Protocadherins control the modular assembly of neuronal columns in the zebrafish optic tectum

Cell–cell recognition guides the assembly of the vertebrate brain during development. δ-Protocadherins comprise a family of neural adhesion molecules that are differentially expressed and have been implicated in a range of neurodevelopmental disorders. Here we show that the expression of δ-protocadherins partitions the zebrafish optic tectum into radial columns of neurons. Using in vivo two-photon imaging of bacterial artificial chromosome transgenic zebrafish, we show that pcdh19 is expressed in discrete columns of neurons, and that these columnar modules are derived from proliferative pcdh19⁺ neuroepithelial precursors. Elimination of pcdh19 results in both a disruption of columnar organization and defects in visually guided behaviors. These results reveal a fundamental mechanism for organizing the developing nervous system: subdivision of the early neuroepithelium into precursors with distinct molecular identities guides the autonomous development of parallel neuronal units, organizing neural circuit formation and behavior.     

Viability and Isolation of Marine Bacteria by Dilution Culture: Theory, Procedures, and Initial Results

Dilution culture, a method for growing the typical small bacteria from natural aquatic assemblages, has been developed. Each of 11 experimental trials of the technique was successful. Populations are measured, diluted to a small and known number of cells, inoculated into unamended sterilized seawater, and examined three times for the presence of 10⁴ or more cells per ml over a 9-week interval. Mean viability for assemblage members is obtained from the frequency of growth, and many of the cultures produced are pure. Statistical formulations for determining viability and the frequency of pure culture production are derived. Formulations for associated errors are derived as well. Computer simulations of experiments agreed with computed values within the expected error, which verified the formulations. These led to strategies for optimizing viability determinations and pure culture production. Viabilities were usually between 2 and 60% and decreased with >5 mg of amino acids per liter as carbon. In view of difficulties in growing marine oligobacteria, these high values are noteworthy. Significant differences in population characteristics during growth, observed by high-resolution flow cytometry, suggested substantial population diversity. Growth of total populations as well as of cytometry-resolved subpopulations sometimes were truncated at levels of near 10⁴ cells per ml, showing that viable cells could escape detection. Viability is therefore defined as the ability to grow to that population; true viabilities could be even higher. Doubling times, based on whole populations as well as individual subpopulations, were in the 1-day to 1-week range. Data were examined for changes in viability with dilution suggesting cell-cell interactions, but none could be confirmed. The frequency of pure culture production can be adjusted by inoculum size if the viability is known. These apparently pure cultures produced retained the size and apparent DNA-content characteristic of the bulk of the organisms in the parent seawater. Three cultures are now available, two of which have been carried for 3 years. The method is thus seen as a useful step for improving our understanding of typical aquatic organisms.     

越南、老挝蕨类植物新记载 （3）——凤尾蕨属

通过野外调查和标本研究,报道越南、老挝两地凤尾蕨属Pteris 7个新记载的物种,它们分别是P.argyraea、四川凤尾蕨P.sichuanensis、两广凤尾蕨P.maclurei、 单叶凤尾蕨P.pseudopellucida、 有刺凤尾蕨P.setuloso-costulata、 琼南凤尾蕨P.morii、 栗轴凤尾蕨P.wangiana.     

Cutting edge: CD47 controls the in vivo proliferation and homeostasis of peripheral CD4+ CD25+ Foxp3+ regulatory T cells that express CD103

Peripheral CD103(+)Foxp3(+) regulatory T cells (Tregs) can develop both from conventional naive T cells upon cognate Ag delivery under tolerogenic conditions and from thymic-derived, expanded/differentiated natural Tregs. We here show that CD47 expression, a marker of self on hematopoietic cells, selectively regulated CD103(+)Foxp3(+) Treg homeostasis at the steady state. First, the proportion of effector/memory-like (CD44(high)CD62L(low)) CD103(+)Foxp3(+) Tregs rapidly augmented with age in CD47-deficient mice (CD47(-/-)) as compared with age-matched control littermates. Yet, the percentage of quiescent (CD44(low)CD62L(high)) CD103(-)Foxp3(+) Tregs remained stable. Second, the increased proliferation rate (BrdU incorporation) observed within the CD47(-/-)Foxp3(+) Treg subpopulation was restricted to those Tregs expressing CD103. Third, CD47(-/-) Tregs maintained a normal suppressive function in vitro and in vivo and their increased proportion in old mice led to a decline of Ag-specific T cell responses. Thus, sustained CD47 expression throughout life is critical to avoid an excessive expansion of CD103(+) Tregs that may overwhelmingly inhibit Ag-specific T cell responses.     

A New Species of Poorly Known Mayfly Genus Paegniodes Eaton (Ephemeroptera: Heptageniidae) from Vietnam

A heptageniid mayfly species Paegniodes dao sp. nov. (Ephemeroptera: Heptageniidae) is described from Vietnam. The larva is distinguished by round anterolateral margins of pronotum, triangular‐shaped labrum, rudimentary lamella and well developed fibrillae of gills 1, and light brown body that lacks distinct markings. Diagnosis, line drawings, material, and habitat data are provided.     

3H]spiroperidol binding on lymphocytes: changes in two different groups of schizophrenic patients and effect of neuroleptic treatment

[3H]spiroperidol binding to lymphocytes was measured in untreated paranoid or disorganized and treated paranoid schizophrenic patients. An increase in the Bmax was detected in untreated paranoid patients but a decrease was found in the disorganized patients. No difference was detected in the KD value. Neuroleptic treatment produced a decrease in the Bmax without affecting the KD value. Such results did not comply with the down regulation but might be explained by a change in membrane viscosity as [3H]spiroperidol binding sites on lymphocytes were coupled to phospholipid methylation.     

Effects of galactomannan as carbon source on production of α-galactosidase by Thermomyces lanuginosus: Fermentation,purification and partial characterisation

Thermophilic fungus Thermomyces lanuginosus CBS 395.62/b strain is able to grow and synthesise extracellular α-galactosidase in media containing galactomannan such as locust bean gum (LBG) or guar gum (GG). Production of extracellular α-galactosidase was enhanced from 1.2 U/mL to 4–6 U/mL meaning about 3–5 times increase by optimisation of medium composition. This enzyme was purified to homogeneity by partial precipitation with 2-propanol and different liquid chromatographical steps. The developed purification protocol yielded 22% of enzyme activity with 900 purified fold. Molecular mass of the purified α-galactosidase enzyme was estimated to be 53 kDa. Maximal catalytic activity of the enzyme was obtained in the acidic pH range between pH 4.6 and 4.8 and in the temperature range 60–66 °C. More than 95% of enzyme activity was remaining after 1-day incubation at 70 °C and on pH in the range from 4.0 to 7.0. The enzyme activity was significantly stimulated by Mg²⁺, Mn²⁺ and K⁺ ions, while considerably inhibited by the presence of Ca²⁺, Ag⁺ and Hg²⁺.     

Quantification of Cellular NEMO Content and Its Impact on NF-κB Activation by Genotoxic Stress

NF-κB essential modulator, NEMO, plays a key role in canonical NF-κB signaling induced by a variety of stimuli, including cytokines and genotoxic agents. To dissect the different biochemical and functional roles of NEMO in NF-κB signaling, various mutant forms of NEMO have been previously analyzed. However, transient or stable overexpression of wild-type NEMO can significantly inhibit NF-κB activation, thereby confounding the analysis of NEMO mutant phenotypes. What levels of NEMO overexpression lead to such an artifact and what levels are tolerated with no significant impact on NEMO function in NF-κB activation are currently unknown. Here we purified full-length recombinant human NEMO protein and used it as a standard to quantify the average number of NEMO molecules per cell in a 1.3E2 NEMO-deficient murine pre-B cell clone stably reconstituted with full-length human NEMO (C5). We determined that the C5 cell clone has an average of 4 x 10⁵ molecules of NEMO per cell. Stable reconstitution of 1.3E2 cells with different numbers of NEMO molecules per cell has demonstrated that a 10-fold range of NEMO expression (0.6–6x10⁵ molecules per cell) yields statistically equivalent NF-κB activation in response to the DNA damaging agent etoposide. Using the C5 cell line, we also quantified the number of NEMO molecules per cell in several commonly employed human cell lines. These results establish baseline numbers of endogenous NEMO per cell and highlight surprisingly normal functionality of NEMO in the DNA damage pathway over a wide range of expression levels that can provide a guideline for future NEMO reconstitution studies.