Epibatidine isomers and analogues: structure-activity relationships

Binding affinities for a range of epibatidine isomers and analogues at the alpha4beta2 and alpha3beta4 nAChR subtypes are reported; compounds having similar N-N distances to epibatidine show similar, high potencies.     

Kin discrimination and altruism in the larvae of a solitary insect

Kin selection theory predicts altruism between related individuals, which requires the ability to recognize kin from non-kin. In insects, kin discrimination associated with altruistic behaviour is well-known in clonal and social species but in very few solitary insects. Here, we report that the solitary larvae of a non-social insect Aleochara bilineata Gyll. (Coleoptera; Staphylinidae) show kin discrimination and sibling-directed altruistic behaviour. Larvae superparasitize more frequently the hosts parasitized by non-kin individuals than those hosts parasitized by siblings. Kin discrimination probably occurs by self-referent phenotype matching, where an individual compares its own phenotype with that of a non-familiar related individual, a mechanism rarely demonstrated in animals. The label used to recognize kin from non-kin corresponds to substances contained in the plug placed on the hosts by the resident larvae during the parasitization process. Kin competition induced by a limited larval dispersion may have favoured the evolution of kin recognition in this solitary species.     

Perception and modification of plant flavonoid signals by rhizosphere microorganisms

Flavonoids are a diverse class of polyphenolic compounds that are produced as a result of plant secondary metabolism. They are known to play a multifunctional role in rhizospheric plant-microbe and plant-plant communication. Most familiar is their function as a signal in initiation of the legume-rhizobia symbiosis, but, flavonoids may also be signals in the establishment of arbuscular mycorrhizal symbiosis and are known agents in plant defence and in allelopathic interactions. Flavonoid perception by, and impact on, their microbial targets (e.g. rhizobia, plant pathogens) is relatively well characterized. However, potential impacts on 'non-target' rhizosphere inhabitants ('non-target' is used to distinguish those microorganisms not conventionally known as targets) have not been thoroughly investigated. Thus, this review first summarizes the conventional roles of flavonoids as nod gene inducers, phytoalexins and allelochemicals before exploring questions concerning 'non-target' impacts. We hypothesize that flavonoids act to shape rhizosphere microbial community structure because they represent a potential source of carbon and toxicity and that they impact on rhizosphere function, for example, by accelerating the biodegradation of xenobiotics. We also examine the reverse question, 'how do rhizosphere microbial communities impact on flavonoid signals?' The presence of microorganisms undoubtedly influences the quality and quantity of flavonoids present in the rhizosphere, both through modification of root exudation patterns and microbial catabolism of exudates. Microbial alteration and attenuation of flavonoid signals may have ecological consequences for below-ground plant-microbe and plant-plant interaction. We have a lack of knowledge concerning the composition, concentration and bioavailability of flavonoids actually experienced by microbes in an intact rhizosphere, but this may be addressed through advances in microspectroscopic and biosensor techniques. Through the use of plant mutants defective in flavonoid biosynthesis, we may also start to address the question of the significance of flavonoids in shaping rhizosphere community structure and function.     

Transthyretin, a new cryptic protease

Transthyretin (TTR) is a plasma homotetrameric protein that acts physiologically as a transporter of thyroxine (T(4)) and retinol, in the latter case through binding to retinol-binding protein (RBP). A fraction of plasma TTR is carried in high density lipoproteins by binding to apolipoprotein AI (apoA-I). We further investigated the nature of the TTR-apoA-I interaction and found that TTR from different sources (recombinant and plasmatic) is able to process proteolytically apoA-I, cleaving its C terminus after Phe-225. TTR-mediated proteolysis was inhibited by serine protease inhibitors (phenylmethanesulfonyl fluoride, Pefabloc, diisopropyl fluorophosphate, chymostatin, and N(alpha)-p-tosyl-l-phenylala-nine-chloromethyl ketone), suggesting a chymotrypsin-like activity. A fluorogenic substrate corresponding to an apoA-I fragment encompassing amino acid residues 223-228 (Abz-ESFKVS-EDDnp) was used to characterize the catalytic activity of TTR, including optimum reaction conditions (37 degrees C and pH 6.8) and catalytic constant (K(m) = 29 microm); when complexed with RBP, TTR activity was lost, whereas when complexed with T(4), only a slight decrease was observed. Cell lines expressing TTR were able to degrade Abz-ESFKVS-EDDnp 2-fold more efficiently than control cells lacking TTR expression; this effect was reversed by the presence of RBP in cell culture media, therefore proving a TTR-specific proteolytic activity. TTR can act as a novel plasma cryptic protease and might have a new, potentially important role under physiological and/or pathological conditions.     

Neurons expressing the highest levels of gamma-synuclein are unaffected by targeted inactivation of the gene

Homologous recombination in ES cells was employed to generate mice with targeted deletion of the first three exons of the gamma-synuclein gene. Complete inactivation of gene expression in null mutant mice was confirmed on the mRNA and protein levels. Null mutant mice are viable, are fertile, and do not display evident phenotypical abnormalities. The effects of gamma-synuclein deficiency on motor and peripheral sensory neurons were studied by various methods in vivo and in vitro. These two types of neurons were selected because they both express high levels of gamma-synuclein from the early stages of mouse embryonic development but later in the development they display different patterns of intracellular compartmentalization of the protein. We found no difference in the number of neurons between wild-type and null mutant animals in several brain stem motor nuclei, in lumbar dorsal root ganglia, and in the trigeminal ganglion. The survival of gamma-synuclein-deficient trigeminal neurons in various culture conditions was not different from that of wild-type neurons. There was no difference in the numbers of myelinated and nonmyelinated fibers in the saphenous nerves of these animals, and sensory reflex thresholds were also intact in gamma-synuclein null mutant mice. Nerve injury led to similar changes in sensory function in wild-type and mutant mice. Taken together, our data suggest that like alpha-synuclein, gamma-synuclein is dispensable for the development and function of the nervous system.     

Effect of the Colorless non-ripening mutation on cell wall biochemistry and gene expression during tomato fruit development and ripening

The Colorless non-ripening (Cnr) mutation in tomato (Solanum lycopersicum) results in mature fruits with colorless pericarp tissue showing an excessive loss of cell adhesion (A.J. Thompson, M. Tor, C.S. Barry, J. Vrebalov, C. Orfila, M.C. Jarvis, J.J. Giovannoni, D. Grierson, G.B. Seymour [1999] Plant Physiol 120: 383-390). This pleiotropic mutation is an important tool for investigating the biochemical and molecular basis of cell separation during ripening. This study reports on the changes in enzyme activity associated with cell wall disassembly in Cnr and the effect of the mutation on the program of ripening-related gene expression. Real-time PCR and biochemical analysis demonstrated that the expression and activity of a range of cell wall-degrading enzymes was altered in Cnr during both development and ripening. These enzymes included polygalacturonase, pectinesterase (PE), galactanase, and xyloglucan endotransglycosylase. In the case of PE, the protein product of the ripening-related isoform PE2 was not detected in the mutant. In contrast with wild type, Cnr fruits were rich in basic chitinase and peroxidase activity. A microarray and differential screen were used to profile the pattern of gene expression in wild-type and Cnr fruits. They revealed a picture of the gene expression in the mutant that was largely consistent with the real-time PCR and biochemical experiments. Additionally, these experiments demonstrated that the Cnr mutation had a profound effect on many aspects of ripening-related gene expression. This included a severe reduction in the expression of ripening-related genes in mature fruits and indications of premature expression of some of these genes in immature fruits. The program of gene expression in Cnr resembles to some degree that found in dehiscence or abscission zones. We speculate that there is a link between events controlling cell separation in tomato, a fleshy fruit, and those involved in the formation of dehiscence zones in dry fruits.     

Inhibition of myostatin in adult mice increases skeletal muscle mass and strength

A human therapeutic that specifically modulates skeletal muscle growth would potentially provide a benefit for a variety of conditions including sarcopenia, cachexia, and muscular dystrophy. Myostatin, a member of the TGF-beta family of growth factors, is a known negative regulator of muscle mass, as mice lacking the myostatin gene have increased muscle mass. Thus, an inhibitor of myostatin may be useful therapeutically as an anabolic agent for muscle. However, since myostatin is expressed in both developing and adult muscles, it is not clear whether it regulates muscle mass during development or in adults. In order to test the hypothesis that myostatin regulates muscle mass in adults, we generated an inhibitory antibody to myostatin and administered it to adult mice. Here we show that mice treated pharmacologically with an antibody to myostatin have increased skeletal muscle mass and increased grip strength. These data show for the first time that myostatin acts postnatally as a negative regulator of skeletal muscle growth and suggest that myostatin inhibitors could provide a therapeutic benefit in diseases for which muscle mass is limiting.     

Identification of human and mouse CatSper3 and CatSper4 genes: Characterisation of a common interaction domain and evidence for expression in testis

Background  

CatSper1 and CatSper2 are two recently identified channel-like proteins, which show sperm specific expression patterns. Through targeted mutagenesis in the mouse, CatSper1 has been shown to be required for fertility, sperm motility and for cAMP induced Ca²⁺ current in sperm. Both channels resemble a single pore forming repeat from a four repeat voltage dependent Ca²⁺ /Na⁺ channel. However, neither CatSper1 or CatSper2 have been shown to function as cation channels when transfected into cells, singly or in conjunction. As the pore forming units of voltage gated cation channels form a tetramer it has been suggested that the known CatSper proteins require additional subunits and/or interaction partners to function.