Lactate dehydrogenase (LDH) gene duplication during chordate evolution: the cDNA sequence of the LDH of the tunicate Styela plicata

L-Lactate dehydrogenase (L-LDH, E.C. 1.1.1.27) is encoded by two or three loci in all vertebrates examined, with the exception of lampreys, which have a single LDH locus. Biochemical characterizations of LDH proteins have suggested that a gene duplication early in vertebrate evolution gave rise to Ldh-A and Ldh-B and that an additional locus, Ldh-C arose in a number of lineages more recently. Although some phylogenetic studies of LDH protein sequences have supported this pattern of gene duplication, others have contradicted it. In particular, a number of studies have suggested that Ldh-C represents the earliest divergence among vertebrate LDHs and that it may have diverged from the other loci well before the origin of vertebrates. Such hypotheses make explicit statements about the relationship of vertebrate and invertebrate LDHs, but to date, no closely related invertebrate LDH sequences have been available for comparison. We have attempted to provide further data on the timing of gene duplications leading to multiple vertebrate LDHs by determining the cDNA sequence of the LDH of the tunicate Styela plicata. Phylogenetic analyses of this and other LDH sequences provide strong support for the duplications giving rise to multiple vertebrate LDHs having occurred after vertebrates diverged from tunicates. The timing of these LDH duplications is consistent with data from a number of other gene families suggesting widespread gene duplication near the origin of vertebrates. With respect to the relationships among vertebrate LDHs, our data are not consistent with previous claims that Ldh-C represented the earliest divergence. However, the precise relationships among some of the main lineages of vertebrate LDHs were not resolved in our analyses.      

Effect of cholesterol on the membrane interaction of Modelin-5 isoforms

Modelin-5 isoforms were used to gain an insight into the effects of amidation on antimicrobial selectivity. When tested against Escherichia coli, amidation increased toxicity 10-fold (MIC = 31.25 μM) while showing limited increased hemolytic activity (2% lysis). Our results show that both the amidated and non-amidated peptides had a disordered structure in aqueous solution (<18% helical) and folded to form helices at the membrane interface (for example, >43% in the presence of DMPC). The stabilization of the helical structure by amidation has previously been shown to play a key role in increasing antibacterial efficacy. The presence of cholesterol in the membrane increases the packing density (C(s)(-1) values 25-33 mN m(-1)) and so prevents the peptide from forming stable association with the membrane, which is evidenced by the higher binding coefficient (K(d)) in the presence of cholesterol: 57.70 μM for Modelin-5-COOH and 35.64 μM for Modelin-5-CONH(2) compared to the presence of E. coli lipid extract (10 μM), which would prevent local concentration of the peptide at the bilayer interface as seen by reduction in monolayer interaction. This in turn would be predicted to inhibit activity.     

Sexual behavior of laboratory- and wild-born male Rhesus monkeys.

Rearing method has an important influence on the sexual performance of adult male rhesus monkeys. Testosterone levels and sexual behavior of laboratory-born and -reared adult males were compared with those of males born in the wild and brought to the laboratory as adults. The mean sexual performance level of colony-reared animals was significantly below that of males born in the wild but testosterone levels in the two groups did not differ significantly. Not all laboratory-reared males were sexually inadequate but less than 30% ejaculated in tests of sexual behavior. The sexual behavior and testosterone levels of adult males treated prenatally with testosterone propionate were found not to differ from those of untreated males reared in the same way. Conclusions about sexual adequacy based on 10-mm tests of sexual behavior differed very little from those based on 1-hr tests.     

Negotiating British Muslim belonging: a qualitative longitudinal study

British Muslims are often viewed as holding values incompatible with Britishness, regarded with suspicion and sometimes subjected to gendered forms of racism. Research projects have found that identifiably Muslim women face everyday microaggressions, yet little is known about how they negotiate both this and their identities over time. This article addresses this gap by reporting the results of qualitative longitudinal research that explores the narratives of two young British Muslim women over a seven-year period. The women were first interviewed when they were single undergraduates in 2010 and followed up as married young professionals in 2017. On both occasions they were negotiating their identities and sense of belonging in a climate of heightened scrutiny of Muslims. The paper examines their reflections on: “fitting in” with Britishness, their religious identities and the complexity of belonging. Methodologically, it contributes to qualitative longitudinal narrative research.     

Gap junction structures: Analysis of the x-ray diffraction data

Models for the spatial distribution of protein, lipid and water in gap junction structures have been constructed from the results of the analysis of X-ray diffraction data described here and the electron microscope and chemical data presented in the preceding paper (Caspar, D. L. D., D. A. Goodenough, L. Makowski, and W.C. Phillips. 1977. 74:605-628). The continuous intensity distribution on the meridian of the X-ray diffraction pattern was measured, and corrected for the effects of the partially ordered stacking and partial orientation of the junctions in the X-ray specimens. The electron density distribution in the direction perpendicular to the plane of the junction was calculated from the meridional intensity data. Determination of the interference function for the stacking of the junctions improved the accuracy of the electron density profile. The pair-correlation function, which provides information about the packing of junctions in the specimen, was calculated from the interference function. The intensities of the hexagonal lattice reflections on the equator of the X-ray pattern were used in coordination with the electron microscope data to calculate to the two-dimensional electron density projection onto the plane of the membrane. Differences in the structure of the connexons as seen in the meridional profile and equatorial projections were shown to be correlated to changes in lattice constant. The parts of the junction structure which are variable have been distinguished from the invariant parts by comparison of the X-ray data from different specimens. The combination of these results with electron microscope and chemical data provides low resolution three- dimensional representations of the structures of gap junctions.     

Interactions of an anionic antimicrobial peptide with Staphylococcus aureus membranes

The antimicrobial activity of the anionic peptide, AP1 (GEQGALAQFGEWL), was investigated. AP1 was found to kill Staphylococcus aureus with an MLC of 3 mM and to induce maximal surface pressure changes of 3.8 mN m⁻¹ over 1200 s in monolayers formed from lipid extract of S. aureus membranes. FTIR spectroscopy showed the peptide to be α-helical (100%) in the presence of vesicles formed from this lipid extract and to induce increases in their fluidity (Δν circa 0.5 cm⁻¹). These combined data show that AP1 is able to function as an α-helical antimicrobial peptide against Gram-positive bacteria and suggest that the killing mechanism used by the peptide involves interactions with the membrane lipid headgroup region. Moreover, this killing mechanism differs strongly from that previously reported for AP1 against Gram-negative bacteria, indicating the importance of considering the effects of membrane lipid composition when investigating the structure/function relationships of antimicrobial peptides.     

Magnetic resonance imaging of structure, diffusivity, and copper immobilization in a phototrophic biofilm

Magnetic resonance imaging (MRI) was used to spatially resolve structure, water diffusion, and copper transport and fate in a phototrophic biofilm [corrected]. MRI was able to resolve considerable structural heterogeneity, ranging from classical laminations approximately 500 mum thick to structures with no apparent ordering. Pulsed-field gradient (PFG) analysis spatially resolved water diffusion coefficients which exhibited relatively little or no attenuation (diffusion coefficients ranged from 1.7 x 10(-9) m(2) s(-1) to 2.2 x 10(-9) m(2) s(-1)). The biofilm was then reacted with a 10-mg liter(-1) Cu(2+) solution, and transverse relaxation time parameter maps [corrected].were used to spatially and temporally map copper immobilization within the biofilm. Significantly, a calibration protocol similar to that used in biomedical research successfully quantified copper concentrations throughout the biofilm. Variations in Cu concentrations were controlled by the biofilm structure. Copper immobilization was most rapid (approximately 5 mg Cu liter(-1) h(-1)) over the first 20 to 30 h and then much slower for the remaining 60 h of the experiment. The transport of metal within the biofilm is controlled by both diffusion and immobilization. This was explored using a Bartlett and Gardner model which examined both diffusion and adsorption through a hypothetical film exhibiting properties similar to those of the phototrophic biofilm. Higher adsorption constants (K) resulted in longer lag times until the onset of immobilization at depth but higher actual adsorption rates. MRI and reaction transport models are versatile tools which can significantly improve our understanding of heavy metal immobilization in naturally occurring biofilms.     

The impacts of UV-B radiation on the regeneration of a sub-arctic heath community

The effects of ultraviolet-B radiation on regeneration after disturbance of a natural sub-arctic heathland have been investigated. Areas of pristine dwarf shrub heath were denuded of all above ground biomass in 1992 and exposed to enhanced UV-B (simulating a 15% depletion of the ozone layer). The resulting regenerated stem and leaf growth parameters were measured after four years on three dwarf shrubs, Vaccinium myrtillus, V. uliginosum and V. vitis-idaea and the grass Calamagrostis lapponica; leaves of the three dwarf shrubs were also analysed for UV-absorbing compounds and carbohydrates. Regeneration irrespective of treatment was slow, with Empetrum hermaphroditum failing to regenerate at all. Vaccinium myrtillus showed the most rapid regeneration attaining much of its original biomass in four years. There was a significant interaction between UV-B and year of regeneration in V. myrtillus; annual stem length increment showed an initial stimulation of 75% under enhanced UV-B in the first year of regeneration while a reduction of 16% was observed in the fourth year. Both V. uliginosum and V. vitis-idaea showed a reduction in annual stem length increment as regeneration progressed with a greater than 50% reduction in stem increment in the fourth year of regeneration compared to the first. Vaccinium uliginosum also showed an initial reduction in stem length increment of 40% under enhanced UV-B. None of the species were affected by enhanced UV-B in terms of total regenerated stem and leaf biomass or UV-absorbing compounds in regenerated leaf tissue. Total leaf carbohydrate and the ethanol/water soluble fraction in V. uliginosum were significantly increased by 29% and 31% respectively under enhanced UV-B. This suggests either a stimulation of photosynthesis or a reduction in sink size for photo-assimilates. Results are discussed in the context of the extremely slow regeneration of sub-arctic heath communities and the implications of contrasting UV-B effects on the regenerative ability of different species.