A cysteine-rich CCG domain contains a novel [4Fe-4S] cluster binding motif as deduced from studies with subunit B of heterodisulfide reductase from Methanothermobacter marburgensis

Heterodisulfide reductase (HDR) of methanogenic archaea with its active-site [4Fe-4S] cluster catalyzes the reversible reduction of the heterodisulfide (CoM-S-S-CoB) of the methanogenic coenzyme M (CoM-SH) and coenzyme B (CoB-SH). CoM-HDR, a mechanistic-based paramagnetic intermediate generated upon half-reaction of the oxidized enzyme with CoM-SH, is a novel type of [4Fe-4S]3+ cluster with CoM-SH as a ligand. Subunit HdrB of the Methanothermobacter marburgensis HdrABC holoenzyme contains two cysteine-rich sequence motifs (CX31-39CCX35-36CXXC), designated as CCG domain in the Pfam database and conserved in many proteins. Here we present experimental evidence that the C-terminal CCG domain of HdrB binds this unusual [4Fe-4S] cluster. HdrB was produced in Escherichia coli, and an iron-sulfur cluster was subsequently inserted by in vitro reconstitution. In the oxidized state the cluster without the substrate exhibited a rhombic EPR signal (gzyx = 2.015, 1.995, and 1.950) reminiscent of the CoM-HDR signal. 57Fe ENDOR spectroscopy revealed that this paramagnetic species is a [4Fe-4S] cluster with 57Fe hyperfine couplings very similar to that of CoM-HDR. CoM-33SH resulted in a broadening of the EPR signal, and upon addition of CoM-SH the midpoint potential of the cluster was shifted to values observed for CoM-HDR, both indicating binding of CoM-SH to the cluster. Site-directed mutagenesis of all 12 cysteine residues in HdrB identified four cysteines of the C-terminal CCG domain as cluster ligands. Combined with the previous detection of CoM-HDR-like EPR signals in other CCG domain-containing proteins our data indicate a general role of the C-terminal CCG domain in coordination of this novel [4Fe-4S] cluster. In addition, Zn K-edge X-ray absorption spectroscopy identified an isolated Zn site with an S3(O/N)1 geometry in HdrB and the HDR holoenzyme. The N-terminal CCG domain is suggested to provide ligands to the Zn site.     

Food web dynamics affect Northeast Arctic cod recruitment

Proper management of ecosystems requires an understanding of both the species interactions as well as the effect of climate variation. However, a common problem is that the available time-series are of different lengths. Here, we present a general approach for studying the dynamic structure of such interactions. Specifically, we analyse the recruitment of the world's largest cod stock, the Northeast Arctic cod. Studies based on data starting in the 1970-1980s indicate that this stock is affected by temperature through a variety of pathways. However, the value of such studies is somewhat limited by the fact that they are based on a quite specific ecological and climatic situation. Recently, this stock has consisted of fairly young fish and the spawning stock has consisted of relatively few age groups. In this study, we develop a model for the effect of capelin (the cod's main prey) and herring on cod recruitment since 1973. Based on this model, we analyse data on cod, herring and temperature going back to 1921 and find that food-web effects explain a significant part of the cod recruitment variation back to around 1950.     

O2 reactions at the six-iron active site (H-cluster) in [FeFe]-hydrogenase

Irreversible inhibition by molecular oxygen (O(2)) complicates the use of [FeFe]-hydrogenases (HydA) for biotechnological hydrogen (H(2)) production. Modification by O(2) of the active site six-iron complex denoted as the H-cluster ([4Fe4S]-2Fe(H)) of HydA1 from the green alga Chlamydomonas reinhardtii was characterized by x-ray absorption spectroscopy at the iron K-edge. In a time-resolved approach, HydA1 protein samples were prepared after increasing O(2) exposure periods at 0 °C. A kinetic analysis of changes in their x-ray absorption near edge structure and extended X-ray absorption fine structure spectra revealed three phases of O(2) reactions. The first phase (τ(1) ≤ 4 s) is characterized by the formation of an increased number of Fe-O,C bonds, elongation of the Fe-Fe distance in the binuclear unit (2Fe(H)), and oxidation of one iron ion. The second phase (τ(2) ≈ 15 s) causes a ～50% decrease of the number of ～2.7-? Fe-Fe distances in the [4Fe4S] subcluster and the oxidation of one more iron ion. The final phase (τ(3) ≤ 1000 s) leads to the disappearance of most Fe-Fe and Fe-S interactions and further iron oxidation. These results favor a reaction sequence, which involves 1) oxygenation at 2Fe(H(+)) leading to the formation of a reactive oxygen species-like superoxide (O(2)(-)), followed by 2) H-cluster inactivation and destabilization due to ROS attack on the [4Fe4S] cluster to convert it into an apparent [3Fe4S](+) unit, leading to 3) complete O(2)-induced degradation of the remainders of the H-cluster. This mechanism suggests that blocking of ROS diffusion paths and/or altering the redox potential of the [4Fe4S] cubane by genetic engineering may yield improved O(2) tolerance in [FeFe]-hydrogenase.     

Phagocytosis and Respiratory Burst Activity in Lumpsucker (Cyclopterus lumpus L.) Leucocytes Analysed by Flow Cytometry

In the present study, we have isolated leucocytes from peripheral blood, head kidney and spleen from lumpsucker (Cyclopterus lumpus L.), and performed functional studies like phagocytosis and respiratory burst, as well as morphological and cytochemical analyses. Different leucocytes were identified, such as lymphocytes, monocytes/macrophages and polymorphonuclear cells with bean shaped or bilobed nuclei. In addition, cells with similar morphology as described for dendritic cells in trout were abundant among the isolated leucocytes. Flow cytometry was successfully used for measuring phagocytosis and respiratory burst activity. The phagocytic capacity and ability were very high, and cells with different morphology in all three leucocyte preparations phagocytised beads rapidly. Due to lack of available cell markers, the identity of the phagocytic cells could not be determined. The potent non-specific phagocytosis was in accordance with a high number of cells positive for myeloperoxidase, an enzyme involved in oxygen-dependent killing mechanism present in phagocytic cells. Further, high respiratory burst activity was present in the leucocytes samples, verifying a potent oxygen- dependent degradation. At present, the specific antibody immune response could not be measured, as immunoglobulin or B-cells have not yet been isolated. Therefore, analyses of the specific immune response in this fish species await further clarification. The present study presents the first analyses of lumpsucker immunity and also the first within the order Scopaeniformes.     

Effects of group size and group density on trade-offs in resource selection by a group-territorial central-place foraging woodpecker

Trade-offs in resource selection by central-place foragers are driven by the need to balance the benefits of selecting resources against the costs of travel from the central place. For group-territorial central-place foraging birds, trade-offs in resource selection are likely to be complicated by a competitive advantage for larger groups at high group density that may limit accessibility of high-quality distant resources to small groups. We used the group-territorial, central-place foraging Red-cockaded Woodpecker Leuconotopicus borealis (RCW) as a case study to test predictions that increases in group density lead to differences in foraging distances and resource selection for groups of different sizes. We used GPS tracking and LiDAR-derived habitat data to model effects of group size on foraging distances and selection for high-quality pines (≥ 35.6 cm diameter at breast height (dbh)) and lower quality pines (25.4–35.6 cm dbh) by RCW groups across low (n = 14), moderate (n = 10) and high group density (n = 10) conditions. At low and moderate group density, all RCW groups selected distant high-quality pines in addition to those near the central place because competition for resources was low. In contrast, at high group density, larger groups travelled further to select high-quality pines, whereas smaller groups selected high-quality pines only when they were close to the central place and, conversely, were more likely to select lower quality pines at greater distances from the central place. Selection for high-quality pines only when close to the cavity tree cluster at high group density is important to long-term fitness of small RCW groups because it allows them to maximize benefits from both territorial defence and selecting high-quality resources while minimizing costs of competition. These relationships suggest that intraspecific competition at high group density entails substantive costs to smaller groups of territorial central-place foragers by limiting accessibility of distant high-quality foraging resources.     

Cholinesterase Activities in Uterus of Normal and Fenchlorphos Treated Blue Foxes (Alopex Lagopus) during Various Reproductive States

The uterine acetylcholinesterase and total cholinesterase (acetylcholinesterase plus butyrylcholinesterase) activities in normal and fenchlorphos treated blue fox vixens were determined during various reproductive states. AChE and Total-ChE of non-medicated vixens in oestrus were about one half of those in anoestrus. In pregnant uteri (luteal phase) the activities were 25 % and 30% compared to anoestrus. In vixens given 100 mg/kg fenchlorphos for 3 weeks during anoestrus, the remaining activity of AChE in uterus were in average 37%. Pregnant and non-pregnant vixens in the luteal phase medicated prior to mating and during time of implantation, displayed AChE activities which were only moderabely reduced (remaining activities 83% and 72% compared to medicated animals in anoestrus: remaining activity 37%). Plasma ChE-activity increased during pregnancy in the controls while enzyme activity was strongly reduced in animals given 100 mg/kg fenchlorphos daily through the whole pregnancy. It was concluded that the previous reported embryotoxic effect of fenchlorphos in the blue fox did not seem to be directed towards the moderate inhibition of the uterine cholinesterases.     

DNA replication and H5 histone exchange during reactivation of chick erythrocyte nuclei in heterokaryons

Fusion of terminally differentiated chick erythrocytes (CE) with replicating quail myoblasts or established L6J1 rat myoblasts results in reactivation of DNA synthesis in the dormant CE nuclei and in suppression of DNA synthesis in the myoblast nuclei. The nuclei of primary quail myoblasts are more effectively inhibited than the nuclei of established rat myoblasts. Inhibition of DNA replication occurs not only by preventing G1 nuclei from entering S-phase but also by blocking nuclei in S-phase and by delaying nuclei in G2 from undergoing mitosis and starting a new DNA replication cycle. No inhibition of DNA synthesis could be observed when mouse erythrocytes, i.e., erythrocytes lacking nuclei, were fused with rat myoblasts to generate mouse-globin-containing L6J1 cybrids. — Reactivation of CE nuclei is associated with a loss of the tissuespecific H5 histone variant. Complete elimination of H5 histone, however, does not seem to be a necessary prerequisite for the initiation or completion of DNA replication in CE nuclei since H5 antigens are found on reactivated G1, S, and G2 nuclei.     

The size at reproduction of canopy tree species in central Africa

Size at reproduction is a key aspect of species life history that is relatively understudied for long‐lived tropical trees. Here, we quantified reproductive diameter for 31 major timber species across 11 sites in Cameroon, Congo, and Central African Republic. Specifically, we examined whether (1) between‐species variability is correlated with other species traits; (2) reproductive diameter varies within‐species among sites; (3) reproductive status varies with crown exposure; and (4) the minimum cutting diameter limits (MCDL) imposed by national forest regulations enable seed trees to persist after logging operations. Consistent with studies conducted elsewhere in the tropics, we found great variability in diameter at reproduction among species, which correlated with adult stature (maximum diameter and height). For some species, reproductive diameter thresholds substantially varied between sites, and crown exposure had a significant positive effect on reproductive status. Most MCDLs were found to be suitable, with trees having a high probability of being seed trees at MCDL. Our findings have implications for the sustainable management of production forests, and they highlight questionable MCDLs for some species and between‐site variation in reproductive diameter. The study also highlights the need for long‐term phenological monitoring of tree species spanning a large range of ecological strategies to address both theoretical (species life history, allocation trade‐offs) and practical questions (MCDL).