for sustained periods in culture. To date, the quiescent state has been considered stable, since it has been reversed only by provision of herpesviral proteins, such as ICP0, not by alteration of the cell physiological state. We report that the interaction of HSV-1 with human fibroblasts can be altered significantly by transient treatment of cultures with sodium arsenite, an inducer of heat shock and oxidative stress, or gramicidin D, a toxin that selectively permeabilizes cell membranes, prior to infection. These regimens stimulated gene expression from IE-deficient HSV-1 mutants in a promoter sequence-independent manner and also overcame the replication defect of ICP0-null mutants. Reactivation of gene expression from quiescent HSV-1 genomes and the resumption of virus replication were observed following addition of arsenite or gramicidin D to cultures. Both agents induced reorganization of nuclear domain 10 structures, the sites of quiescent genomes, but appeared to do so through different mechanisms. The results demonstrate that the physiological state of the cell is important in determining the outcome of infection with IE-deficient HSV-1 and show novel methods for reactivating quiescent HSV-1 in fibroblasts with a high efficiency.
Baseline Resistance of Primary Human Immunodeficiency Virus Type 1 Strains to the CXCR4 Inhibitor AMD3100
Authors: Harrison Jessamina E. , Lynch Jonathan B. , Sierra Luz-Jeannette , Blackburn Leslie A. , Ray Neelanjana , Collman Ronald G. , Doms Robert W.
Published: September 2008

We screened a panel of R5X4 and X4 human immunodeficiency virus type 1 (HIV-1) strains for their sensitivities to AMD3100, a small-molecule CXCR4 antagonist that blocks HIV-1 infection via this coreceptor. While no longer under clinical development, AMD3100 is a useful tool with which to probe interactions between the viral envelope (Env) protein and CXCR4 and to identify pathways by which HIV-1 may become resistant to this class of antiviral agents. While infection by most virus strains was completely blocked by AMD3100, we identified several R5X4 and X4 isolates that exhibited plateau effects: as the AMD3100 concentration was increased, virus infection and membrane fusion diminished to variable degrees. Once saturating concentrations of AMD3100 were achieved, further inhibition was not observed, indicating a noncompetitive mode of viral resistance to the drug. The magnitude of the plateau varied depending on the virus isolate, as well as the cell type used, with considerable variation observed when primary human T cells from different human donors were used. Structure-function studies indicated that the V1/V2 region of the R5X4 HIV-1 isolate DH12 was necessary for AMD3100 resistance and could confer this property on two heterologous Env proteins. We conclude that some R5X4 and X4 HIV-1 isolates can utilize the AMD3100-bound conformation of CXCR4, with the efficiency being influenced by both viral and host factors. Baseline resistance to this CXCR4 antagonist could influence the clinical use of such compounds.
Mapping of the Tacaribe Arenavirus Z-Protein Binding Sites on the L Protein Identified both Amino Acids within the Putative Polymerase Domain and a Region at the N Terminus of L That Are Critically Involved in Binding
Authors: Wilda Maximiliano , Lopez Nora , Casabona Juan Cruz , Franze-Fernandez Maria T.
Published: September 2008

Tacaribe virus (TacV) is the prototype of the New World group of arenaviruses. The TacV genome encodes four proteins: the nucleoprotein (N), the glycoprotein precursor, the polymerase (L), and a RING finger protein (Z). Using a reverse genetics system, we demonstrated that TacV N and L are sufficient to drive transcription and replication mediated by TacV-like RNAs and that Z is a powerful inhibitor of these processes (Lopez et al., J. Virol.
65:
12241-12251, 2001). More recently, we provided the first evidence of an interaction between Z and L and showed that Z's inhibitory activity was dependent on its ability to bind to L (Jácamo et al., J. Virol.
77:
10383-10393, 2003). In the present study, we mapped the TacV Z-binding sites on the 2,210-amino-acid L polymerase. To that end, we performed deletion analysis and point mutations of L and studied the Z-L interaction by coimmunoprecipitation with specific sera. We found that the C-terminal region of L was not essential for the interaction and identified two noncontiguous regions that were critical for binding: one at the N-terminus of L between residues 156 and 292 and a second one in the polymerase domain (domain III). The importance of domain III in binding was revealed by substitutions in D1188 and H1189 within motif A and in each residue of the conserved SDD sequence (residues 1328, 1329, and 1330) within motif C. Our results showed that of the substituted residues, only H1189 and D1329 appeared to be critically involved in binding Z.
Biochemical and Biophysical Characterization of a Chimeric TRIM21-TRIM5α Protein
Authors: Kar Alak Kanti , Diaz-Griffero Felipe , Li Yuan , Li Xing , Sodroski Joseph
Published: September 2008

The tripartite motif (TRIM) protein, TRIM5α, is an endogenous factor in primates that recognizes the capsids of certain retroviruses after virus entry into the host cell. TRIM5α promotes premature uncoating of the capsid, thus blocking virus infection. Low levels of expression and tendencies to aggregate have hindered the biochemical, biophysical, and structural characterization of TRIM proteins. Here, a chimeric TRIM5α protein (TRIM5
Rh
-21R) with a RING domain derived from TRIM21 was expressed in baculovirus-infected insect cells and purified. Although a fraction of the TRIM5
Rh
-21R protein formed large aggregates, soluble fractions of the protein formed oligomers (mainly dimers), exhibited a protease-resistant core, and contained a high percentage of helical secondary structure. Cross-linking followed by negative staining and electron microscopy suggested a globular structure. The purified TRIM5
Rh
-21R protein displayed E3-ligase activity in vitro and also self-ubiquitylated in the presence of ubiquitin-activating and -conjugating enzymes. The purified TRIM5
Rh
-21R protein specifically associated with human immunodeficiency virus type 1 capsid-like complexes; a deletion within the V1 variable region of the B30.2(SPRY) domain decreased capsid binding. Thus, the TRIM5
Rh
-21R restriction factor can directly recognize retroviral capsid-like complexes in the absence of other mammalian proteins.
Identification and Characterization of PWWP Domain Residues Critical for LEDGF/p75 Chromatin Binding and Human Immunodeficiency Virus Type 1 Infectivity
Authors: Shun Ming-Chieh , Botbol Yaïr , Li Xiang , Di Nunzio Francesca , Daigle Janet E. , Yan Nan , Lieberman Judy , Lavigne Marc , Engelman Alan
Published: September 2008

Lens epithelium-derived growth factor (LEDGF)/p75 functions as a bimodal tether during lentiviral DNA integration: its C-terminal integrase-binding domain interacts with the viral preintegration complex, whereas the N-terminal PWWP domain can bind to cellular chromatin. The molecular basis for the integrase-LEDGF/p75 interaction is understood, while the mechanism of chromatin binding is unknown. The PWWP domain is homologous to other protein interaction modules that together comprise the Tudor clan. Based on primary amino acid sequence and three-dimensional structural similarities, 24 residues of the LEDGF/p75 PWWP domain were mutagenized to garner essential details of its function during human immunodeficiency virus type 1 (HIV-1) infection. Mutating either Trp-21 or Ala-51, which line the inner wall of a hydrophobic cavity that is common to Tudor clan members, disrupts chromatin binding and virus infectivity. Consistent with a role for chromatin-associated LEDGF/p75 in stimulating integrase activity during infection, recombinant W21A protein is preferentially defective for enhancing integration into chromatinized target DNA in vitro. The A51P mutation corresponds to the S270P change in DNA methyltransferase 3B that causes human immunodeficiency, centromeric instability, and facial anomaly syndrome, revealing a critical role for this amino acid position in the chromatin binding functions of varied PWWP domains. Our results furthermore highlight the requirement for a conserved Glu in the hydrophobic core that mediates interactions between other Tudor clan members and their substrates. This initial systematic mutagenesis of a PWWP domain identifies amino acid residues critical for chromatin binding function and the consequences of their changes on HIV-1 integration and infection.
Estimating the Impact of Vaccination on Acute Simian-Human Immunodeficiency Virus/Simian Immunodeficiency Virus Infections
Authors: Petravic Janka , Ribeiro Ruy M. , Casimiro Danilo R. , Mattapallil Joseph J. , Roederer Mario , Shiver John W. , Davenport Miles P.
Published: September 2008

The dynamics of HIV infection have been studied in humans and in a variety of animal models. The standard model of infection has been used to estimate the basic reproductive ratio of the virus, calculated from the growth rate of virus in acute infection. This method has not been useful in studying the effects of vaccination, since, for the vaccines developed so far, early growth rates of virus do not differ between control and vaccinated animals. Here, we use the standard model of viral dynamics to derive the reproductive ratio from the peak viral load and nadir of target cell numbers in acute infection. We apply this method to data from studies of vaccination in SHIV and SIV infection and demonstrate that vaccination can reduce the reproductive ratio by 2.3- and 2-fold, respectively. This method allows the comparison of vaccination efficacies among different viral strains and animal models in vivo.
Recruitment Limitation as a Consequence of Natural Selection for Use of Restricted Feeding Habitats and Predation Risk Taking by Juvenile Fishes
Authors: Walters Carl J. , Juanes Francis
Published: September 2008

Juvenile fishes generally have spatial refuges from predation, and forage in limited but risky areas near refuges. Models of food density dynamics within such limited foraging areas predict that food availability and consumption per time spent feeding should depend strongly on Juvenile density. Selection should act on the time that Juveniles spend foraging, so as to strike a balance between growth and predation risk; we predict that optimum balance will occur at foraging times proportional to the minimum needed to reach viable sizes for later survival and reproduction plus an additional time inversely proportional to the predation risk per time. Combining the food availability and optimal foraging time predictions leads to stock–recruitment patterns similar to classic Beverton–Holt and Ricker forms, depending on how food organisms respond over time. Very strong density-dependent mortality can occur even without noticeable changes in juvenile growth rates or average food densities over large spatial scales, but intraspecific competitive effects should always be evident through density-dependent changes in foraging time.
Replacement of Sulfur, Carbon, and Nitrogen in Tissue of Growing Broad Whitefish (Coregonus nasus) in Response to a Change in Diet Traced by δ34S, δ13C, and δ15N
Authors: Hesslein R.H. , Hallard K.A. , Ramlal P.
Published: September 2008

We monitored the change in the isotope composition of sulfur, carbon, and nitrogen in broad whitefish (Coregonus nasus) tissues in response to a change in the isotope composition of their food. One of two batches of 2.5-yr-old fish raised in the laboratory were given a new food source with different δ34S, δ13C, and δ15N, which were monitored in muscle and liver tissue for 1 yr. A model including change due to tissue accumulation (growth) and metabolic replacement was developed. For all three isotopes, most of the change could be attributed to growth. Metabolic replacement expressed as a turnover rate was only 0.1 −0.2%∙d−1 and was similar for the three isotopes. Although liver tissue was −4.4 and −4.1‰, respectively, for δ34S and δ13C relative to muscle tissue, the response over time to the new food was the same as for muscle. We expect that the complete change in the isotope composition of fish tissue in response to a change in food could take years in slow-growing wild populations. The stable isotope composition would represent a long-term average of the food. In fast-growing fish the rate of change would directly reflect the growth rate.
Ballast Wafer Exchange as a Means of Controlling Dispersal of Freshwater Organisms by Ships
Authors: Locke A. , Reid D. M. , van Leeuwen H. C. , Sprules W.G. , Carlton J. T.
Published: September 2008

During May–December 1990 and March–May 1991, 546 foreign ocean-going vessels entered the Laurentian Great Lakes and upper St. Lawrence River, areas protected by the Great Lakes Ballast Water Control Guidelines. Between 88 and 94% of the vessels exchanged their ballast water with seawater as required by the guidelines. Living representatives of 11 invertebrate phyla were sampled from ballast tanks. Between 14 and 33% of ships that exchanged freshwater ballast in midocean carried living freshwater-tolerant zooplankton at the time of entry to the Seaway, although these included many taxa already found in the Great Lakes. Four freshwater-tolerant zooplankton species that were identified as living specimens in ballast water have apparently not been recorded from the Great Lakes. Voluntary ballast water controls reduced but did not eliminate the risk of species invasion, since some ships did not comply with the guidelines, and even ships that did exchange ballast water could introduce viable freshwater-tolerant organisms into the Great Lakes. About half of the ballast water carried into the Seaway by ocean-going vessels and lakers each year originates from the St. Lawrence River, portions of which are not yet protected by any ballast controls.
Determinants of Postmolt Size in the American Lobster (Homarus americanus). I. D13is the Critical Stage
Authors: Cheng Jin-Hua , Chang Ernest S.
Published: September 2008

Juvenile American lobster (Homarus americanus) were subjected to induced limb autotomy, eyestalk ablation, and manipulation of container space at different molt stages to test the hypothesis that postmolt size is predetermined at the time of deposition of the epicuticle. Molt increments in autotomized animals that regenerated limbs were reduced by 30–40%. Molt increments were reduced by only 5–10% in autotomized animals that did not regenerate limbs. The critical stage for limb regeneration is D13. Depending on the time of limb autotomy, molt intervals were decreased (stages B–C), increased (stages D11and D12), or not affected (stages D0, D13, D2, and D3). Eyestalk ablation shortened the molt interval; the change in molt interval was less when ablation was performed near the end of the current molt cycle. Eyestalk-ablated lobsters had larger molt increments compared with intact controls, especially in the chelipeds. This growth-promoting effect disappeared if ablation was conducted at or after D13. Transferring lobsters from small to large container spaces shortened molt intervals and increased molt increments. Increases in molt increments only occurred when transfers were made before D13. We conclude that postmolt size of lobsters is predetermined at D13when the new epicuticle is produced.
Use of Oceanographic Forecasts and Echosounders to Guide and Enhance an Inshore Gillnet Fishery for Atlantic Cod (Gadus morhua)
Authors: Rose G. A. , Leggett W.C.
Published: September 2008

Vessels fishing with gill nets for Atlantic cod (Gadus morhua) in the northern Gulf of St. Lawrence in 1985 and 1986 concentrated their fishing effort in shallow waters (<50 m) where cod densities were highest (to 0.5/m3). In both years, seasonal trends (June–August) in mean daily deployment depths of gill nets were positively correlated with mean depths at which cod were surveyed (1985, r = 0.71; 1986, r = 0.51, Ps < 0.05). Daily catch rate variability of individual vessels was accounted for by fish "flux" adjacent to nets (44%), vessel operator skill (19%), and "flux"–skill interaction (8%; total R2 = 0.71). A guided vessel directed to fish at sites predicted to have high fish flux (located down-current from high-density cod aggregations identified by echosounding within depth ranges forecast to be favorable to cod by "rule of thumb" wind-based oceanographic models) had higher catch rates (mean 1.3 t/d) than the fleet average and its own average fishing without guidance (means 0.4 t/d, P's < 0.05). Directed searches were of shorter duration (mean 0.5 h) than searches conducted at random (1.5 h). Combined use of echosounders and air–sea-based forecasts of cod distribution could help stabilize catch rates, especially at times of poor fish availability inshore.
A Reappraisal of Phenotypic Variation inDaphnia galeata mendotae: The Role of Interspecific Hybridization
Authors: Taylor Derek J. , Hebert Paul D. N.
Published: September 2008

Seasonal, within-lake, and among-lake variation in helmet size and shape has been reported for Daphnia galeata mendotae. The genetic component of this variation was assessed by an allozyme and morphological reanalysis of nine previously studied sites. Genetic associations with morphological variation were found, but all were attributed to D. galeata mendotae × Daphnia rosea hybridization or to taxonomic confusion with D. rosea. The existence of hybrids accounted for spatial polymorphism both within and among lakes. In addition, hybrids contributed to seasonal variation as they increased in abundance from summer to winter. Nevertheless, cyclomorphosis in D. galeata mendotae s.s was real and due largely to phenotypic plasticity. Local hybridization between D. galeata mendotae and D. rosea was widespread from the midwestern states to Vermont. The distributions and habitat associations of D. galeata mendotae and D. rosea indicated that ecological segregation at the lake level acts as an important reproductive isolating mechanism. Reinterpretation of previous studies, however, suggests that disturbance followed by one or a few hybridization events often leads to hybrid domination of a lake.
Reproductive Cycle and Fecundity of Primiparous and Multiparous Female Snow Crab, Chionoecetes opilio, in the Northwest Gulf of Saint Lawrence
Authors: Sainte-Marie Bernard
Published: September 2008

A total of 1 691 mature female Chionoecetes opilio was collected by beam trawl in Baie Sainte-Marguerite on seven occasions from April 1991 to May 1992. Quantitative analyses of ovaries, brood, eggs, and spermathecal contents were performed on a subsample of 318 females. Females moulted to maturity in March and April. Development of ovaries and brood were phased and lasted 24–27 mo. Eggs hatched mainly from April to June. Spermathecae contained zero to three ejaculates, and the mean blotted weight of freshly deposited ejaculates was significantly greater in primiparous (0.513 g) than in multiparous (0.249 g) females. The weight of stored ejaculates decreased over time independently of spawning events. Fecundity was correlated positively with carapace width and negatively with mean egg diameter. Eggs of primiparous females were on average 1.4–2.7% larger, but 16.4–22.7% fewer per brood, than those of multiparous females. Some large multiparous females had small broods, due possibly to lesser fertility. Females probably hatch only two broods in a lifetime and produce a total of 81 630 to 83 143 larvae at 57.4 mm carapace width. Primiparous females apparently contribute >40% of larvae produced by the population.
Use of Parasites for Separating Stocks of Greenland Halibut (Reinhardtius hippoglossoides) in the Canadian Northwest Atlantic
Authors: Arthur J. R. , Albert E.
Published: September 2008

The usefulness of parasites as biological tags for stocks of Greenland halibut (Reinharditius hippoglossoides) occurring off the Atlantic coast of Canada and in the Gulf of St. Lawrence was investigated. Nonparametric discriminant function analyses of eight collections comprising a total of 231 large fish (>39 cm) using counts for selected parasite taxa (Corynosoma strumosum juvenile, Otodistomum sp. metacercaria, Contracaecinea spp. larva, Anisakis simplex larva, and Pseudoterranova decipiens larva) gave accurate results (almost 100% correct classification) for the separation of fish from the Gulf of St. Lawrence from those collected from adjacent areas of the Saguenay Fjord and the Atlantic Ocean off Labrador. Little, if any, mixing of subadult or adult fish apparently occurs among these areas.
Gammarus lawrencianus (Amphipoda) as Intermediate Host of Echinorhynchus salmonis (Acanthocephala) in an Estuarine Environment
Authors: Measures Lena N. , Bossé Luci
Published: September 2008

Echinorhynchus salmonis (Acanthocephala) were found in the stomach of a rainbow smelt, Osmerus mordax (Salmoniformes), collected at the confluence of the Aux-Outardes River and St. Lawrence River in Quebec. One acanthocephalan occupied the haemocoel of a recently ingested amphipod (Gammarus lawrencianus), while two other specimens were free in the stomach. This is the first report of G. lawrencianus, an estuarine amphipod, as intermediate host of E. salmonis. This is also the first report of transmission of E. salmonis in an estuarine environment in North America.
Assessment of Phytoplankton Nutrient Limitation in Productive Waters: Application of Dilution Bioassays
Authors: Carrick Hunter J. , Schelske Claire L. , Aldridge Frederick J. , Coveney Michael F.
Published: September 2008

Excessive nutrient loads to aquatic systems can complicate otherwise predictable relationships between nutrient concentrations and phytoplankton biomass. We conducted six bioassays on surface phytoplankton assemblages collected from productive Lake Apopka, Florida, to measure the effect of nutrient reduction on phytoplankton growth and nutritional state. Lake water was mixed with one of three diluents to create a gradient of ambient nutrient concentrations; nitrogen (N) and phosphorus (P) limitation at each level of dilution was evaluated in a 2 × 2 factorial design. While the addition of N clearly increased the growth of phytoplankton in undiluted Lake Apopka water, the phytoplankton became more P limited with the reduction of particles (30–60% dilution). Regression of algal yields onto total P concentrations from our bottle experiments indicated that an 8 μg∙L−1change in P leads to only a 1 μg∙L−1change in chlorophyll yield, probably due to the high concentration of P in the lake. Because dilution influences factors in addition to ambient nutrient concentrations, results obtained with the technique must be carefully evaluated. Despite this, reduction of particles to improve water quality may, in concept, be a reasonable management scheme in lakes where a large fraction of the nutrients is particulate.
Importance of Runoff and Winter Anoxia to the P and N Dynamics of a Beaver Pond
Authors: Devito Kevin J. , Dillon Peter J.
Published: September 2008

Annual retentions of total phosphorus (TP) (−11%) and total nitrogen (TN) (−5%) of a headwater beaver pond in central Ontario were low during 1987–88. Annually, inputs exceeded outputs of total reactive P (71%) and NO3–N (35%), and outputs exceeded inputs of total unreactive P (−33%) and total organic N (−26%), and inputs approximated outputs of NH4–N (−8%). Seasonal trends in P and N retention were inversely correlated with runoff. Monthly retention was weakly positively related to average water temperature and redox potential (ORP). Positive monthly retention coincided with low runoff and high biotic assimilation during the growing season. Winter ice cover was associated with undetectable dissolved oxygen and low ORP and increased levels of P and N, particularly NH4–N. High levels of P and N in the surface water during winter were coupled with increased runoff and potentially low biotic assimilation resulting in a net release of TP and TN. Large flow-through of waterborne inputs and flushing of regenerated P and N occurred during peak snowmelt resulting in low annual retention. Initial accumulation of flooded forest material and input of organic matter by beaver may be very important to P and N dynamics, representing a long-term source of nutrients to the pond water and outflow.
Photosynthetic Carbon Assimilation and Allocation by Surface and Deep Phytoplankton in Colpoys Bay (Western Georgian Bay)
Authors: Smith Ralph E. H. , Maly Julie A.
Published: September 2008

We used reciprocal shift experiments to test the hypothesis that surface (i.e., epilimnetic) and deep (i.e., metalimnetic) phytoplankton from thermally stratified Colpoys Bay (Georgian Bay) would differ in their photosynthetic and biosynthetic patterns. A deep chlorophyll maximum existed in the metalimnion during the first half of the stratified season. Surface phytoplankton always had higher light-saturated rates of photosynthesis [Formula: see text] and photoadaptation parameters (Ik) than did deep phytoplankton, but most of the difference was due to the proximate effect of temperature on [Formula: see text], and not to true photoadaptation. Neither sensitivity to photoinhibition (limited) nor light-utilization efficiency (α) varied between surface and deep populations. Depth-related differences in photosynthate allocation to protein, total lipid, neutral lipid, and phospholipid were significant according to ANOVA but were nonetheless small compared with other sources of variation. Photosynthetic rates were generally low, while allocation to polysaccharide and neutral lipid was large, suggesting a relatively poor physiological condition of the phytoplankton. Short-term dynamics (days to weeks) of water masses in Colpoys Bay, and probably many other nearshore Great Lakes locations, appear sufficient to preclude strong vertical differentiation of phytoplankton physiology.
Emerging Role of Atmospheric Nitrogen Deposition in Coastal Eutrophication: Biogeochemical and Trophic Perspectives
Authors: Paerl Hans W.
Published: September 2008

Approximately half of the world's oceanic primary production originates in coastal waters which are often nitrogen (N) limited and sensitive to externally supplied N. Among anthropogenically generated N sources, atmospheric deposition of N (ADN) (as wet- and dryfall) contributes 10–50% of the total external N load at present (20–100 mmol N∙m−2∙yr−1), and this will increase. At current levels, atmospheric N inputs lead to a significant enhancement of CO2 fixation and chlorophyll a production. ADN may play a central role in mediating "new" production in coastal oceans downwind of emissions. Alterations in productivity, nutrient flux, and trophic changes have recently been documented from the Baltic and North seas, the Northwest Atlantic seaboard, and the Western Mediterranean. The recently noted "epidemic" of toxic and nuisance phytoplankton blooms is possibly due to accelerating nutrient loading inshore, especially from atmospheric sources. A need exists to spatially and temporally evaluate, quantify, and potentially manage this nutrient-production interaction on regional and larger synoptic scales. Aircraft- and satellite-based multiband ocean color remote sensing (SeaWiFS) will help characterize and link the spatial-temporal interactions of this (and other) "new" nutrient inputs with biogeochemical and trophic changes in coastal and offshore waters.
Aggregative Mating of Tanner Crabs,Chionoecetes bairdi
Authors: Stevens Bradley G. , Haaga Jan A. , Donaldson William E.
Published: September 2008

In April 1991, a high-density mating aggregation of Tanner crabs, Chionoecetes bairdi, was discovered at 150 m depth near Kodiak, Alaska, with the research submersible DSRV Delta. The aggregation consisted primarily of oldshell, multiparous female crabs which formed mounds 1–2 m in diameter, 0.5–1.0 m high, and spaced at intervals of 1–2 m. Mounds contained hundreds of crabs each at densities > 100∙m−2. Male crabs were found mating with females at the periphery of the aggregation, with sex ratios (male to female) varying from 1:10 to 1:100. The entire aggregation covered an area of about 2.2 ha and included approximately 100 000 crabs. No mounds were observed in May 1992, but recently spawned female crabs were found buried in the sediment at high densities (up to 2.2∙m−2) over an area of about 25 ha, suggesting that aggregation had reoccurred at the same location. Newly ovigerous females exhibited a cycle of nocturnal activity and diurnal burial whereas unmated crabs remained exposed on the sediment surface. Many body parts and presumably dead crabs were observed. We conclude that aggregative mating is a major mode of reproduction in the family Majidae.
Stable Carbon and Nitrogen Isotopic Analysis of the Plankton Food Web in a Subarctic Lake
Authors: Gu Binhe , Schell Donald M. , Alexander Vera
Published: September 2008

Carbon and nitrogen stable isotopes (δ13C and δ15N) were used to track energy flow and nutrient cycling pathways in the plankton food web of a subarctic Alaskan lake. Results indicated that planktonic primary production was the major energy source fueling the zooplankton community. In spring, δ15N of Daphnia was strongly influenced by atmospheric nitrogen derived from a N2-fixing blue-green algal bloom. In winter, δ13C evidence suggested that phytoplankton comprised a small fraction (~15%) of particulate organic matter (POM) in the water column, largely due to low primary productivity. The disparity between δ13C of POM and Daphnia in winter may result from preferential assimilation of isotopically light algal carbon from POM. Nitrogen isotope values showed that Heterocope, a presumed carnivore, probably relied heavily on POM as a nutrient source. In common with some arctic lakes, the δ15N data showed less than three trophic levels in this plankton food web. The energy transfer pathways and trophic levels revealed a simple plankton trophic structure in this subarctic lacustrine system.
Cadmium, Metal-binding Proteins, and Growth in Bluegill (Lepomis macrochirus) Exposed to Contaminated Sediments from the Upper Mississippi River Basin
Authors: Cope W. Gregory , Wiener James G. , Steingraeber Mark T. , Atchison Gary J.
Published: September 2008

We exposed juvenile bluegill (Lepomis macrochirus) to ~1000 mg∙L−1 of continuously suspended river sediment in a 28-d test with six treatments (randomized block with one sediment-free control and five sediments ranging from 1.3 to 21.4 μg Cd∙g dry weight−1). Each treatment had three replicates, each with 25 fish. Growth was reduced by exposure to suspended sediment, probably due to physical effects of sediment on feeding and to toxicity in the treatment with the greatest concentrations of metals. Mean whole-body concentrations of cadmium (0.04–0.14 μg∙g wet weight−1) were correlated with cadmium concentration in filtered water (8–72 ng∙L−1), suspended sediment (0.61–16.8 μg∙L−1), and bulk sediment. The concentration of hepatic nonthionein cytosolic cadmium (cadmium not bound by metal-binding proteins, MBP) in fish exposed to the two most contaminated sediments exceeded that in controls. The mean concentration of hepatic MBP was correlated with cadmium concentration in filtered water, suspended sediment, bulk sediment, and whole fish. Whole-body cadmium concentration was the most sensitive indicator of cadmium exposure, with lowest observed effect concentrations of 1.9 μg Cd∙L−1 for suspended sediment and 13 ng Cd∙L−1 for filtered water. Sediment-associated cadmium was less available than waterborne cadmium for uptake by fish.
PCB Concentration Trends in Lake Michigan Coho (Oncorhynchus kisutch) and Chinook Salmon (O.tshawytscha)
Authors: Stow Craig A. , Carpenter Stephen R. , Amrhein James F.
Published: September 2008

Lake Michigan coho (Oncorhynchus kisutch) and chinook salmon (O. tshawytscha) PCB concentration data from 1974 to 1990 were examined. The mean and variance of the concentrations exhibited a generally decreasing pattern in each species. Three alternative models, an exponential decay model, a double exponential decay model, and an exponential decay model with a nonzero asymptote, were fit to the data. The double exponential model provided the best fit for both species. Estimated rate coefficients for this model indicated that an early rapid decrease in PCBs has slowed and that PCBs may currently be increasing in coho and chinook salmon. These PCB increases may be the indirect result of a decline in the alewife (Alosa pseudoharengus) forage base that occurred during the 1980s. Parameter estimates from the nonzero asymptote model suggested that PCB levels may not decrease substantially below the current 2 mg/kg FDA action level. Significant decreases of PCB concentrations in coho and chinook salmon may depend on fisheries management practices that promote higher fish growth rates.
Arctic char (Salvelinus alpinus) and Rainbow Trout (Oncorhynchus mykiss) Differ in Their Growth and Lipid Metabolism in Response to Dietary Polyunsaturated Fatty Acids
Authors: Yang Xiuwei , Dick Terry A.
Published: September 2008

Juvenile Arctic char (Salvelinus alpinus) and rainbow trout (Oncorhynchus mykiss) were fed one commercial and three casein-based diets varying in amounts of 18:2n-6 and 18:3n-3, two polyunsaturated fatty acids (PUFA), for 12 wk at 10 °C. Both species on the diet without PUFA had lower specific growth rate (SGR) but higher percentages of liver neutral lipids and 20:3n-9 in liver polar lipids than those fed high PUFA content or commercial diets. Regardless of the amount of 18:3n-3 in the diets, the percentages of 20:3n-9 and 20:3n-9/22:6n-3 in liver or muscle polar lipids were significantly lower in char than in trout; values in trout were lower than reported in other studies. These findings question the use of 20:3n-9/22:6n-3 as an indicator of essential fatty acid status for both species. Different levels of 20:3n-9, 22:6n-3, and other PUFAs in tissue polar lipids of the two species suggest that Δ6 and/or Δ5 desaturases are less efficient in char. While trout fed test diets had slightly less 20:4n-6 than wild trout, char had 1/10 less 20:4n-6 in muscle polar and total lipids than wild char, suggesting that char may require n-6 and n-3 PUFA in their diet for optimal performance.
Substrate-induced changes in the density of peptide transporter PEPT1 expressed inXenopusoocytes
Authors: Mertl Manuela , Daniel Hannelore , Kottra Gabor
Published: September 2008

The adaptation of the capacity of the intestinal peptide transporter PEPT1 to varying substrate concentrations may be important with respect to its role in providing bulk quantities of amino acids for growth, development, and other nutritional needs. In the present study, we describe a novel phenomenon of the regulation of PEPT1 in the Xenopus oocyte system. Using electrophysiological and immunofluorescence methods, we demonstrate that a prolonged substrate exposure of rabbit PEPT1 (rPEPT1) caused a retrieval of transporters from the membrane. Capacitance as a measure of membrane surface area was increased in parallel with the increase in rPEPT1-mediated transport currents with a slope of ∼5% of basal surface per 100 nA. Exposure of oocytes to the model peptide Gly-l-Gln for 2 h resulted in a decrease in maximal transport currents with no change of membrane capacitance. However, exposure to substrate for 5 h decreased transport currents but also, in parallel, surface area by endocytotic removal of transporter proteins from the surface. The reduction of the surface expression of rPEPT1 was confirmed by presteady-state current measurements and immunofluorescent labeling of rPEPT1. A similar simultaneous decrease of current and surface area was also observed when endocytosis was stimulated by the activation of PKC. Cytochalasin D inhibited all changes evoked by either dipeptide or PKC stimulation, whereas the PKC-selective inhibitor bisindolylmaleimide only affected PKC-stimulated endocytotic processes but not substrate-dependent retrieval of rPEPT1. Coexpression experiments with human Na+-glucose transporter 1 (hSGLT1) revealed that substrate exposure selectively affected PEPT1 but not the activity of hSGLT1.
A fluorimetry-based ssYFP secretion assay to monitor vasopressin-induced exocytosis in LLC-PK1 cells expressing aquaporin-2
Authors: Nunes Paula , Hasler Udo , McKee Mary , Lu Hua A. J. , Bouley Richard , Brown Dennis
Published: September 2008

Vasopressin (VP)-induced exocytosis was dissected in native and aquaporin-2 (AQP2)-expressing renal LLC-PK1 cells by a fluorimetric exocytosis assay based on soluble secreted yellow fluorescent protein (ssYFP). YFP was targeted to the secretory pathway by addition of an 18-amino acid signal peptide from hen egg white lysozyme. Immunofluorescence labeling, together with analysis of Alexa 555-dextran internalization, revealed that ssYFP is exclusively located in the secretory pathway. Immunofluorescence and immunogold electron microscopy showed significant colocalization of ssYFP and AQP2. Fluorimetry and Western blot analysis demonstrated similar constitutive ssYFP secretion in native LLC-PK1 and AQP2-expressing cells. In AQP2-expressing cells, a twofold increase in ssYFP secretion was observed within 15 min of VP stimulation. This transient burst of ssYFP secretion was abolished by the PKA inhibitor H-89 and was not observed in native cells. The endocytotic inhibitor methyl-β-cyclodextrin, which also promotes membrane accumulation of AQP2, had no effect on ssYFP secretion. Although cells expressing phosphorylation-deficient AQP2-S256A showed significantly lower baseline levels of constitutive secretion, VP induced a significant increase in exocytosis. Our data indicate that 1) this assay can monitor exocytosis in cultured epithelial cells, 2) VP has an acute stimulatory effect on ssYFP secretion in AQP2-expressing, but not native, cells, and 3) phosphorylation of AQP2 at S256 may be involved in the regulation of constitutive AQP2 exocytosis and play only a minor role in the VP-induced burst. These results support the idea that, in addition to its role in reducing AQP2 endocytosis, VP increases AQP2 exocytosis.