Mountain birch under multiple stressors – heavy metal‐resistant populations co‐resistant to biotic stress but maladapted to abiotic stress

Stress adaptations often include a trade‐off of weakened performance in nonlocal conditions, resulting in divergent selection, and potentially, genetic differentiation and evolutionary adaptation. Results of a two‐phase (greenhouse and field) common garden experiment demonstrated adaptation of mountain birch (Betula pubescens subsp. czerepanovii) populations from industrially polluted areas of the Kola Peninsula, north‐western Russia, to heavy metals (HM), whereas no adaptations to wind or drought stress were detected in populations from wind‐exposed sites. HM‐adapted seedlings were maladapted to drought but less palatable (co‐resistant) to insect herbivores, even under background HM concentrations. The absence of adaptations to harsh microclimate and the generally high adaptive potential of mountain birch, a critical forest forming tree in subarctic Europe, need to be accounted for in models predicting consequences of human‐driven environmental changes, including the projected climate change.     

Heterogeneity in the structural dynamics of Sulfolobus solfataricus beta-glycosidase revealed by electron paramagnetic resonance and frequency domain fluorometry

The local and global dynamics of the Sulfolobus solfataricus beta-glycosidase were studied by electron spin resonance and time-resolved fluorescence techniques. For electron paramagnetic resonance (EPR) investigations, the protein was covalently modified by the maleimido nitroxide spin label, which is specific for cysteine -SH groups, at position 344 and at position 101, where Ser-101 was changed into a cysteine by site-directed mutagenesis. The greater reactivity of exposed Cys-101 suggested the exclusive modification of this amino acid compared with Cys-344. The labeled proteins underwent temperature perturbation in the range 290-335 K and the values of the spin-label rotation correlation frequencies (nu(c)) ranged from 6 x 10(7) to 2 x 10(8) sec(-1) for the protein labeled at position C344 and from 5.62 x 10(7) to 1.10 x 10(8) sec(-1) for the protein labeled at C101. These rotation correlation values are related to the local dynamic characteristics of the protein matrix. The temperature dependence of rotation correlation frequencies expressed in terms of Arrhenius coordinates (log (nu(c)) vs. 1/T) for the protein labeled at C344 exhibited a linear dependence but with a change in the slope at 311 K. For the protein labeled at C101, no change in the slope was observed at the same temperature. General dynamic information was deduced from the analysis of the fluorescence emission decay of the tryptophanyl residues that are present in each region of the protein structure. Fluorescence data analysis highlighted a bimodal distribution of fluorescence lifetimes arising from the contribution of two emitting groups: one consisting of closely clustered tryptophans responsible for the long-lived emission component (7.1 nsec) and the other composed of tryptophans nearer to the protein surface, which can be associated to the short-lived component (2.5 nsec). The temperature dependence of lifetime distribution parameters linked to the long-lived and short-lived components, expressed in Arrhenius coordinates, showed two different points in which the change in the slope occurred (i.e., 328 K and 338 K, respectively). The Arrhenius analysis of data provided the activation energy relative to the conformational changes characterizing the local and global movements running through the protein matrix.     

Molecular dynamics investigation of an antibody binding site by the fluorescence-photochrome method

A combined fluorescence-photochrome approach was used for investigation of the molecular dynamics antiDNP antibody binding site and its cavity. A 4-(N-2,4-dinitrophenylamino)-4'-(N,N'-dimethylamino)stilbene (StDNP) fluorescence DNP analog was incorporated into the antibody binding site. This was followed by measurements of fluorescence and photochrome parameters such as the StDNP excitation and emission spectra, fluorescence lifetime, steady-state and time-resolved fluorescence polarization, kinetics of trans-cis and cis-trans photoisomerization, and fluorescence quenching by nitroxide radicals freely diffused in solution. In parallel, computational modeling studies on the location and dynamics of DNP/TEMPO spin-label (NslDNP) and StDNP guests within a model of the binding site were performed. When all the experimental evidence is considered (including data from the antibody X-ray study), one can conclude that wobbling of the Trp 91 L/Trp 96 H binding-site.bound-hapten moiety (StDNP), can be responsible for the label's nanosecond dynamics monitored by fluorescence polarization techniques. A similar conclusion may be reached as a result of data analysis on NslDNP mobility within the antibody binding site. The mobility of Trp 91 L and Trp 96 H moieties provides the induced fit needed for effective stacking and release of the DNP epitope. Analysis of the above-mentioned data allows one to explore the mechanism of the probe's movement within the binding site and enables one to discuss the local dynamics of the binding site region. The combined fluorescence-photochrome approach can be used for investigation of local medium molecular dynamics in the immediate vicinity of specific sites of proteins and nucleic acids, as well as for other biologically important structures and synthetic analogues.     

Direct cord implantation in brachial plexus avulsions: revised technique using a single stage combined anterior (first) posterior (second) approach and end-to-side side-to-side grafting neurorrhaphy

Background

The superiority of a single stage combined anterior (first) posterior (second) approach and end-to-side side-to-side grafting neurorrhaphy in direct cord implantation was investigated as to providing adequate exposure to both the cervical cord and the brachial plexus, as to causing less tissue damage and as to being more extensible than current surgical approaches.

Methods

The front and back of the neck, the front and back of the chest up to the midline and the whole affected upper limb were sterilized while the patient was in the lateral position; the patient was next turned into the supine position, the plexus explored anteriorly and the grafts were placed; the patient was then turned again into the lateral position, and a posterior cervical laminectomy was done. The grafts were retrieved posteriorly and side grafted to the anterior cord. Using this approach, 5 patients suffering from complete traumatic brachial plexus palsy, 4 adults and 1 obstetric case were operated upon and followed up for 2 years. 2 were C5,6 ruptures and C7,8T1 avulsions. 3 were C5,6,7,8T1 avulsions. C5,6 ruptures were grafted and all avulsions were cord implanted.

Results

Surgery in complete avulsions led to Grade 4 improvement in shoulder abduction/flexion and elbow flexion. Cocontractions occurred between the lateral deltoid and biceps on active shoulder abduction. No cocontractions occurred after surgery in C5,6 ruptures and C7,8T1 avulsions, muscle power improvement extended into the forearm and hand; pain disappeared.

Limitations include

spontaneous recovery despite MRI appearance of avulsions, fallacies in determining intraoperative avulsions (wrong diagnosis, wrong level); small sample size; no controls rule out superiority of this technique versus other direct cord reimplantation techniques or other neurotization procedures; intra- and interobserver variability in testing muscle power and cocontractions.

Conclusion

Through providing proper exposure to the brachial plexus and to the cervical cord, the single stage combined anterior (first) and posterior (second) approach might stimulate brachial plexus surgeons to go more for direct cord implantation. In this study, it allowed for placing side grafts along an extensive donor recipient area by end-to-side, side-to-side grafting neurorrhaphy and thus improved results.

Level of evidence

Level IV, prospective case series.     

Intraclonal variation in RNA viruses: generation, maintenance and consequences

This paper explores the evolutionary implications of the enormous variability that characterizes populations of RNA viruses and retroviruses. It begins by examining the magnitude of genetic variation in both natural and experimental populations. In natural populations, differences arise even within individual infected patients, with the per-site nucleotide diversity at this level ranging from < 1% to 6%. In laboratory populations, two viruses sampled from the same clone differed by ∼0.7% in their fitness. Three different mechanisms that may be important in maintaining viral genetic variability were tested: (1) Fisher's fundamental theorem, to compare the observed rate of fitness change with the extent of fitness-related variation within the same experimental populations; (2) magnitude of genomic mutation rate, to assess whether it correlated with fitness-related variation, as predicted by the mutation-selection balance hypothesis; (3) frequency-dependent selection, which affords rare genotypes an advantage. The paper concludes with a discussion of two evolutionary consequences of variability: the fixation of deleterious mutations by drift in small populations and the role of clonal interference in large ones.  © 2003 The Linnean Society of London. Biological Journal of the Linnean Society , 2003, 79 , 17–26.     

Complex asparagine-linked oligosaccharides are required for morphogenic events during post-implantation development.

Complex asparagine (N)-linked oligosaccharides appear late in phylogeny and are highly regulated in vertebrates. Variations in these structures are found on the majority of cell-surface and secreted proteins. Complex N-linked oligosaccharide biosynthesis is initiated in the Golgi apparatus by the action of Mgat-1-encoded UDP-N-acetylglucosamine:alpha-3-D- mannoside beta-1,2-N-acetylglucosaminyltransferase I (GlcNAc-TI). To determine if these structures govern ontogenic processes in mammals, mouse embryos were generated that lacked a functional Mgat-1 gene. Inactivation of both Mgat-1 alleles produced deficiencies in GlcNAc-TI activity and complex N-linked oligosaccharides. Embryonic lethality occurred by day 10.5, thus establishing that complex N-linked oligosaccharides are required during post-implantation development. Remarkably, embryonic development proceeded into day 9 with the differentiation of multiple cell types. Complex N-linked oligosaccharides are important for morphogenic processes as neural tube formation, vascularization and the determination of left-right body plan asymmetry were impaired in the absence of a functional Mgat-1 gene.