Elevated cadmium accumulation in marine organisms from Terra Nova Bay (Antarctica)

 As a contribution towards identification of the principal environmental factors involved in cadmium accumulation in Antarctic marine organisms and the establishment of a baseline near the Italian Antarctic Station “Baia Terra Nova”, surface sediments, plankton and benthic organisms were studied in coastal waters of Terra Nova Bay (Ross Sea). The cadmium content of sediments was similar to that regarded as background in most marine coastal areas, whereas in surface water, phyto- and zooplankton it was similar to values measured in areas of enhanced upwelling. Algal and animal taxa dominating benthic associations had a higher cadmium content than related species from other seas. Very high concentrations of the metal were found in sponges (10–80 μg/g dw) and in the digestive gland of molluscs (up to 345 μg/g in Neobuccinum eatoni). The rapid regeneration of cadmium and its natural occurrence and bioavailability in highly productive coastal waters seem to be responsible for cadmium accumulation in the tissues of marine organisms near the “Baia Terra Nova” station. Received: 6 June 1995/Accepted: 23 November 1995     

Individual differences in aggressiveness and food stealing among wintering oystercatchers, Haematopus ostralegus L

Oystercatchers feeding on mussels foraged in small, overlapping areas and aggressive encounters were common amongst all birds irrespective of their method of opening mussels. Individuals varied consistently in aggressiveness. The more aggressive birds were more frequently present on the mussel bed, fed in those parts where food was most abundant, and were more successful in stealing mussels from other birds. Only the more aggressive birds stole more mussels than they had stolen from them, and stealing increased their rate of food intake.     

Identification of an intermolecular disulfide bond in barley hemoglobin

The present study was designed to investigate properties of ion channels in undifferentiated rabbit mesenchymal stem cells (MSCs) from bone marrow using whole-cell patch-clamp and RT-PCR techniques. It was found that three types of outward currents were present in rabbit MSCs, including an inward rectifier K(+) current (I(Kir)), a noise-like Ca(2+)-activated K(+) current (I(KCa)) co-present with delayed rectifier K(+) current (IK(DR)). I(Kir) was inhibited by Ba(2+), while I(KCa) was inhibited by paxilline (a blocker of big conductance I(KCa) channels) and clotrimazole (an inhibitor of intermediate conductance I(KCa) channels). IK(DR) exhibited a slow inactivation, "U-shaped" voltage-dependent inactivation, and slow recovery from inactivation, and the current was inhibited by tetraethylammonium or 4-aminopyridine. RT-PCR revealed the molecular identities for the functional ionic currents, including Kir1.1 (possibly responsible for I(Kir)), KCa1.1 and KCa3.1 (possibly responsible for I(KCa)), and Kv1.2, Kv2.1, and Kv2.2 (possibly responsible for IK(DR)). These results demonstrate for the first time that three types of functional ion channel currents (i.e., I(Kir), I(KCa), and IK(DR)) are present in rabbit MSCs from bone marrow.     

The Role of the ��DELSEED-loop of ATP Synthase

ATP synthase uses a unique rotational mechanism to convert chemical energy into mechanical energy and back into chemical energy. The helix-turn-helix motif, termed “DELSEED-loop,” in the C-terminal domain of the β subunit was suggested to be involved in coupling between catalysis and rotation. Here, the role of the DELSEED-loop was investigated by functional analysis of mutants of Bacillus PS3 ATP synthase that had 3–7 amino acids within the loop deleted. All mutants were able to catalyze ATP hydrolysis, some at rates several times higher than the wild-type enzyme. In most cases ATP hydrolysis in membrane vesicles generated a transmembrane proton gradient, indicating that hydrolysis occurred via the normal rotational mechanism. Except for two mutants that showed low activity and low abundance in the membrane preparations, the deletion mutants were able to catalyze ATP synthesis. In general, the mutants seemed less well coupled than the wild-type enzyme, to a varying degree. Arrhenius analysis demonstrated that in the mutants fewer bonds had to be rearranged during the rate-limiting catalytic step; the extent of this effect was dependent on the size of the deletion. The results support the idea of a significant involvement of the DELSEED-loop in mechanochemical coupling in ATP synthase. In addition, for two deletion mutants it was possible to prepare an α₃β₃γ subcomplex and measure nucleotide binding to the catalytic sites. Interestingly, both mutants showed a severely reduced affinity for MgATP at the high affinity site.F₁F₀-ATP synthase catalyzes the final step of oxidative phosphorylation and photophosphorylation, the synthesis of ATP from ADP and inorganic phosphate. F₁F₀-ATP synthase consists of the membrane-embedded F₀ subcomplex, with, in most bacteria, a subunit composition of ab₂c₁₀, and the peripheral F₁ subcomplex, with a subunit composition of α₃β₃γδε. The energy necessary for ATP synthesis is derived from an electrochemical transmembrane proton (or, in some organisms, a sodium ion) gradient. Proton flow down the gradient through F₀ is coupled to ATP synthesis on F₁ by a unique rotary mechanism. The protons flow through (half) channels at the interface of the a and c subunits, which drives rotation of the ring of c subunits. The c₁₀ ring, together with F₁ subunits γ and ε, forms the rotor. Rotation of γ leads to conformational changes in the catalytic nucleotide binding sites on the β subunits, where ADP and P_i are bound. The conformational changes result in the formation and release of ATP. Thus, ATP synthase converts electrochemical energy, the proton gradient, into mechanical energy in the form of subunit rotation and back into chemical energy as ATP. In bacteria, under certain physiological conditions, the process runs in reverse. ATP is hydrolyzed to generate a transmembrane proton gradient, which the bacterium requires for such functions as nutrient import and locomotion (for reviews, see Refs. 1–6).F₁ (or F₁-ATPase) has three catalytic nucleotide binding sites located on the β subunits at the interface to the adjacent α subunit. The catalytic sites have pronounced differences in their nucleotide binding affinity. During rotational catalysis, the sites switch their affinities in a synchronized manner; the position of γ determines which catalytic site is the high affinity site (K_d1 in the nanomolar range), which site is the medium affinity site (K_d2 ≈ 1 μm), and which site is the low affinity site (K_d3 ≈ 30–100 μm; see Refs. 7 and 8). In the original crystal structure of bovine mitochondrial F₁ (9), one of the three catalytic sites, was filled with the ATP analog AMP-PNP,² a second was filled with ADP (plus azide) (see Ref. 10), and the third site was empty. Hence, the β subunits are referred to as β_TP, β_DP, and β_E. The occupied β subunits, β_TP and β_DP, were in a closed conformation, and the empty β_E subunit was in an open conformation. The main difference between these two conformations is found in the C-terminal domain. Here, the “DELSEED-loop,” a helix-turn-helix structure containing the conserved DELSEED motif, is in an “up” position when the catalytic site on the respective β subunit is filled with nucleotide and in a “down” position when the site is empty (Fig. 1A). When all three catalytic sites are occupied by nucleotide, the previously open β_E subunit assumes an intermediate, half-closed (β_HC) conformation. It cannot close completely because of steric clashes with γ (11).Open in a separate windowFIGURE 1.The βDELSEED-loop. A, interaction of the β_TP and β_E subunits with theγ subunit.β subunits are shown in yellow andγ in blue. The DELSEED-loop (shown in orange, with the DELSEED motif itself in green)of β_TP interacts with the C-terminal helixγ and the short helix that runs nearly perpendicular to the rotation axis. The DELSEED-loop of β_E makes contact with the convex portion of γ, formed mainly by the N-terminal helix. A nucleotide molecule (shown in stick representation) occupies the catalytic site of β_TP, and the subunit is in the closed conformation. The catalytic site on β_E is empty, and the subunit is in the open conformation. This figure is based on Protein Data Bank file 1e79 (32). B, deletions in the βDELSEED-loop. The loop was “mutated” in silico to represent the PS3 ATP synthase. The 3–4-residue segments that are removed in the deletion mutants are color-coded as follows: ³⁸⁰LQDI³⁸³, pink; ³⁸⁴IAIL³⁸⁷, green; ³⁸⁸GMDE³⁹¹, yellow; ³⁹²LSD³⁹⁴, cyan; ³⁹⁵EDKL³⁹⁸, orange; ³⁹⁹VVHR⁴⁰², blue. Residues that are the most involved in contacts with γ are labeled. All figures were generated using the program PyMOL (DeLano Scientific, San Carlos, CA).The DELSEED-loop of each of the three β subunits makes contact with the γ subunit. In some cases, these contacts consist of hydrogen bonds or salt bridges between the negatively charged residues of the DELSEED motif and positively charged residues on γ. The interactions of the DELSEED-loop with γ, its movement during catalysis, the conservation of the DELSEED motif (see 12–14). Thus, the finding that an AALSAAA mutant in the α₃β₃γ complex of ATP synthase from the thermophilic Bacillus PS3, where several hydrogen bonds/salt bridges to γ are removed simultaneously, could drive rotation of γ with the same torque as the wild-type enzyme (14) came as a surprise. On the other hand, it seems possible that it is the bulk of the DELSEED-loop, more so than individual interactions, that drives rotation of γ. According to a model favored by several authors (6, 15, 16) (see also Refs. 17–19), binding of ATP (or, more precisely, MgATP) to the low affinity catalytic site on β_E and the subsequent closure of this site, accompanied by its conversion into the high affinity site, are responsible for driving the large (80–90°) rotation substep during ATP hydrolysis, with the DELSEED-loop acting as a “pushrod.” A recent molecular dynamics (20) study supports this model and implicates mainly the region around several hydrophobic residues upstream of the DELSEED motif (specifically βI386 and βL387)³ as being responsible for making contact with γ during the large rotation substep.

TABLE 1

Conservation of residues in the DELSEED-loop Amino acids found in selected species in the turn region of the DELSEED-loop. Listed are all positions subjected to deletions in the present study. Residue numbers refer to the PS3 enzyme. Consensus annotation: p, polar residue; s, small residue; h, hydrophobic residue; –, negatively charged residue; +, positively charged residue.Open in a separate windowIn the present study, we investigated the function of the DELSEED-loop using an approach less focused on individual residues, by deleting stretches of 3–7 amino acids between positions β380 and β402 of ATP synthase from the thermophilic Bacillus PS3. We analyzed the functional properties of the deletion mutants after expression in Escherichia coli. The mutants showed ATPase activities, which were in some cases surprisingly high, severalfold higher than the activity of the wild-type control. On the other hand, in all cases where ATP synthesis could be measured, the rates where below or equal to those of the wild-type enzyme. In Arrhenius plots, the hydrolysis rates of the mutants were less temperature-dependent than those of wild-type ATP synthase. In those cases where nucleotide binding to the catalytic sites could be tested, the deletion mutants had a much reduced affinity for MgATP at high affinity site 1. The functional role of the DELSEED-loop will be discussed in light of the new information.     

FLUCTUATING SELECTION AND THE MAINTENANCE OF INDIVIDUAL AND SEX‐SPECIFIC DIET SPECIALIZATION IN FREE‐LIVING OYSTERCATCHERS

Fluctuating and disruptive selection are important mechanisms for maintaining intrapopulation trait variation. Nonetheless, few field studies quantify selection pressures over long periods and identify what causes them to fluctuate. Diet specialists in oystercatchers differ in short‐term payoffs (intake), but their long‐term payoffs are hypothesized to be condition dependent. We test whether phenotypic selection on diet specialization fluctuates between years due to the frequency of specialists, competitor density, prey abundance, and environmental conditions. Short‐term payoffs proved to be poor predictors of long‐term fitness payoffs of specialization. Sex‐differences in diet specialization were maintained by opposing directional fecundity and viability selection between the sexes. Contrasting other studies, selection on individual diet specialization was neither negative frequency‐ or density‐dependent nor dependent on prey abundance. Notwithstanding, viability selection fluctuated strongly (stabilizing?disruptive) over the 26‐year study period: slightly favoring generalists in most years, but strongly disfavoring generalists in rare harsh winters, suggesting generalists cannot cope with extreme conditions. Although selection fluctuated, mean selection on specialists was weak, which can explain how individual specialization can persist over long periods. Because rare events can dramatically affect long‐term selective landscapes, more care should be taken to match the timescale of evolutionary studies to the temporal variability of critical environmental conditions.     

Comparative distribution of human and avian type sialic acid influenza receptors in the pig

Background  

A major determinant of influenza infection is the presence of virus receptors on susceptible host cells to which the viral haemagglutinin is able to bind. Avian viruses preferentially bind to sialic acid α2,3-galactose (SAα2,3-Gal) linked receptors, whereas human strains bind to sialic acid α2,6-galactose (SAα2,6-Gal) linked receptors. To date, there has been no detailed account published on the distribution of SA receptors in the pig, a model host that is susceptible to avian and human influenza subtypes, thus with potential for virus reassortment. We examined the relative expression and spatial distribution of SAα2,3-GalG(1-3)GalNAc and SAα2,6-Gal receptors in the major organs from normal post-weaned pigs by binding with lectins Maackia amurensis agglutinins (MAA II) and Sambucus nigra agglutinin (SNA) respectively.     

Analysis of the wheat and Puccinia triticina (leaf rust) proteomes during a susceptible host-pathogen interaction

Wheat leaf rust is caused by the fungus Puccinia triticina. The genetics of resistance follows the gene-for-gene hypothesis, and thus the presence or absence of a single host resistance gene renders a plant resistant or susceptible to a leaf rust race bearing the corresponding avirulence gene. To investigate some of the changes in the proteomes of both host and pathogen during disease development, a susceptible line of wheat infected with a virulent race of leaf rust were compared to mock-inoculated wheat using 2-DE (with IEF pH 4-8) and MS. Up-regulated protein spots were excised and analyzed by MALDI-QqTOF MS/MS, followed by cross-species protein identification. Where possible MS/MS spectra were matched to homologous proteins in the NCBI database or to fungal ESTs encoding putative proteins. Searching was done using the MASCOT search engine. Remaining unmatched spectra were then sequenced de novo and queried against the NCBInr database using the BLAST and MS BLAST tools. A total of 32 consistently up-regulated proteins were examined from the gels representing the 9-day post-infection proteome in susceptible plants. Of these 7 are host proteins, 22 are fungal proteins of known or hypothetical function and 3 are unknown proteins of putative fungal origin.     

Optimal foraging on perilous prey: risk of bill damage reduces optimal prey size in oystercatchers

Intake rate maximization alone is not always sufficient in explainingprey size selection in predators. For example, bivalve-feedingoystercatchers regularly select smaller prey than expected ifthey aimed to maximize their intake rate. It has been proposedthat to these birds large prey are "risky," in the sense thatbirds may damage their bills when feeding on large bivalves.Large bivalves yield more energy, but according to this hypothesisthis is achieved at the expense of energy yield in the longterm when (1) the risk of bill damage increases with prey sizeand (2) foraging with a damaged bill is less effective. In accordancewith this hypothesis, we show that captive oystercatchers feedingon large cockles experienced a high probability of bill tipdamage, while bill damage was absent when cockles were small.Moreover, among free-living oystercatchers the prevalence ofbill damage was correlated with mean cockle size near the capturesite, and the data on captive birds fit in this pattern. Foodintake of captive oystercatchers feeding exclusively on cockleswas reduced by 23% after bill damage, and free-living birdswith damaged bills had 14 g lower mass. Because lower body masswas associated with higher mortality probability, these resultsindicate long-term costs associated with feeding on large cockles.We conclude that the risk of bill damage can potentially explainwhy oystercatchers avoid large bivalves and that oystercatchersmay maximize long-term intake rate by selecting prey sizes thatare "suboptimal" from a short-term rate-maximizing point ofview.