Alterations of Red Cell Membrane Properties in Nneuroacanthocytosis

Neuroacanthocytosis (NA) refers to a group of heterogenous, rare genetic disorders, namely chorea acanthocytosis (ChAc), McLeod syndrome (MLS), Huntington’s disease-like 2 (HDL2) and pantothenate kinase associated neurodegeneration (PKAN), that mainly affect the basal ganglia and are associated with similar neurological symptoms. PKAN is also assigned to a group of rare neurodegenerative diseases, known as NBIA (neurodegeneration with brain iron accumulation), associated with iron accumulation in the basal ganglia and progressive movement disorder. Acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions, is frequently observed in ChAc and MLS patients but less prevalent in PKAN (about 10%) and HDL2 patients. The pathological factors that lead to the formation of the acanthocytic red blood cell shape are currently unknown. The aim of this study was to determine whether NA/NBIA acanthocytes differ in their functionality from normal erythrocytes. Several flow-cytometry-based assays were applied to test the physiological responses of the plasma membrane, namely drug-induced endocytosis, phosphatidylserine exposure and calcium uptake upon treatment with lysophosphatidic acid. ChAc red cell samples clearly showed a reduced response in drug-induced endovesiculation, lysophosphatidic acid-induced phosphatidylserine exposure, and calcium uptake. Impaired responses were also observed in acanthocyte-positive NBIA (PKAN) red cells but not in patient cells without shape abnormalities. These data suggest an “acanthocytic state” of the red cell where alterations in functional and interdependent membrane properties arise together with an acanthocytic cell shape. Further elucidation of the aberrant molecular mechanisms that cause this acanthocytic state may possibly help to evaluate the pathological pathways leading to neurodegeneration.     

Fitness Consequences of Northward Dispersal as Possible Adaptation to Climate Change,Using Experimental Translocation of a Migratory Passerine

Climate change leads to rapid, differential changes in phenology across trophic levels, often resulting in temporal mismatches between predators and their prey. If a species cannot easily adjust its timing, it can adapt by choosing a new breeding location with a later phenology of its prey. In this study, we experimentally investigated whether long-distance dispersal to northern breeding grounds with a later phenology could be a feasible process to restore the match between timing of breeding and peak food abundance and thus improve reproductive success. Here, we report the successful translocation of pied flycatchers (Ficedula hypoleuca) to natural breeding sites 560 km to the Northeast. We expected translocated birds to have a fitness advantage with respect to environmental phenology, but to potentially pay costs through the lack of other locally adapted traits. Translocated individuals started egg laying 11 days earlier than northern control birds, which were translocated only within the northern site. The number of fledglings produced was somewhat lower in translocated birds, compared to northern controls, and fledglings were in lower body condition. Translocated individuals were performing not significantly different to control birds that remained at the original southern site. The lack of advantage of the translocated individuals most likely resulted from the exceptionally cold spring in which the experiment was carried out. Our results, however, suggest that pied flycatchers can successfully introduce their early breeding phenotype after dispersing to more northern areas, and thus that adaptation through dispersal is a viable option for populations that get locally maladapted through climate change.     

Body adiposity index and other indexes of body composition in the SAPHIR study: Association with cardiovascular risk factors

Objective:

The accuracy of anthropometric surrogate markers such as the body adiposity index (BAI) and other common indexes like the body mass index (BMI), waist‐to‐hip ratio (WHR) and waist‐to‐height ratio (WHtR) to predict metabolic sequelae is essential for its use in clinical practice.

Design and Methods:

Thus, we evaluated the strength of BAI and other indexes to relate with anthropometric parameters, adipocytokines, blood lipids, parameters of glucose‐homeostasis and blood pressure in 1,770 patients from the Salzburg Atherosclerosis Prevention Program in Subjects at High Individual Risk (SAPHIR) study in a crosssectional design. Measurements were BAI, BMI, WHR, WHtR, abdominal subcutaneous and visceral adipose tissue (aSAT and VAT), total body adipose tissue mass, body weight, waist‐ and hip circumference (WC and HC), leptin, adiponectin, high‐density lipoprotein‐cholesterol (HDL‐C), low‐density lipoprotein‐cholesterol (LDL‐C), triglycerides (TG), fasting plasma glucose, fasting plasma insulin, the homeostasis model assessment of insulin resistance (HOMAIR), systolic and diastolic blood pressure.

Results and Conclusions:

BAI was significantly associated with leptin and HC. We conclude that BAI was the best calculator for leptin. BAI was inferior to BMI to predict anthropometric parameters other than HC, adiponectin, blood lipids, parameters of glucose homeostasis, and blood pressure in this cross‐sectional study.     

The Role of Glia in Addiction: Dopamine as a Modulator of Glial Responses in Addiction

Cellular and Molecular Neurobiology - Addiction is a chronic and potentially deadly disease considered a global health problem. Nevertheless, there is still no ideal treatment for its management....     

Drug Effect Unveils Inter-head Cooperativity and Strain-dependent ADP Release in Fast Skeletal Actomyosin

Amrinone is a bipyridine compound with characteristic effects on the force-velocity relationship of fast skeletal muscle, including a reduction in the maximum shortening velocity and increased maximum isometric force. Here we performed experiments to elucidate the molecular mechanisms for these effects, with the additional aim to gain insight into the molecular mechanisms underlying the force-velocity relationship. In vitro motility assays established that amrinone reduces the sliding velocity of heavy meromyosin-propelled actin filaments by 30% at different ionic strengths of the assay solution. Stopped-flow studies of myofibrils, heavy meromyosin and myosin subfragment 1, showed that the effects on sliding speed were not because of a reduced rate of ATP-induced actomyosin dissociation because the rate of this process was increased by amrinone. Moreover, optical tweezers studies could not detect any amrinone-induced changes in the working stroke length. In contrast, the ADP affinity of acto-heavy meromyosin was increased about 2-fold by 1 mm amrinone. Similar effects were not observed for acto-subfragment 1. Together with the other findings, this suggests that the amrinone-induced reduction in sliding velocity is attributed to inhibition of a strain-dependent ADP release step. Modeling results show that such an effect may account for the amrinone-induced changes of the force-velocity relationship. The data emphasize the importance of the rate of a strain-dependent ADP release step in influencing the maximum sliding velocity in fast skeletal muscle. The data also lead us to discuss the possible importance of cooperative interactions between the two myosin heads in muscle contraction.Muscle contraction, as well as several other aspects of cell motility, results from cyclic interactions between myosin II motors and actin filaments. These force-generating interactions are driven by the hydrolysis of ATP at the myosin active site as outlined in Scheme 1 (1–3). In the absence of actin, the P_i and ADP release steps (k₄ and k₅) are rate-limiting for the entire cycle at high (>12 °C) and low temperatures, respectively (4–6). In the presence of actin, the rate of P_i release increases significantly, and the overall cycle is accelerated more than 2 orders of magnitude. The sliding velocity of myosin-propelled motors is generally believed to be rate-limited by actomyosin dissociation (rate constant k′₅, k′₆, or k′₂ in Scheme 1) (7). Alternatively, some studies (8, 9) have suggested that the sliding velocity is determined by the fraction of myosin heads in the weak-binding states, AM⁴ ATP and AM ADP P_i. However, it is worth emphasizing that K_T is very low under physiological conditions (1, 3) with low population of these states. For the same reason, the rate of dissociation of the AM complex is governed by K′₁ and k′₂.Open in a separate windowSCHEME 1.Simplified kinetics scheme for MgATP turnover by myosin (lower row) and actomyosin (upper row). Inorganic phosphate is denoted by P_i; MgATP is denoted by ATP, and MgADP is denoted by ADP; myosin is denoted by M. The states AM*ADP and AM ADP correspond to myosin heads with their nucleotide binding pocket in a partially closed and open conformation, respectively (7, 52). Rate constants are indicated by lowercase letters (rightward transitions, k₂ − k₅ and k′₂ − k′₅, or leftward transitions, k₋₂ − k₋₅ and k′₋₂ − k′₋₅) and equilibrium constants by uppercase letters (K₁, K′₁, K_T, K₃, K′₃, K₆, k′₆, and K_DP). The equilibrium constants are association constants except for simple bimolecular reactions where they are defined as k_i/k_−i.For the study of contractile mechanisms in both muscle and other types of cells, drugs may be useful as pharmacological tools affecting different transitions or states in the force-generating cycle. Whereas the use of drugs as tools may be less specific than site-directed mutagenesis, it also has advantages. The motor protein function may be studied in vivo, with maintained ordering of the protein components, e.g. as in the muscle sarcomere, allowing more insight into the relationship between specific molecular events and contractile properties of muscle. A drug that has been used quite extensively in this context is butanedione monoxime. The usefulness of this drug is based on firm characterization of its effect on actomyosin function on the molecular level (3, 10–13). More recently other drugs, like N-benzyl-p-toluene sulfonamide (14, 15) and blebbistatin (16), have been found to affect myosin function, and their effects at the molecular level have also been elucidated in some detail (14, 15, 17, 18). Both these drugs appear to affect the actomyosin interaction in a similar way as butanedione monoxime by inhibiting a step before (or very early in) the myosin power stroke, leading to the inhibition of actomyosin cross-bridge formation and force production.In contrast to the reduced isometric force, caused by the above mentioned drugs, the bipyridine compound amrinone (Fig. 1A) has been found to increase the isometric force production of fast intact skeletal muscles of the frog (19, 20) and mouse (21) and also of fast (but much less slow) skinned muscle fibers of the rat (22). In all the fast myosin preparations, the effect of about 1 mm amrinone on isometric force was associated with characteristic changes of the force-velocity relationship (Fig. 1B), including a reduced maximum velocity of shortening (19–22) and a reduced curvature of the force-velocity relationship (19–22). The latter effect was accompanied (20, 21) by a less pronounced deviation of the force-velocity relationship from the hyperbolic shape (23) at high loads. There have been different interpretations of the drug effects. It has been proposed (20–22) that amrinone might competitively inhibit the MgATP binding by myosin. However, more recently, results from in vitro motility assay experiments (24) challenged this idea. These results showed that amrinone reduces the sliding velocity (V_max) at saturating MgATP concentrations but not at MgATP concentrations close to, or below, the K_m value for the hyperbolic relationship between MgATP concentration and sliding velocity. Such a combination of effects is consistent with a reduced MgADP release rate (24) but not with competitive inhibition of substrate binding. However, effects of amrinone on the MgADP release rate have not been directly demonstrated. Additionally, in view of the uncertainty about what step actually determines the sliding velocity at saturating [MgATP] (see above and Refs. 7–9), it is of interest to consider other possible drug effects that could account for the data of Klinth et al. (24). These include the following: 1) an increased drag force, e.g. because of enhancement of weak actomyosin interactions; 2) a reduced step length; and 3) effects of the drug on the rate of MgATP-induced dissociation of actomyosin.Open in a separate windowFIGURE 1.A, structure of amrinone. B, experimental force-velocity data obtained in the presence (filled symbols) and absence (open symbols) of 1.1 mm amrinone. The data, from intact single frog muscle fibers, were obtained at 2 °C and fitted by Hill''s (42) hyperbola (lines) for data truncated at 80% of the maximum isometric force. Filled line, equation fitted to control data, a/P₀* = 0.185; P₀*/P₀ = 1.196. Dashed line, amrinone, a/P₀* = 0.347; P₀*/P₀ = 1.009. Force-velocity data were obtained in collaboration with Professor K. A. P. Edman. Same data as in Fig. 8 of Ref. 20. Note a decrease in maximum sliding velocity and curvature of the force-velocity relationship at low force, in response to amrinone. Also note that amrinone caused increased isometric force and a reduced deviation of the force-velocity relationship from the Hill''s hyperbola at high force. All changes of the force-velocity relationship were statistically significant (20), and similar changes were later also observed in intact mouse muscle and skinned rat muscle fibers. Data in Fig. 1 are published by agreement with Professor K. A. P. Edman.To differentiate between these hypotheses for the amrinone effects, and to gain more general insight into fundamental aspects of muscle function (e.g. mechanisms underlying the force-velocity relationship), we here study the molecular effects of amrinone on fast skeletal muscle myosin preparations in the presence and absence of actin.In vitro motility assay studies at different ionic strengths suggest that drag forces, caused by increased fraction of myosin heads in weak binding states, are not important for the effect of amrinone on sliding velocity. Likewise, optical tweezers studies showed no effect of the drug on the myosin step length. Finally, ideas that amrinone should reduce sliding velocity by reduced rate of MgATP-induced dissociation could be discarded because the drug actually increased the rate of this process. Instead, we found an amrinone-induced increase in the MgADP affinity of heavy meromyosin (HMM) in the presence of actin. Interestingly, similar effects of amrinone were not observed using myosin S1. As discussed below, this result and other results point to an amrinone-induced reduction in the rate of a strain-dependent MgADP release step. Simulations, using a model modified from that of Edman et al. (25), support this proposed mechanism of action. The results are discussed in relation to fundamental mechanisms underlying the force-velocity relationship of fast skeletal muscle, including which step determines shortening velocity and the possible importance of inter-head cooperativity.     

Potential mineralization of nitrogen from organic wastes to ryegrass and wheat crops

Two-pot experiments with ryegrass and wheat plants were conducted in a Cambic Arenosol to test the reliability of N fate predicted by incubation experiments previously performed, with the same soil, to assess potentially mineralizable nitrogen from six organic wastes (municipal solid waste compost, secondary pulp mill sludge, horn meal, poultry manure, solid phase from pig slurry and composted pig manure). Two treatments, corresponding to 80 and 160 kgN/ha were tested, with or without mineral N fertilization. Experimental data obtained in the pot trials was consistent with nitrogen net mineralization trend observed in the aerobic incubations with all the wastes tested. Values of potentially mineralizable nitrogen (N(0)) from the equations obtained by model fitting, to the incubation data, were well correlated to ryegrass and wheat N uptake. Poultry manure was the most efficient N supplier to crops.     

A freshwater cyanophage whose genome indicates close relationships to photosynthetic marine cyanomyophages

Bacteriophage S-CRM01 has been isolated from a freshwater strain of Synechococcus and shown to be present in the upper Klamath River valley in northern California and Oregon. The genome of this lytic T4-like phage has a 178,563 bp circular genetic map with 297 predicted protein-coding genes and 33 tRNA genes that represent all 20-amino-acid specificities. Analyses based on gene sequence and gene content indicate a close phylogenetic relationship to the 'photosynthetic' marine cyanomyophages infecting Synechococcus and Prochlorococcus. Such relatedness suggests that freshwater and marine phages can draw on a common gene pool. The genome can be considered as being comprised of three regions. Region 1 is populated predominantly with structural genes, recognized as such by homology to other T4-like phages and by identification in a proteomic analysis of purified virions. Region 2 contains most of the genes with roles in replication, recombination, nucleotide metabolism and regulation of gene expression, as well as 5 of the 6 signature genes of the photosynthetic cyanomyophages (hli03, hsp20, mazG, phoH and psbA; cobS is present in Region 3). Much of Regions 1 and 2 are syntenic with marine cyanomyophage genomes, except that a segment encompassing Region 2 is inverted. Region 3 contains a high proportion (85%) of genes that are unique to S-CRM01, as well as most of the tRNA genes. Regions 1 and 2 contain many predicted late promoters, with a combination of CTAAATA and ATAAATA core sequences. Two predicted genes that are unusual in phage genomes are homologues of cellular spoT and nusG.     

Zinc stabilizes the SecB binding site of SecA

The molecular chaperone SecB targets preproteins to SecA at the translocation sites in the cytoplasmic membrane of Escherichia coli. SecA recognizes SecB via its carboxyl-terminal 22 aminoacyl residues, a highly conserved domain that contains 3 cysteines and 1 histidine residue that could potentially be involved in the coordination of a metal ion. Treatment of SecA with a zinc chelator resulted in a loss of the stimulatory effect of SecB on the SecA translocation ATPase activity, while the activity could be restored by the addition of ZnCl2. Interaction of SecB with the SecB binding domain of SecA is disrupted by chelators of divalent cations, and could be restored by the addition of Cu2+ or Zn2+. Atomic absorption and electrospray mass spectrometry revealed the presence of one zinc atom per monomeric carboxyl terminus of SecA. It is concluded that the SecB binding domain of SecA is stabilized by a zinc ion that promotes the functional binding of SecB to SecA.     

Ecophysiology in Relation to Exposure of Pendant Epiphytic Bryophytes in the Canopy of a Tropical Montane Oak Forest1

We investigated ecophysiological and morphological traits affecting light and water use of four commercially important pendant epiphytic bryophyte species in a montane oak‐bamboo forest in Costa Rica. Predictions based mostly on ecophysiological studies of temperate bryophytes and vascular plants were experimentally tested on tropical montane bryophyte species ranked on the basis of their distributions in the canopy from the most protected (1) to the most exposed sites (4): (1) Phyllogonium viscosum; (2) Pilotrichella flexilis; (3) Dendropogonella rufescens; and (4) Frullania convoluta. As predicted, highly exposed species tended to have higher light saturation and compensation points, higher dark respiration rates, more chlorophyll, higher chlorophyll a:b ratios, and higher nitrogen concentrations. Contrary to predicted trends, exposed‐site species had lower water contents at full saturation; there were no detectable differences among species in the rate of water loss. Rates of carbon gain in all species reached asymptotes with increasing water content, but the moisture compensation point for carbon uptake of the moss D. rufescens was substantially higher than in the other species. Observed patterns result from interactions among processes related to water storage and transport operating at different scales and are determined by various morphological traits including the density, size, and disposition of phylloids, as well as by whole‐clump architecture.     

Cellulase immobilization on magnetic nanoparticles encapsulated in polymer nanospheres

Immobilization of cellulases on magnetic nanoparticles, especially magnetite nanoparticles, has been the main approach studied to make this enzyme, economically and industrially, more attractive. However, magnetite nanoparticles tend to agglomerate, are very reactive and easily oxidized in air, which has strong impact on their useful life. Thus, it is very important to provide proper surface coating to avoid the mentioned problems. This study aimed to investigate the immobilization of cellulase on magnetic nanoparticles encapsulated in polymeric nanospheres. The support was characterized in terms of morphology, average diameter, magnetic behavior and thermal decomposition analyses. The polymer nanospheres containing encapsulated magnetic nanoparticles showed superparamagnetic behavior and intensity average diameter about 150 nm. Immobilized cellulase exhibited broader temperature stability than in the free form and great reusability capacity, 69% of the initial enzyme activity was maintained after eight cycles of use. The magnetic support showed potential for cellulase immobilization and allowed fast and easy biocatalyst recovery through a single magnet.