Ultrastructure and molecular phylogeny of thaumatomonads (Cercozoa) with emphasis on Thaumatomastix salina from Oslofjorden, Norway

A culture of Thaumatomastix was isolated from a sediment sample collected in Oslofjorden and established as a monospecific strain (UIO286). Based on this culture, light and transmission electron microscopy and phylogenetic analyses were carried out. Thaumatomastix species are confined within the order Thaumatomonadida of the class Imbricatea and phylum Cercozoa. They are heterotrophic and their cell bodies are covered with silica scales. Observations of thin sections as well as whole mounts indicate that the morphology and ultrastructure of UIO286 is identical to T. salina, which was initially described from salt pools in Denmark. Detailed examination revealed some new features such as the presence of pseudopodia and silica deposition vesicles producing spine scales. The phylogeny presented here includes ribosomal DNA sequences from both imbricatean cultures and environmental samples. The 18S rDNA phylogenetic tree suggests that (i) Thaumatomastix is paraphyletic within the Thaumatomonadida clade, (ii) there is no close affinity between T. salina and other cultured and sequenced strains, but it is closely related to a sequence obtained from environmental DNA; we propose the present strain to serve as a reference culture of Thaumatomastix species and T. salina. Further, we discuss the distribution, habitats, and evolution of scale formation among euglyphids and thaumatomonads.     

Glutamine as a precursor for transmitter glutamate, aspartate and GABA in the cerebellum: a role for phosphate-activated glutaminase

Phosphate-activated glutaminase is present at high levels in the cerebellar mossy fiber terminals. The role of this enzyme for the production of glutamate from glutamine in the parallel-fiber terminals is unclear. In order to address this, we used light miroscopic immunoperoxidase and electron microscopic immunogold methods to study the localization of glutamate in rat cerbellar slices incubated with physiological K+ (3 mmol/L) and depolarizing K+ (40 mmol/L) concentrations, and during depolarizing conditions with the addition of glutamine and the glutaminase inhibitor 6-diazo-5-oxo-l-norleucine. During K+-induced depolarization glutamate labeling was redistributed from parallel-fiber terminals to glial cells. The nerve terminal content of glutamate was sustained when the slices were supplied with glutamine, which also reduced the accumulation of glutamate in glia. In spite of glutamine supplementation, the depolarized slices treated with 6-diazo-5-oxo-l-norleucine showed depletion of glutamate from parallel-fiber terminals and accumulation in glial cells. We conclude that cerebellar parallel-fiber terminals contain a glutaminase activity enabling them to synthesize glutamate from glutamine. Our results confirm that this is also true for the mossy fiber terminals. In addition, we show that, like for glutamate, the levels of aspartate in parallel-fiber terminals and GABA in Golgi fiber terminals can be maintained during depolarization if glutamine is present. This process is dependent on the activity of a glutaminase, as it can be inhibited by 6-diazo-5-oxo-l-norleucine, suggesting that the glutaminase reaction is important for glutamine to act as a precursor also for aspartate and GABA. The low levels of the kidney type of glutaminase that previously has been shown to be present in the parallel and Golgi fiber terminals could be sufficient to produce the transmitter amino acids. Alternatively, the amino acids could be produced from the liver type of glutaminase, which is not yet localized on the cellular level, or from an unknown glutminase.     

The formin mDia2 stabilizes microtubules independently of its actin nucleation activity

A critical microtubule (MT) polarization event in cell migration is the Rho/mDia-dependent stabilization of a subset of MTs oriented toward the direction of migration. Although mDia nucleates actin filaments, it is unclear whether this or a separate activity of mDia underlies MT stabilization. We generated two actin mutants (K853A and I704A) in a constitutively active version of mDia2 containing formin homology domains 1 and 2 (FH1FH2) and found that they still induced stable MTs and bound to the MT TIP proteins EB1 and APC, which have also been implicated in MT stabilization. A dimerization-impaired mutant of mDia2 (W630A) also generated stable MTs in cells. We examined whether FH1FH2mDia2 had direct activity on MTs in vitro and found that it bound directly to MTs, stabilized MTs against cold- and dilution-induced disassembly, and reduced the rates of growth and shortening during MT assembly and disassembly, respectively. These results indicate that mDia2 has a novel MT stabilization activity that is separate from its actin nucleation activity.     

森林土壤氮素转换及其对氮沉降的响应

近几十年人类活动向大气中排放的含氮化合物激增 ,并引起大气氮沉降也成比例增加。目前 ,氮沉降的增加使一些森林生态系统结构和功能发生改变 ,甚至衰退。近 2 0 a欧洲和北美有关氮沉降及其对森林生态系统的影响方面的研究较多 ,而我国少有涉及。森林土壤氮素转换是森林生态系统氮素循环的一个重要的组成部分 ,而矿化、硝化和反硝化作用是其核心过程 ,氮沉降作为驱动因子势必改变森林土壤氮素转换速度、方向和通量。根据国外近 2 0 a有关研究 ,首先介绍了森林土壤氮素转换过程和强度 ,论述森林土壤氮素在生态系统氮素循环中的作用 ,然后在此基础上 ,介绍了氮沉降对森林土壤氮素循环的研究途径 ,探讨了氮沉降对森林土壤氮素矿化、硝化和反硝化作用的影响及其机理     

南亚热带森林土壤有效氮含量及其对模拟氮沉降增加的初期响应

研究了南亚热带主要森林类型 (马尾松林、混交林和季风常绿阔叶林 )土壤有效氮含量对模拟氮沉降的初期响应。结果显示 :(1)马尾松林、混交林和阔叶林 0～ 10 cm和 10～ 2 0 cm两个土层有效氮 (铵态氮 硝态氮 )含量总平均分别为 6 .2 4、6 .2 2和14 .77m g/kg,其中铵态氮占 4 5 .3%、4 8.7%和 14 .5 %。 (2 )外加氮处理使 3个森林两个土层的有效氮含量都在增加 ,但其影响程度取决于土层、氮处理水平、氮处理时间和森林类型。总体而言 ,0～ 10 cm土层略比 10～ 2 0 cm土层敏感 ;氮处理水平越高土壤有效氮增加越多 ;外加氮处理时间越长 ,处理样方与对照样方的差距越大 ;阔叶林的响应稍落后于马尾松林和混交林     

Antimicrobial peptides with stability toward tryptic degradation

The inherent instability of peptides toward metabolic degradation is an obstacle on the way toward bringing potential peptide drugs onto the market. Truncation can be one way to increase the proteolytic stability of peptides, and in the present study the susceptibility against trypsin, which is one of the major proteolytic enzymes in the gastrointestinal tract, was investigated for several short and diverse libraries of promising cationic antimicrobial tripeptides. Quite surprisingly, trypsin was able to cleave very small cationic antimicrobial peptides at a substantial rate. Isothermal titration calorimetry studies revealed stoichiometric interactions between selected peptides and trypsin, with dissociation constants ranging from 1 to 20 microM. Introduction of hydrophobic C-terminal amide modifications and likewise bulky synthetic side chains on the central amino acid offered an effective way to increased half-life in our assays. Analysis of the degradation products revealed that the location of cleavage changed when different end-capping strategies were employed to increase the stability and the antimicrobial potency. This suggests that trypsin prefers a bulky hydrophobic element in S1' in addition to a positively charged side chain in S1 and that this binding dictates the mode of cleavage for these substrates. Molecular modeling studies supported this hypothesis, and it is shown that small alterations of the tripeptide result in two very different modes of trypsin binding and degradation. The data presented allows for the design of stable cationic antibacterial peptides and/or peptidomimetics based on several novel design principles.     

Export from Pericentriolar Endocytic Recycling Compartment to Cell Surface Depends on Stable, Detyrosinated (Glu) Microtubules and Kinesin

A significant fraction of internalized transferrin (Tf) concentrates in the endocytic recycling compartment (ERC), which is near the microtubule-organizing center in many cell types. Tf then recycles back to the cell surface. The mechanisms controlling the localization, morphology, and function of the ERC are not fully understood. We examined the relationship of Tf trafficking with microtubules (MTs), specifically the subset of stable, detyrosinated Glu MTs. We found some correlation between the level of stable Glu MTs and the distribution of the ERC; in cells with low levels of Glu MTs concentrated near to the centriole, the ERC was often tightly clustered, whereas in cells with higher levels of Glu MTs throughout the cell, the ERC was more dispersed. The clustered ERC in Chinese hamster ovary cells became dispersed when the level of Glu MTs was increased with taxol treatment. Furthermore, in a temperature-sensitive Chinese hamster ovary cell line (B104-5), the cells had more Glu MTs when the ERC became dispersed at elevated temperature. Microinjecting purified anti-Glu tubulin antibody into B104-5 cells at elevated temperature induced the redistribution of the ERC to a tight cluster. Microinjection of anti-Glu tubulin antibody slowed recycling of Tf to the cell surface without affecting Tf internalization or delivery to the ERC. Similar inhibition of Tf recycling was caused by microinjecting anti-kinesin antibody. These results suggest that stable Glu MTs and kinesin play a role in the organization of the ERC and in facilitating movement of vesicles from the ERC to the cell surface.     

Improved nitrogen removal by application of new nitrogen-cycle bacteria

In order to meet increasingly stringentEuropean discharge standards, new applicationsand control strategies for the sustainableremoval of ammonia from wastewater have to beimplemented. In this paper we discuss anitrogen removal system based on the processesof partial nitrification and anoxic ammoniaoxidation (anammox). The anammox process offersgreat opportunities to remove ammonia in fullyautotrophic systems with biomass retention. Noorganic carbon is needed in such nitrogenremoval system, since ammonia is used aselectron donor for nitrite reduction. Thenitrite can be produced from ammonia inoxygen-limited biofilm systems or in continuousprocesses without biomass retention. Forsuccessful implementation of the combinedprocesses, accurate biosensors for measuringammonia and nitrite concentrations, insight inthe complex microbial communities involved, andnew control strategies have to be developed andevaluated.     

Specific cytosol binding of diethylstilbestrol in rat skeletal muscle

Rat skeletal muscle cytosol proteins bound ³H-diethylstilbestrol (³H-DES). More than 90% of this binding was high capacity and low affinity. Serum albumin accounted for roughly 50–60% of the binding, as evidenced by its precipitation with anti-rat albumin IgG. About half of the binding was distinguishable from albumin and other serum proteins by its precipitation in 40% saturated ammonium sulfate. This material sedimented at 4–5S in high-salt sucrose gradients, and resolved into two components (8S and 4–5S) in low-salt. Following incubation at 23–27°C for one hour, 2% of the bound ³H-DES in whole cytosol (approximately 2 fmole/mg cytosol protein) was retained by DNA-cellulose, and was eluted with 0.6 M KCl. This small fraction of the total binding was inhibited by estrogens and DES analogues: estradiol-17β, DES, dienestrol, and hexestrol were strong inhibitors; isodienestrol, dimethylstilbestrol, estradiol-17α, estrone, tamoxifen, MER-25, CI-628, and nafoxidine were weak inhibitors; dihydrotestosterone, testosterone, and prednisone did not compete. These observations indicate that specific estrogen-binding sites exist in rat skeletal muscle.