The monofunctional glycosyltransferase of Escherichia coli localizes to the cell division site and interacts with penicillin-binding protein 3, FtsW, and FtsN

The monofunctional peptidoglycan glycosyltransferase (MtgA) catalyzes glycan chain elongation of the bacterial cell wall. Here we show that MtgA localizes at the division site of Escherichia coli cells that are deficient in PBP1b and produce a thermosensitive PBP1a and is able to interact with three constituents of the divisome, PBP3, FtsW, and FtsN, suggesting that MtgA may play a role in peptidoglycan assembly during the cell cycle in collaboration with other proteins.     

Patterns of sequence variability and divergence at the diminutive gene region of Drosophila melanogaster: complex patterns suggest an ancestral selective sweep

To identify putatively swept regions of the Drosophila melanogaster genome, we performed a microsatellite screen spanning a 260-kb region of the X chromosome in populations from Zimbabwe, Ecuador, the United States, and China. Among the regions identified by this screen as showing a complex pattern of reduced heterozygosity and a skewed frequency spectrum was the gene diminutive (dm). To investigate the microsatellite findings, nucleotide sequence polymorphism data were generated in populations from both China and Zimbabwe spanning a 25-kb region and encompassing dm. Analysis of the sequence data reveals strongly reduced nucleotide variation across the entire gene region in both the non-African and the African populations, an extended haplotype pattern, and structured linkage disequilibrium, as well as a rejection of neutrality in favor of selection using a composite likelihood-ratio test. Additionally, unusual patterns of synonymous site evolution were observed at the second exon of this locus. On the basis of simulation studies as well as recently proposed methods for distinguishing between selection and nonequilibrium demography, we find that this "footprint" is best explained by a selective sweep in the ancestral population, the signal of which has been somewhat blurred via founder effects in the non-African samples.     

Reorganization of microtubules in endosperm cells and cell fragments of the higher plant Haemanthus in vivo

The reorganization of the microtubular meshwork was studied in intact Haemanthus endosperm cells and cell fragments (cytoplasts). This higher plant tissue is devoid of a known microtubule organizating organelle. Observations on living cells were correlated with microtubule arrangements visualized with the immunogold method. In small fragments, reorganization did not proceed. In medium and large sized fragments, microtubular converging centers formed first. Then these converging centers reorganized into either closed bushy microtubular spiral or chromosome-free cytoplasmic spindles/phragmoplasts. Therefore, the final shape of organized microtubular structures, including spindle shaped, was determined by the initial size of the cell fragments and could be achieved without chromosomes or centrioles. Converging centers elongate due to the formation of additional structures resembling microtubular fir trees. These structures were observed at the pole of the microtubular converging center in anucleate fragments, accessory phragmoplasts in nucleated cells, and in the polar region of the mitotic spindle during anaphase. Therefore, during anaphase pronounced assembly of new microtubules occurs at the polar region of acentriolar spindles. Moreover, statistical analysis demonstrated that during the first two-thirds of anaphase, when chromosomes move with an approximately constant speed, kinetochore fibers shorten, while the length of the kinetochore fiber complex remains constant due to the simultaneous elongation of their integral parts (microtubular fir trees). The half-spindle shortens only during the last one-third of anaphase. These data contradict the presently prevailing view that chromosome-to-pole movements in acentriolar spindles of higher plants are concurrent with the shortening of the half-spindle, the self-reorganizing property of higher plant microtubules (tubulin) in vivo. It may be specific for cells without centrosomes and may be superimposed also on other microtubule-related processes.     

Kinetic analysis of the mass transport limited interaction between the tyrosine kinase lck SH2 domain and a phosphorylated peptide studied by a new cuvette-based surface plasmon resonance instrument

We explored the use of a newly developed cuvette-based surface plasmon resonance (SPR) instrument (IBIS) to study peptide-protein interactions. We studied the interaction between the SH2 domain of lck and a phosphotyrosine peptide EPQY*EEIPIYL which was immobilized on a sensor chip. No indications for mass transport limitation (MTL) were observed when standard kinetic approaches were used. However, addition of competing peptide during dissociation revealed a high extent of rebinding. A dissociation rate constant (k(d)) of 0.6+/-0.1 s(-1) was obtained in the presence of large amounts of peptide. A simple bimolecular binding model, applying second-order kinetics for the cuvette system, could not adequately describe the data. Fits were improved upon including a step in the model which describes diffusion of the SH2 domain from the bulk to the sensor, especially for a surface with high binding capacity. From experiments in glycerol-containing buffers, it appeared that the diffusion rate decreased with higher viscosity. It is demonstrated that MTL during association and dissociation can be described by the same diffusion rate. A binding constant (K(D)) of 5.9+/-0.8 nM was obtained from the SPR equilibrium signals by fitting to a Langmuir binding isotherm, with correction for loss of free analyte due to binding. An association rate constant k(a) of 1.1(+/-0.2)x10(8) M(-1) x s(-1) was obtained from k(d)/K(D). The values for k(a) and k(d) obtained in this way were 2-3 orders larger than that from standard kinetic analysis, ignoring MTL. We conclude that in a cuvette the extent of MTL is comparable to that in a flow system.     

Interlaboratory Comparison of Methods To Quantify Microsclerotia of Verticillium dahliae in Soil

In a comparison of different methods for estimating Verticillium dahliae in soil, 14 soil samples were analyzed in a blinded fashion by 13 research groups in seven countries, using their preferred methods. One group analyzed only four samples. Twelve soil samples were naturally infested, and two had known numbers of microsclerotia of V. dahliae added to them. In addition, a control was included to determine whether transport had an effect on the results. Results differed considerably among the research groups. There was a 118-fold difference between the groups with the lowest and highest mean estimates. Results of the other groups were evenly distributed between these extremes. In general, methods based on plating dry soil samples gave higher numbers of V. dahliae than did plating of an aqueous soil suspension. Recovery of V. dahliae from samples with added microsclerotia varied from 0 to 59%. Most of the variability within each analysis was at the petri dish level. The results indicate the necessity to check the performance of detection assays regularly by comparing recoveries with other laboratories, using a common set of soil samples. We conclude that wet plating assays are less accurate than dry plating assays.     

Evidence for new C-terminally truncated variants of α- and β-tubulins

Cellular α-tubulin can bear various carboxy-terminal sequences: full-length tubulin arising from gene neosynthesis is tyrosinated, and two truncated variants, corresponding to detyrosinated and Δ2 α‑tubulin, result from the sequential cleavage of one or two C-terminal residues, respectively. Here, by using a novel antibody named 3EG that is highly specific to the –EEEG C-terminal sequence, we demonstrate the occurrence in neuronal tissues of a new αΔ3‑tubulin variant corresponding to α1A/B‑tubulin deleted of its last three residues (EEY). αΔ3‑tubulin has a specific distribution pattern: its quantity in the brain is similar to that of αΔ2-tubulin around birth but is much lower in adult tissue. This truncated α1A/B-tubulin variant can be generated from αΔ2-tubulin by the deglutamylases CCP1, CCP4, CCP5, and CCP6 but not by CCP2 and CCP3. Moreover, using 3EG antibody, we identify a C‑terminally truncated β-tubulin form with the same –EEEG C-terminal sequence. Using mass spectrometry, we demonstrate that β2A/B-tubulin is modified by truncation of the four C-terminal residues (EDEA). We show that this newly identified βΔ4-tubulin is ubiquitously present in cells and tissues and that its level is constant throughout the cell cycle. These new C-terminally truncated α- and β-tubulin variants, both ending with –EEEG sequence, are expected to regulate microtubule physiology. Of interest, the αΔ3-tubulin seems to be related to dynamic microtubules, resembling tyrosinated-tubulin rather than the other truncated variants, and may have critical function(s) in neuronal development.     

Biodiversity, evolution and adaptation of cultivated crops

The human diet depends on very few crops. Current diversity in these crops is the result of a long interaction between farmers and cultivated plants, and their environment. Man largely shaped crop biodiversity from the domestication period 12,000 B.P. to the development of improved varieties during the last century. We illustrate this process through a detailed analysis of the domestication and early diffusion of maize. In smallholder agricultural systems, farmers still have a major impact on crop diversity today. We review several examples of the major impact of man on current diversity. Finally, biodiversity is considered to be an asset for adaptation to current environmental changes. We describe the evolution of pearl millet in West Africa, where average rainfall has decreased over the last forty years. Diversity in cultivated varieties has certainly helped this crop to adapt to climate variation.     

Systematic re-evaluation of intraoperative motor-evoked potential suppression in scoliosis surgery

Background

Motor- (MEP) and somatosensory-evoked potentials (SSEP) are susceptible to the effects of intraoperative environmental factors.

Methods

Over a 5-year period, 250 patients with adolescent idiopathic scoliosis (AIS) who underwent corrective surgery with IOM were retrospectively analyzed for MEP suppression (MEPS).

Results

Our results show that four distinct groups of MEPS were encountered over the study period. All 12 patients did not sustain any neurological deficits postoperatively. However, comparison of groups 1 and 2 suggests that neither the duration of anesthesia nor speed of surgical or anesthetic intervention were associated with recovery to a level beyond the criteria for MEPS. For group 3, spontaneous MEPS recovery despite the lack of surgical intervention suggests that anesthetic intervention may play a role in this process. However, spontaneous MEPS recovery was also seen in group 4, suggesting that in certain circumstances, both surgical and anesthetic intervention was not required. In addition, neither the duration of time to the first surgical manoeuver nor the duration of surgical manoeuver to MEPS were related to recovery of MEPS. None of the patients had suppression of SSEPs intraoperatively.

Conclusion

This study suggests that in susceptible individuals, MEPS may rarely occur unpredictably, independent of surgical or anesthetic intervention. However, our findings favor anesthetic before surgical intervention as a proposed protocol. Early recognition of MEPS is important to prevent false positives in the course of IOM for spinal surgery.