Shoot-derived auxin is essential for early lateral root emergence in Arabidopsis seedlings

Lateral root formation is profoundly affected by auxins. Here we present data which indicate that light influences the formation of indole-3-acetic acid (IAA) in germinating Arabidopsis seedlings. IAA transported from the developing leaves to the root system is detectable as a short-lived pulse in the roots and is required for the emergence of the lateral root primordia (LRP) during early seedling development. LRP emergence is inhibited by the removal of apical tissues prior to detection of the IAA pulse in the root, but this treatment has minimal effects on LRP initiation. Our results identify the first developing true leaves as the most likely source for the IAA required for the first emergence of the LRP, as removal of cotyledons has only a minor effect on LRP emergence in contrast to removal of the leaves. A basipetal IAA concentration gradient with high levels of IAA in the root tip appears to control LRP initiation, in contrast to their emergence. A significant increase in the ability of the root system to synthesize IAA is observed 10 days after germination, and this in turn is reflected in the reduced dependence of the lateral root emergence on aerial tissue-derived auxin at this stage. We propose a model for lateral root formation during early seedling development that can be divided into two phases: (i) an LRP initiation phase dependent on a root tip-localized IAA source, and (ii) an LRP emergence phase dependent on leaf-derived IAA up to 10 days after germination.     

Structural studies of the exopolysaccharide produced by Lactobacillus rhamnosus strain GG (ATCC 53103)

The structure of the extracellular polysaccharide (EPS) from Lactobacillus rhamnosus strain GG has been investigated. In combination with component analysis, NMR spectroscopy shows that the polysaccharide is composed of hexasaccharide repeating units. Sequential information was obtained by two-dimensional (1)H,(1)H-NOESY, and (1)H,(13)C-HMBC NMR techniques. The structure of the repeating unit of the EPS from Lactobacillus rhamnosus strain GG was determined as: [carbohydrate structure: see text]     

FISH-mapping of a 100-kb terminal 22q13 deletion

Both cytogenetically visible and cryptic deletions of the terminal region of chromosome 22q are associated with a clinical phenotype including mental retardation, delay in expressive speech development, hypotonia, normal to accelerated growth and minor facial dysmorphic features. The genes responsible for the development of the phenotype have not yet been identified, but a distal localization is probable, since the cytogenetically visible and the cryptic deletions show a similar pattern of symptoms. We report a 33-year-old woman with a submicroscopic 22q13 deletion, mild mental retardation, speech delay, autistic symptoms and mild facial dysmorphic features. The deletion was mapped by FISH using cosmid probes from terminal 22q13, and the size of the deletion was estimated to be 100 kb. Three genes are affected by the deletion in this patient. ACR and RABL2B are deleted and proSAP2 is disrupted. This observation, together with recently published data, supports the notion that proSAP2 is the most important contributor to the 22q13 deletion phenotype.     

Dramatic stabilization of the native state of human carbonic anhydrase II by an engineered disulfide bond

To find a disulfide pair that could stabilize the enzyme human carbonic anhydrase II (HCA II), we grafted the disulfide bridge from the related and unusually stable carbonic anhydrase form from Neisseria gonorrhoeae (NGCA) into the human enzyme. Thus, the two Cys residues at positions 23 and 203 were engineered into a pseudo-wild-type form of HCA II (C206S), giving the mutant C206S/A23C/L203C. The disulfide bond was not formed spontaneously. The native state of the reduced form of the mutant was markedly destabilized (2.9 kcal/mol) compared to that of HCA II. Formation of a disulfide bridge was achieved by treatment by oxidized glutathione. This led to a significant stabilization of the native conformation. Compared to HCA II the unfolding midpoint for the variant was increased from 0.9 to 1.7 M guanidine HCl, corresponding to a stabilization of 3.7 kcal/mol. This makes the human enzyme almost as stable as the model protein NGCA, for which the unfolding of the native state has a midpoint at 2.1 M guanidine HCl. The stabilized protein underwent, contrary to all other investigated variants of HCA II, an apparent two-state unfolding transition, as judged from intrinsic Trp fluorescence measurements. A molten-globule intermediate is nevertheless formed but is suppressed because of the high denaturant pressure it faces upon rupture of the native state.     

Solution structure of a pentasaccharide representing the repeating unit of the O-antigen polysaccharide from Escherichia coli O142: NMR spectroscopy and molecular simulation studies

Conformational studies have been performed of a pentasaccharide derived from the O-polysaccharide from Escherichia coli O142. The polymer was selectively degraded by anhydrous hydrogen fluoride and reduced to yield an oligosaccharide model of its repeating unit, which in the branching region consists of four aminosugars. A comparison of (1)H and (13)C chemical shifts between the pentasaccharide and the polymer showed only minor differences, except where the cleavage had taken place, indicating that the oligomer is a good model of the repeating unit. Langevin dynamics and molecular dynamics simulations with explicit water molecules were carried out to sample the conformational space of the pentasaccharide. For the glycosidic linkages between the hexopyranoside residues, small but significant changes were observed between the simulation techniques. One-dimensional (1D) (1)H,(1)H double pulsed field gradient spin echo (DPFGSE) transverse rotating-frame Overhauser effect spectroscopy (T-ROESY) experiments were performed, and homonuclear cross-relaxation rates were obtained. Subsequently, a comparison of interproton distances from NMR experiment and the two simulation approaches showed that in all cases the use of explicit water in the simulations resulted in better agreement. Hydrogen-bond analysis of the trajectories from the molecular dynamics simulation revealed interresidue interactions to be important as a cluster of different hydrogen bonds and as a distinct highly populated hydrogen bond. NMR data are consistent with the presence of hydrogen bonding within the model of the repeating unit.     

Pine Forest Floor Carbon Accumulation in Response to N and PK Additions: Bomb <Superscript>14</Superscript>C Modelling and Respiration Studies

The addition of nitrogen via deposition alters the carbon balance of temperate forest ecosystems by affecting both production and decomposition rates. The effects of 20 years of nitrogen (N) and phosphorus and potassium (PK) additions were studied in a 40-year-old pine stand in northern Sweden. Carbon fluxes of the forest floor were reconstructed using a combination of data on soil ¹⁴C, tree growth, and litter decomposition. N-only additions caused an increase in needle litterfall, whereas both N and PK additions reduced long-term decomposition rates. Soil respiration measurements showed a 40% reduction in soil respiration for treated compared to control plots. The average age of forest floor carbon was 17 years. Predictions of future soil carbon storage indicate an increase of around 100% in the next 100 years for the N plots and 200% for the NPK plots. As much as 70% of the increase in soil carbon was attributed to the decreased decomposition rate, whereas only 20% was attributable to increased litter production. A reduction in decomposition was observed at a rate of N addition of 30 kg C ha^–1 y^–1, which is not an uncommon rate of N deposition in central Europe. A model based on the continuous-quality decomposition theory was applied to interpret decomposer and substrate parameters. The most likely explanations for the decreased decomposition rate were a fertilizer-induced increase in decomposer efficiency (production-to-assimilation ratio), a more rapid rate of decrease in litter quality, and a decrease in decomposer basic growth rate.     

Analysis of human alpha-thrombin by hydrophobic interaction high-performance liquid chromatography

The major active form of human thrombin, alpha-thrombin, was analyzed by hydrophobic interaction high-performance liquid chromatography (HIC-HPLC). The chromatographic system included a polymeric phenyl column and elution was performed by a gradient, 2-0M sodium chloride (5-20 min). Total analysis time was 30 min per injection. By this method, a good resolution between alpha-thrombin and the proteolytically modified thrombin forms, beta- and gamma-thrombin, was obtained. In addition, the thrombin preforms, prothrombin, prethrombin 1, and prethrombin 2, were also resolved from alpha-thrombin in the system. The results from the HIC method were compared to those obtained from non-reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By this high-resolution chromatographic method, the rapid analysis of purified alpha-thrombin is possible.     

The influence of volume of exercise on early adaptations to strength training

The purpose of this investigation was to compare the effects of single-set strength training and 3-set strength training during the early phase of adaptation in 18 untrained male subjects (age, 20-30 years). After initial testing, subjects were randomly assigned to either the 3L-1U group (n = 8), which trained 3 sets in leg exercises and 1 set in upper-body exercises, or the 1L-3U group (n = 10), which trained 1 set in leg exercises and 3 sets in upper-body exercises. Testing was conducted at the beginning and at the end of the study and consisted of 2 maximal isometric tests (knee extension and bench press) and 6 maximal dynamic tests (1 repetition maximum [1RM] tests). Subjects trained 3 days per week for 6 weeks. After warm-up, subjects performed 3 leg exercises and 4 upper-body exercises. In both groups, each set consisted of 7 repetitions (reps) with the load supposed to induce muscular failure after the seventh rep (7RM load). After 6 weeks of training, 1RM performance in all training exercises was significantly increased (10-26%, p < 0.01) in both groups. The relative increase in 1RM load in the 3 leg exercises was significantly greater in the 3L-1U group than in the 1L-3U group (21% vs. 14%, p = 0.01). However, the relative increase in 1RM load in the 3 upper-body exercises was similar in the 3L-1U group (16%) and the 1L-3U group (14%). These results show a superior adaptation to 3-set strength training, compared with 1-set strength training, in leg exercises but not in upper-body exercises during the early phase of adaptation.