Purification and characterization of an activator protein for methanol dehydrogenase from thermotolerant Bacillus spp

All thermotolerant methanol-utilizing Bacillus spp. investigated by us possess a NAD-dependent methanol dehydrogenase (MDH) activity which is stimulated by a protein present in the soluble fraction of Bacillus sp. C1 cells. This activator protein was purified to homogeneity from Bacillus sp. C1 cells grown at a low dilution rate in a methanol-limited chemostat culture. The native activator protein (Mr = 50,000) is a dimer of Mr = 27,000 subunits. The N-terminal amino acid sequence revealed no significant similarity with any published sequences. Stimulation of MDH activity by the activator protein required the presence of Mg2+ ions. Plots of specific MDH activity versus activator protein concentration revealed Michaelis-Menten type kinetics. In the presence of activator protein, MDH displayed biphasic kinetics (v versus substrate concentration) toward C1-C4 primary alcohols and NAD. The data suggest that in the presence of activator protein plus Mg2+ ions, MDH possesses a high affinity active site for alcohols and NAD, in addition to an activator- and Mg2(+)-independent low affinity active site. The activation mechanism remains to be elucidated.     

Intermediate sucrose octa-acetate sensitivity suggests a third allele at mouse bitter taste locus Soa and Soa-Rua identity

Mice have been characterized as either tasters or non-tastersof the bitter compound sucrose octa-acetate(SOA). However, 11of 17 supposedly non-taster inbred strains were found to avoid1 mM SOA. All 17 strains were indifferent to 0.1 mM SOA. Tasterstrains avoided both concentrations. The intermediate phenotypewas dubbed demitaster. A consistent phenotypic dominance orderwas found in crosses among both inbred and outbred strains (taster> non-taster > demitaster). Demitasters were found (withtasters) in an outbred strain showing monogenic segregationfor SOA avoidance. This, plus monogenic segregation in a back-crossof taster to demitaster inbred strains, suggested a third alleleat the Soa locus (Soa^c). Demitaster allelism was supported bythe strong associations found in 15 strains between the threeSOA phenotypes and HindIII restriction fragment patterns forthe closely linked Prp (proline rich protein) loci. SOA demitasterstrains were also intermediate in raffinose undeca-acetate (RUA)avoidance. Furthermore, B6.SW-Soa² congenic mice avoided notonly SOA, but RUA and eight other acetylated sugars. A previouslyproposed separate RUA-sensitivity gene (Rua) thus appeared tobe redundant.     

Floral adaptation and diversification under pollen limitation

Pollen limitation (PL) of seed production creates unique conditions for reproductive adaptation by angiosperms, in part because, unlike under ovule or resource limitation, floral interactions with pollen vectors can contribute to variation in female success. Although the ecological and conservation consequences of PL have received considerable attention in recent times, its evolutionary implications are poorly appreciated. To identify general influences of PL on reproductive adaptation compared with those under other seed-production limits and their implications for evolution in altered environments, we derive a model that incorporates pollination and post-pollination aspects of PL. Because PL always favours increased ovule fertilization, even when population dynamics are not seed limited, it should pervasively influence selection on reproductive traits. Significantly, under PL the intensity of inbreeding does not determine whether outcrossing or autonomous selfing can evolve, although it can affect which response is most likely. Because the causes of PL are multifaceted in both natural and anthropogenically altered environments, the possible outcrossing solutions are diverse and context dependent, which may contribute to the extensive variety of angiosperm reproductive characteristics. Finally, the increased adaptive options available under PL may be responsible for positive global associations between it and angiosperm diversity.     

Malaria Parasite Actin Polymerization and Filament Structure

A novel form of acto-myosin regulation has been proposed in which polymerization of new actin filaments regulates motility of parasites of the apicomplexan class of protozoa. In vivo and in vitro parasite F-actin is very short and unstable, but the structural basis and details of filament dynamics remain unknown. Here, we show that long actin filaments can be obtained by polymerizing unlabeled rabbit skeletal actin (RS-actin) onto both ends of the short rhodamine-phalloidin-stabilized Plasmodium falciparum actin I (Pf-actin) filaments. Following annealing, hybrid filaments of micron length and “zebra-striped” appearance are observed by fluorescence microscopy that are stable enough to move over myosin class II motors in a gliding filament assay. Using negative stain electron microscopy we find that pure Pf-actin stabilized by jasplakinolide (JAS) also forms long filaments, indistinguishable in length from RS-actin filaments, and long enough to be characterized structurally. To compare structures in near physiological conditions in aqueous solution we imaged Pf-actin and RS-actin filaments by atomic force microscopy (AFM). We found the monomer stacking to be distinctly different for Pf-actin compared with RS-actin, such that the pitch of the double helix of Pf-actin filaments was 10% larger. Our results can be explained by a rotational angle between subunits that is larger in the parasite compared with RS-actin. Modeling of the AFM data using high-resolution actin filament models supports our interpretation of the data. The structural differences reported here may be a consequence of weaker inter- and intra-strand contacts, and may be critical for differences in filament dynamics and for regulation of parasite motility.     

Comparison of Cobb Angles Measured Manually,Calculated from 3-D Spinal Reconstruction,and Estimated from Torso Asymmetry

While scoliotic spinal deformity is traditionally measured by the Cobb angle, we seek to estimate scoliosis severity from the torso surface without X-ray radiation. Here, we measured the Cobb angle in three ways: by protractor from postero-anterior X-ray, by computer from a 3-D digitized model of the vertebral body line, and by neural-network estimation from indices of torso surface asymmetry. The estimates of the Cobb angle by computer and by neural network were equally accurate in 153 records from 52 patients (standard deviation of 6° from the Cobb angle, r =0.93 ), showing that torso asymmetry reliably predicted spinal deformity. Further improvements in predictive accuracy may require estimation of other 3-D indices of spinal deformity besides the Cobb angle with its wide measurement variability.     

Suicidal process,suicidal communication and psychosocial situation of young suicide attempters in a rural Vietnamese community

The study aimed to explore the suicidal process, suicidal communication and psychosocial situation of young suicide attempters in a rural community in Hanoi, Vietnam. Semi-structured interviews were conducted, in a community setting, with 19 suicide attempters aged 15-24 who had been consecutively hospitalized in an intensive care unit. In 12 of 19 cases, the first pressing, distinct and constant suicidal thoughts appeared less than one day before the suicide attempt in question. However, distress and mild, fleeting suicidal thoughts had been present up to six months before the suicide attempt in 16 cases. Five respondents had a suicide plan one to three days before attempting suicide. Altogether, 13 engaged in some form of suicidal communication before their attempt. This communication was, however, difficult for outsiders to interpret. Twelve of the respondents were victims of regular physical abuse and 16 had suffered psychological violence for at least one year before attempting suicide. Eighteen of the respondents used pesticides or raticides in their suicide attempts. None sought advice or consultation in the community despite long-standing psychosocial problems. The strategy of reducing the availability of suicide means (e.g., pesticides or raticides) in Asian countries should be complemented with a long-term suicide-preventive strategy that targets school dropouts and domestic violence, and promotes coping abilities and communication about psychological and social problems as well as recognition of signs of distress and suicidal communication.     

The evolution of floral nectaries in Disa (Orchidaceae: Disinae): recapitulation or diversifying innovation?

Background and Aims

The Orchidaceae have a history of recurring convergent evolution in floral function as nectar production has evolved repeatedly from an ancestral nectarless state. However, orchids exhibit considerable diversity in nectary type, position and morphology, indicating that this convergence arose from alternative adaptive solutions. Using the genus Disa, this study asks whether repeated evolution of floral nectaries involved recapitulation of the same nectary type or diversifying innovation. Epidermis morphology of closely related nectar-producing and nectarless species is also compared in order to identify histological changes that accompanied the gain or loss of nectar production.

Methods

The micromorphology of nectaries and positionally equivalent tissues in nectarless species was examined with light and scanning electron microscopy. This information was subjected to phylogenetic analyses to reconstruct nectary evolution and compare characteristics of nectar-producing and nectarless species.

Key Results

Two nectary types evolved in Disa. Nectar exudation by modified stomata in floral spurs evolved twice, whereas exudation by a secretory epidermis evolved six times in different perianth segments. The spur epidermis of nectarless species exhibited considerable micromorphological variation, including strongly textured surfaces and non-secreting stomata in some species. Epidermis morphology of nectar-producing species did not differ consistently from that of rewardless species at the magnifications used in this study, suggesting that transitions from rewardlessness to nectar production are not necessarily accompanied by visible morphological changes but only require sub-cellular modification.

Conclusions

Independent nectary evolution in Disa involved both repeated recapitulation of secretory epidermis, which is present in the sister genus Brownleea, and innovation of stomatal nectaries. These contrasting nectary types and positional diversity within types imply weak genetic, developmental or physiological constraints in ancestral, nectarless Disa. Such functional convergence generated by morphologically diverse solutions probably also underlies the extensive diversity of nectary types and positions in the Orchidaceae.