Mitochondrial mislocalization and altered assembly of a cluster of Barth syndrome mutant tafazzins

None of the 28 identified point mutations in tafazzin (Taz1p), which is the mutant gene product associated with Barth syndrome (BTHS), has a biochemical explanation. In this study, endogenous Taz1p was localized to mitochondria in association with both the inner and outer mitochondrial membranes facing the intermembrane space (IMS). Unexpectedly, Taz1p does not contain transmembrane (TM) segments. Instead, Taz1p membrane association involves a segment that integrates into, but not through, the membrane bilayer. Residues 215-232, which were predicted to be a TM domain, were identified as the interfacial membrane anchor by modeling four distinct BTHS mutations that occur at conserved residues within this segment. Each Taz1p mutant exhibits altered membrane association and is nonfunctional. However, the basis for Taz1p dysfunction falls into the following two categories: (1) mistargeting to the mitochondrial matrix or (2) correct localization associated with aberrant complex assembly. Thus, BTHS can be caused by mutations that alter Taz1p sorting and assembly within the mitochondrion, indicating that the lipid target of Taz1p is resident to IMS-facing leaflets.     

Symmetry breaking in interspecific Drosophila hybrids is not due to developmental noise

Hybrids from crosses of different species have been reported to display decreased developmental stability when compared to their pure species, which is conventionally attributed to a breakdown of coadapted gene complexes. Drosophila subobscura and its close relative D. madeirensis were hybridized in the laboratory to test the hypothesis that genuine fluctuating asymmetry, measured as the within-individual variance between right and left wings that results from random perturbations in development, would significantly increase after interspecific hybridization. When sires of D. subobscura were mated to heterospecific females following a hybrid half-sib breeding design, F1 hybrid females showed a large bilateral asymmetry with a substantial proportion of individuals having an asymmetric index larger than 5% of total wing size. Such an anomaly, however, cannot be plainly explained by an increase of developmental instability in hybrids but is the result of some aberrant developmental processes. Our findings suggest that interspecific hybrids are as able as their parents to buffer developmental noise, notwithstanding the fact that their proper bilateral development can be harshly compromised. Together with the low correspondence between the co-variation structures of the interindividual genetic components and the within-individual ones from a Procrustes analysis, our data also suggest that the underlying processes that control (genetic) canalization and developmental stability do not share a common mechanism. We argue that the conventional account of decreased developmental stability in interspecific hybrids needs to be reappraised.     

Multiple interactions of FbsA, a surface protein from Streptococcus agalactiae, with fibrinogen: affinity, stoichiometry, and structural characterization

Streptococcus agalactiae is an etiological agent of several infective diseases in humans. We previously demonstrated that FbsA, a fibrinogen-binding protein expressed by this bacterium, elicits a fibrinogen-dependent aggregation of platelets. In the present communication, we show that the binding of FbsA to fibrinogen is specific and saturable, and that the FbsA-binding site resides in the D region of fibrinogen. In accordance with the repetitive nature of the protein, we found that FbsA contains multiple binding sites for fibrinogen. By using several biophysical methods, we provide evidence that the addition of FbsA induces extensive fibrinogen aggregation and has noticeable effects on thrombin-catalyzed fibrin clot formation. Fibrinogen aggregation was also found to depend on FbsA concentration and on the number of FbsA repeat units. Scanning electron microscopy evidentiated that, while fibrin clot is made of a fine fibrillar network, FbsA-induced Fbg aggregates consist of thicker fibers organized in a cage-like structure. The structural difference of the two structures was further indicated by the diverse immunological reactivity and capability to bind tissue-type plasminogen activator or plasminogen. The mechanisms of FbsA-induced fibrinogen aggregation and fibrin polymerization followed distinct pathways since Fbg assembly was not inhibited by GPRP, a specific inhibitor of fibrin polymerization. This finding was supported by the different sensitivity of the aggregates to the disruptive effects of urea and guanidine hydrochloride. We suggest that FbsA and fibrinogen play complementary roles in contributing to thrombogenesis associated with S. agalactiae infection.     

Role of loop bundle hydrogen bonds in the maturation and activity of (Pro)caspase-3

During maturation, procaspase-3 is cleaved at D175, which resides in a linker that connects the large and small subunits. The intersubunit linker also connects two active site loops that rearrange following cleavage and, in part, form the so-called loop bundle. As a result of chain cleavage, new hydrogen bonds and van der Waals contacts form among three active site loops. The new interactions are predicted to stabilize the active site. One unresolved issue is the extent to which the loop bundle residues also stabilize the procaspase active site. We examined the effects of replacing four loop bundle residues (E167, D169, E173, and Y203) on the biochemical and structural properties of the (pro)caspase. We show that replacing the residues affects the activity of the procaspase as well as the mature caspase, with D169A and E167A replacements having the largest effects. Replacement of D169 prevents caspase-3 autoactivation, and its cleavage at D175 no longer leads to an active enzyme. In addition, the E173A mutation, when coupled to a second mutation in the procaspase, D175A, may alter the substrate specificity of the procaspase. The mutations affected the active site environment as assessed by changes in fluorescence emission, accessibility to quencher, and cleavage by either trypsin or V8 proteases. High-resolution X-ray crystallographic structures of E167A, D173A, and Y203F caspases show that changes in the active site environment may be due to the increased flexibility of several residues in the N-terminus of the small subunit. Overall, the results show that these residues are important for stabilizing the procaspase active site as well as that of the mature caspase.     

Economy and environment of Bronze Age settlements – Terramaras – on the Po Plain (Northern Italy): first results from the archaeobotanical research at the Terramara di Montale

The paper presents a synthesis of the on-site archaeobotanical investigations of the Terramara di Montale, one of the most important sites of the Terramara cultural system which characterised the Po Plain in the Middle-Late Bronze Age (1650–1200 b.c.). Samples for pollen analysis and macroremains, including seed/fruit and wood/charcoal records, were collected from stratigraphic sequences and occupation levels during the excavations 1996–2001. The results permitted the reconstruction of the main characteristics of the landscape which at the onset of the Terramara rapidly passed from a natural, more forested landscape with mixed oak wood and conifers to a more open and anthropic landscape characterised by cereal fields, pastures and meadows. People felled oaks and other trees such as Populus/Salix and Fraxinus to make piles or walls for houses. Wood from these species was also recorded as charcoal in the hearths. Palynological and carpological data show that the inhabitants of the Terramara largely founded their economy on cereals (mainly Triticum aestivum/durum, T. dicoccum and Hordeum vulgare). They also grew a few legumes (Vicia faba var. minor, Vicia sp. and Lens culinaris). There was also grazing by domestic animals, mainly ovicaprines but also pigs and cattle, and these were fed exploiting wild plants such as Carpinus. In the paper the four main steps of the history of the Terramara are described (before the Terramara, the onset, the Terramara phase, the decline) during which both human influence and climatic changes were important. At the onset of the Terramara (around 1600 b.c.) a warm and possibly dry phase occurred. The intense use of the territory and a climatic deterioration at around 1300 b.c. might have triggered the decline of the Terramara di Montale.     

Supraspinal Gβγ-dependent stimulation of PLCβ3 originating from G inhibitory protein-μ opioid receptor-coupling is necessary for morphine induced acute hyperalgesia

Although alterations in μ-opioid receptor (μOR) signaling mediate excitatory effects of opiates in opioid tolerance, the molecular mechanism for the excitatory effect of acute low dose morphine, as it relates to μOR coupling, is presently unknown. A pronounced coupling of μOR to the α subunit of G inhibitory protein emerged in periaqueductal gray (PAG) from mice systemically administered with morphine at a dose producing acute thermal hyperalgesia. This coupling was abolished in presence of the selective μOR antagonist d -Phe–Cys–Tyr– d -Trp–Orn–Thr–Pen–Thr–NH₂ administered at the PAG site, showing that the low dose morphine effect is triggered by μOR activated G inhibitory protein at supraspinal level. When Gβγ downstream signalling was blocked by intra-PAG co-administration of 2-(3,4,5-trihydroxy-6-oxoxanthen-9-yl)cyclohexane-1-carboxylic acid, a compound that inhibits Gβγ dimer-dependent signaling, a complete prevention of low dose morphine induced acute thermal hyperalgesia was obtained. Phospholipase C β3, an enzyme necessary to morphine hyperalgesia, was revealed to be associated with Gβγ in PAG. Although opioid administration induces a shift in μOR-G protein coupling from Gi to Gs after chronic administration, our data support that this condition is not realized in acute treatment providing evidence that a separate molecular mechanism underlies morphine induced acute excitatory effect.     

Transduction mechanism(s) of Na-saccharin in the blowfly <Emphasis Type="Italic">Protophormia terraenovae</Emphasis>: evidence for potassium and calcium conductance involvement

The study on transduction mechanisms underlying bitter stimuli is a particularly intriguing challenge for taste researchers. The present study investigates, in the labellar chemosensilla of the blowfly Protophormia terraenovae, the transduction mechanism by which saccharin evokes the response of the “deterrent” cell, with particular attention to the contribution of K⁺ and Ca²⁺ current and the role of cyclic nucleotides, since second messengers modulate Ca²⁺, Cl⁻ and K⁺ currents to different extents. As assessed by extracellular single-sensillum recordings, our results show that the addition of a Ca²⁺ chelator such as EGTA or the Ca²⁺ current blockers SK&F-96365, Mibefradil, Nifedipine and W-7 decrease the response of the “deterrent” cell to saccharin. A similar decreasing effect was also obtained following the addition of 4-aminopyridine, a K⁺ current blocker. On the contrary, the membrane-permeable cyclic nucleotide 8-bromoguanosine 3′,5′-cyclic monophosphate (8Br-cGMP) activates this cell and shows an additive effect when presented mixed with saccharin. Our results are consistent with the hypothesis that in the labellar chemosensilla of the blowfly both Ca²⁺ and K⁺ ions are involved in the transduction mechanism of the “deterrent” cell in response to saccharin. Our results also suggest a possible pathway common to saccharin and 8Br-cGMP.     

Cell wall adaptations of planktonic and biofilm <Emphasis Type="Italic">Rhodococcus erythropolis</Emphasis> cells to growth on C5 to C16 <Emphasis Type="Italic">n</Emphasis>-alkane hydrocarbons

Rhodococcus erythropolis was found to utilize C5 to C16 n-alkane hydrocarbons as sole source of carbon and energy when growing as planktonic or biofilm cells attached to polystyrene surfaces. Growth rates on even numbered were two- to threefold increased relatively to odd-numbered n-alkanes and depended on the chain length of the n-alkanes. C10-, C12-, C14-, and C16-n-alkanes gave rise to two- to fourfold increased maximal growth rates relative to C5- to C9-hydrocarbons. In reaction to the extremely poor water solubility of the n-alkanes, both planktonic and biofilm cells exhibited distinct adaptive changes. These included the production of surface active compounds and substrate-specific modifications of the physicochemical cell surface properties as expressed by the microbial adhesion to hydrocarbon- and contact angle-based hydrophobicity, as well as the zeta potential of the cells. By contrast, n-alkane-specific alterations of the cellular membrane composition were less distinct. The specificity of the observed autecological changes suggest that R. erythropolis cells may be used in the development and application of sound biocatalytic processes using n-alkanes as substrates or substrate reservoirs or for target-specific bioremediation of oils and combustibles, respectively.     

Omega-3 consumption and sudden cardiac death in schizophrenia

People with schizophrenia show a two- to three-fold increased risk to die prematurely. Mortality is accounted for by a combination of factors (patients’ life style, suicide, premature cardiovascular disease, metabolic syndromes and, not so often mentioned, sudden death). The cause of sudden death in schizophrenia is unknown, but cardiac arrhythmia plays a potential role. Patients with schizophrenia are at high risk for cardiovascular disease, and some antipsychotics may be associated with cardiovascular adverse events (e.g., electrocardiograph QT interval prolongation), suggesting that this could lead to sudden cardiac death. Animal and clinical studies have shown that omega-3 fatty acids could be useful in the prevention and treatment of schizophrenia. As omega-3 fatty acids have been considered a cardioprotector agent, reducing cardiac arrhythmias and hence sudden cardiac deaths and given their relative safety and general health benefits, our update article summarizes the knowledge by the possible positive effects of omega-3 supplementation and fish consumption against sudden cardiac death in patients with schizophrenia. However, fish species should be selected with caution due to contamination with toxic methylmercury.