Haplotype and nucleotide characteristics of photobiont and mycobiont gene sequences in the lichen Cetraria aculeata

Lichens, symbiotic associations of fungi (mycobionts) and green algae or cyanobacteria (photobionts), are poikilohydric organisms that are particularly well adapted to withstand adverse environmental conditions. Terrestrial ecosystems of the Antarctic are therefore largely dominated by lichens. The effects of global climate change are especially pronounced in the maritime Antarctic and it may be assumed that the lichen vegetation will profoundly change in the future. The genetic diversity of populations is closely correlated to their ability to adapt to changing environmental conditions and to their future evolutionary potential. In this study, we present evidence for low genetic diversity in Antarctic mycobiont and photobiont populations of the widespread lichen Cetraria aculeata. We compared between 110 and 219 DNA sequences from each of three gene loci for each symbiont. A total of 222 individuals from three Antarctic and nine antiboreal, temperate and Arctic populations were investigated. The mycobiont diversity is highest in Arctic populations, while the photobionts are most diverse in temperate regions. Photobiont diversity decreases significantly towards the Antarctic but less markedly towards the Arctic, indicating that ecological factors play a minor role in determining the diversity of Antarctic photobiont populations. Richness estimators calculated for the four geographical regions suggest that the low genetic diversity of Antarctic populations is not a sampling artefact. Cetraria aculeata appears to have diversified in the Arctic and subsequently expanded its range into the Southern Hemisphere. The reduced genetic diversity in the Antarctic is most likely due to founder effects during long-distance colonization.

Nicotinamide nucleotide and adenylate concentrations in mante, siphon and gill tissue of old and young Laternula elliptica individuals under control and experimental conditions

Future oceans are predicted to contain less oxygen than at present. This is because oxygen is less soluble in warmer water and predicted stratification will reduce mixing. Hypoxia in marine environments is thus likely to become more widespread in marine environments and understanding species-responses is important to predicting future impacts on biodiversity. This study used a tractable model, the Antarctic clam, Laternula elliptica, which can live for 36 years, and has a well-characterized ecology and physiology to understand responses to hypoxia and how the effect varied with age. Younger animals had a higher condition index, higher adenylate energy charge and transcriptional profiling indicated that they were physically active in their response to hypoxia, whereas older animals were more sedentary, with higher levels of oxidative damage and apoptosis in the gills. These effects could be attributed, in part, to age-related tissue scaling; older animals had proportionally less contractile muscle mass and smaller gills and foot compared with younger animals, with consequential effects on the whole-animal physiological response. The data here emphasize the importance of including age effects, as large mature individuals appear to be less able to resist hypoxic conditions and this is the size range that is the major contributor to future generations. Thus, the increased prevalence of hypoxia in future oceans may have marked effects on benthic organisms' abilities to persist and this is especially so for long-lived species when predicting responses to environmental perturbation.

Environmental analyses of cyclic Middle Eocene sediments and palynomorph records of ODP Hole 189-1172A

The influence of orbital precession on early Paleogene climate and ocean circulation patterns in the southeast Pacific region is investigated by combining environmental analyses of cyclic Middle Eocene sediments and palynomorph records recovered from ODP Hole 1172A on the East Tasman Plateau with climate model simulations. Integration of results indicates that in the marine realm, direct effects of precessional forcing are not pronounced, although increased precipitation/runoff could have enhanced dinoflagellate cyst production. On the southeast Australian continent, the most pronounced effects of precessional forcing were fluctuations in summer precipitation and temperature on the Antarctic Margin. These fluctuations resulted in vegetational changes, most notably in the distribution of Nothofagus (subgenus Brassospora). The climate model results suggest significant fluctuations in sea ice in the Ross Sea, notably during Austral summers. This is consistent with the influx of Antarctic heterotrophic dinoflagellates in the early part of the studied record. The data demonstrate a strong precessionally driven climate variability and thus support the concept that precessional forcing could have played a role in early Antarctic glaciation via changes in runoff and/or precipitation.

Cyclic behaviour of saturated sands subject to previous horizontal shear stresses

The dynamic behaviour of saturated sands has been studied from different perspectives. However, most experimental research on this field does not take into account the shear stress conditions existing prior to the application of dynamic loads; i.e., a null initial static shear stress (τo = 0) is assumed. The main objective of this work is to report on the influence that static shear stresses (τo) have on the behaviour of saturated sands under cyclic shear loads. This article presents the results and analysis of part of a wider experimental programme involving 30 monotonic and 26 cyclic simple shear tests for different combinations of static shear stress (τo) and cyclic shear stress (τc) (all undrained), besides identification and classification tests. The tested samples have been taken from the area of the North Entrance Mouth at the Port of Barcelona (Spain).

Dynamic & Cyclic behaviour of ballast in the long term as determined in Cedex's track box

The 6 cylinder servo-hydraulic loading system of CEDEX's track box (250 kN, 50 Hz) has been recently implemented with a new piezoelectric loading system (±20 kN, 300 Hz) allowing the incorporation of low amplitude high frequency dynamic load time histories to the high amplitude low frequency quasi-static load time histories used so far in the CEDEX's track box to assess the inelastic long term behavior of ballast under mixed traffic in conventional and high- speed lines. This presentation will discuss the results obtained in the first long-duration test performed at CEDEX's track box using simultaneously both loading systems, to simulate the pass-by of 6000 freight vehicles (1M of 225 kN axle loads) travelling at a speed of 120 km/h over a line with vertical irregularities corresponding to a medium quality lin3e level. The superstructure of the track tested at full scale consisted of E 60 rails, stiff rail pads (mayor que 450 kN/mm), B90.2 sleepers with USP 0.10 N/mm and a 0.35 m thick ballast layer of ADIF first class. A shear wave velocity of 250 m/s can be assumed for the different layers of the track sub-base. The ballast long-term settlements will be compared with those obtained in a previous long-duration quasi- static test performed in the same track, for the RIVAS [EU co-funded] project, in which no dynamic loads where considered. Also, the results provided by a high diameter cyclic triaxial cell with ballast tested in full size will be commented. Finally, the progress made at CEDEX's Geotechnical Laboratory to reproduce numerically the long term behavior of ballast will be discussed.

Health monitoring of web plastifying dampers subjected to cyclic loading through vibration tests

This article investigates experimentally the application of health monitoring techniques to assess the damage on a particular kind of hysteretic (metallic) damper called web plastifying dampers, which are subjected to cyclic loading. In general terms, hysteretic dampers are increasingly used as passive control systems in advanced earthquake-resistant structures. Nonparametric statistical processing of the signals obtained from simple vibration tests of the web plastifying damper is used here to propose an area index damage. This area index damage is compared with an alternative energy-based index of damage proposed in past research that is based on the decomposition of the load?displacement curve experienced by the damper. Index of damage has been proven to accurately predict the level of damage and the proximity to failure of web plastifying damper, but obtaining the load?displacement curve for its direct calculation requires the use of costly instrumentation. For this reason, the aim of this study is to estimate index of damage indirectly from simple vibration tests, calling for much simpler and cheaper instrumentation, through an auxiliary index called area index damage. Web plastifying damper is a particular type of hysteretic damper that uses the out-of-plane plastic deformation of the web of I-section steel segments as a source of energy dissipation. Four I-section steel segments with similar geometry were subjected to the same pattern of cyclic loading, and the damage was evaluated with the index of damage and area index damage indexes at several stages of the loading process. A good correlation was found between area index damage and index of damage. Based on this correlation, simple formulae are proposed to estimate index of damage from the area index damage.

Evidence of a role for cyclic ADP-ribose in long-term synaptic depression in hippocampus

Ca2+ released from presynaptic and postsynaptic intracellular stores plays important roles in activity-dependent synaptic plasticity, including long-term depression (LTD) of synaptic strength. At Schaffer collateral–CA1 synapses in the hippocampus, presynaptic ryanodine receptor-gated stores appear to mobilize some of the Ca2+ necessary to induce LTD. Cyclic ADP-ribose (cADPR) has recently been proposed as an endogenous activator of ryanodine receptors in sea urchin eggs and several mammalian cell types. Here, we provide evidence that cADPR-mediated signaling pathways play a key role in inducing LTD. We show that biochemical production of cGMP increases cADPR concentration in hippocampal slices in vitro, and that blockade of cGMP-dependent protein kinase, cADPR receptors, or ryanodine-sensitive Ca2+ stores each prevent the induction of LTD at Schaffer collateral–CA1 synapses. A lack of effect of postsynaptic infusion of either cADPR antagonist indicates a probable presynaptic site of action.

Identification of a nucleotide binding site in HIV-1 integrase

HIV-1 integrase is essential for viral replication and can be inhibited by antiviral nucleotides. Photoaffinity labeling with the 3′-azido-3′-deoxythymidine (AZT) analog 3′,5-diazido-2′,3′-dideoxyuridine 5′-monophosphate (5N3-AZTMP) and proteolytic mapping identified the amino acid 153–167 region of integrase as the site of photocrosslinking. Docking of 5N3-AZTMP revealed the possibility for strong hydrogen bonds between the inhibitor and lysines 156, 159, and 160 of the enzyme. Mutation of these residues reduced photocrosslinking selectively. This report elucidates the binding site of a nucleotide inhibitor of HIV-1 integrase, and possibly a component of the enzyme polynucleotide binding site.

Brefeldin A-inhibited guanine nucleotide-exchange activity of Sec7 domain from yeast Sec7 with yeast and mammalian ADP ribosylation factors

The Saccharomyces cerevisiae Sec7 protein (ySec7p), which is an important component of the yeast secretory pathway, contains a sequence of ≈200 amino acids referred to as a Sec7 domain. Similar Sec7 domain sequences have been recognized in several guanine nucleotide-exchange proteins (GEPs) for ADP ribosylation factors (ARFs). ARFs are ≈20-kDa GTPases that regulate intracellular vesicular membrane trafficking and activate phospholipase D. GEPs activate ARFs by catalyzing the replacement of bound GDP with GTP. We, therefore, undertook to determine whether a Sec7 domain itself could catalyze nucleotide exchange on ARF and found that it exhibited brefeldin A (BFA)-inhibitable ARF GEP activity. BFA is known to inhibit ARF GEP activity in Golgi membranes, thereby causing reversible apparent dissolution of the Golgi complex in many cells. The His6-tagged Sec7 domain from ySec7p (rySec7d) synthesized in Escherichia coli enhanced binding of guanosine 5′-[γ-[35S]thio]triphosphate by recombinant yeast ARF1 (ryARF1) and ryARF2 but not by ryARF3. The effects of rySec7d on ryARF2 were inhibited by BFA in a concentration-dependent manner but not by inactive analogues of BFA (B-17, B-27, and B-36). rySec7d also promoted BFA-sensitive guanosine 5′-[γ-thio]triphosphate binding by nonmyristoylated recombinant human ARF1 (rhARF1), rhARF5, and rhARF6, although the effect on rhARF6 was very small. These results are consistent with the conclusion that the yeast Sec7 domain itself contains the elements necessary for ARF GEP activity and its inhibition by BFA.

Cyclic AMP regulates potassium channel expression in C6 glioma by destabilizing Kv1.1 mRNA

The tissue distributions and physiological properties of a variety of cloned voltage-gated potassium channel genes have been characterized extensively, yet relatively little is known about the mechanisms controlling expression of these genes. Here, we report studies on the regulation of Kv1.1 expressed endogenously in the C6 glioma cell line. We demonstrate that elevation of intracellular cAMP leads to the accelerated degradation of Kv1.1 RNA. The cAMP-induced decrease in Kv1.1 RNA is followed by a decrease in Kv1.1 protein and a decrease in the whole cell sustained K+ current amplitude. Dendrotoxin-I, a relatively specific blocker of Kv1.1, blocks 96% of the sustained K+ current in glioma cells, causing a shift in the resting membrane potential from −40 mV to −7 mV. These data suggest that expression of Kv1.1 contributes to setting the resting membrane potential in undifferentiated glioma cells. We therefore suggest that receptor-mediated elevation of cAMP reduces outward K+ current density by acting at the translational level to destabilize Kv1.1 RNA, an additional mechanism for regulating potassium channel gene expression.

In vitro repair of oxidative DNA damage by human nucleotide excision repair system: Possible explanation for neurodegeneration in Xeroderma pigmentosum patients

Xeroderma pigmentosum (XP) patients fail to remove pyrimidine dimers caused by sunlight and, as a consequence, develop multiple cancers in areas exposed to light. The second most common sign, present in 20–30% of XP patients, is a set of neurological abnormalities caused by neuronal death in the central and peripheral nervous systems. Neural tissue is shielded from sunlight-induced DNA damage, so the cause of neurodegeneration in XP patients remains unexplained. In this study, we show that two major oxidative DNA lesions, 8-oxoguanine and thymine glycol, are excised from DNA in vitro by the same enzyme system responsible for removing pyrimidine dimers and other bulky DNA adducts. Our results suggest that XP neurological disease may be caused by defective repair of lesions that are produced in nerve cells by reactive oxygen species generated as by-products of an active oxidative metabolism.

The signal recognition particle receptor of Escherichia coli (FtsY) has a nucleotide exchange factor built into the GTPase domain

Targeting of many secretory and membrane proteins to the inner membrane in Escherichia coli is achieved by the signal recognition particle (SRP) and its receptor (FtsY). In E. coli SRP consists of only one polypeptide (Ffh), and a 4.5S RNA. Ffh and FtsY each contain a conserved GTPase domain (G domain) with an α-helical domain on its N terminus (N domain). The nucleotide binding kinetics of the NG domain of the SRP receptor FtsY have been investigated, using different fluorescence techniques. Methods to describe the reaction kinetically are presented. The kinetics of interaction of FtsY with guanine nucleotides are quantitatively different from those of other GTPases. The intrinsic guanine nucleotide dissociation rates of FtsY are about 105 times higher than in Ras, but similar to those seen in GTPases in the presence of an exchange factor. Therefore, the data presented here show that the NG domain of FtsY resembles a GTPase–nucleotide exchange factor complex not only in its structure but also kinetically. The I-box, an insertion present in all SRP-type GTPases, is likely to act as an intrinsic exchange factor. From this we conclude that the details of the GTPase cycle of FtsY and presumably other SRP-type GTPases are fundamentally different from those of other GTPases.

Nucleotide-dependent conformational change near the fulcrum region in Dictyostelium myosin II

In skeletal muscle myosin, the reactive thiols (SH1 and SH2) are close to a proposed fulcrum region that is thought to undergo a large conformational change. The reactive thiol region is thought to transmit the conformational changes induced by the actin–myosin–ATP interactions to the lever arm, which amplifies the power stroke. In skeletal muscle myosin, SH1 and SH2 can be chemically cross-linked in the presence of nucleotide, trapping the nucleotide in its pocket. Although the flexibility of the reactive thiol region has been well studied in skeletal muscle myosin, crystal structures of truncated nonmuscle myosin II from Dictyostelium in the presence of various ATP analogs do not show changes at the reactive thiol region that would be consistent with the SH1–SH2 cross-linking observed for muscle myosin. To examine the dynamics of the reactive thiol region in Dictyostelium myosin II, we have examined a modified myosin II that has cysteines at the muscle myosin SH1 and SH2 positions. This myosin is specifically cross-linked at SH1–SH2 by a chemical cross-linker in the presence of ADP, but not in its absence. Furthermore, the cross-linked species traps the nucleotide, as in the case of muscle myosin. Thus, the Dictyostelium myosin II shares the same dynamic behavior in the fulcrum region of the molecule as the skeletal muscle myosin. This result emphasizes the importance of nucleotide-dependent changes in this part of the molecule.