Estrogen receptor beta selective ligands: discovery and SAR of novel heterocyclic ligands

A series of ligands with varying heterocyclic cores and substituents that display a range of selectivity’s (up to >100x) for ER-β over ER- are reported.     

Morphological Changes of <Emphasis Type="Italic">Pseudomonas pseudoalcaligenes</Emphasis> in Response to Temperature Selection

Adaptation to novel environments usually entails morphological changes. The cell morphology of six experimental populations of Pseudomonas pseudoalcaligenes and their common ancestor were examined with scanning electron microscopy (SEM). The six experimental populations were propagated under different temperatures for 10 months: three of them cultured at constant normal temperature (35°C) forming the control group, and the other three cultured at incremental higher temperatures (from 41° to 47°C) as the HT group. SEM showed the deformed and elongated cells in the 6-h cultures of both ancestral and control populations at 45°C, indicating that 45°C is stressful for the ancestral and the control populations. In contrast, the HT populations retained normal cell shape in the 6-h cultures at both 35°C and 45°C. The mean cell volumes of control and HT populations increased 29% and 34%, respectively, relative to the ancestor at their respective thermal regimens, suggestion that the culturing conditions might favor larger cells. Received: 27 March 2002 / Accepted: 30 April 2002     

Development and evaluation of a broadly reactive reverse transcription recombinase polymerase amplification assay for rapid detection of murine norovirus

Background

Murine norovirus (MNV) is recognized as the most prevalent viral pathogen in captive mouse colonies. The rapid detection assay for MNV would be a useful tool for monitoring and preventing MNV infection. A recombinase polymerase amplification (RPA) assay was established in this study to provide a solution for rapid and sensitive detection of MNV.

Results

The detection limit of the RT-RPA assay for the detection of MNV was 1?×?10² copies of RNA molecules per reaction. The assay was specific since there was no cross-reaction with other common murine viruses. In addition, the broad reactivity of the RT-RPA assay was validated using the synthesized template carrying seven point mutations among several MNV strains. The MNV RT-RPA assay could detect as few as 1?×?10² copies of the mutant per reaction, suggesting the assay could be broadly reactive against a large diversity of MNV strains. Forty eight clinical samples including 16 gastric tissue specimens, 16 cecal tissue specimens and 16 fecal specimens were tested for the validation of the new developed RT-RPA assay. The detection results of RT-RPA and RT-qPCR for clinical samples were very similar, except that a gastric tissue sample which was positive by RT-qPCR, with a RNA titer of 27 copies, was negative by RT-RPA.

Conclusions

A broadly reactive RT-RPA assay was successfully established for MNV detection.

     

Selenium nanoparticles-loaded chitosan/citrate complex and its protection against oxidative stress in <Emphasis Type="SmallCaps">d</Emphasis>-galactose-induced aging mice

Background

Selenium (Se) is an indispensable trace element required for animals and humans, and extra Se-supplement is necessary, especially for those having Se deficiency. Recently, selenium nanoparticles (SeNPs), as a special form of Se supplement, have attracted worldwide attention due to their distinguished properties and excellent bioactivities. In this present study, an eco-friendly and economic way to prepare stable SeNPs was introduced. SeNPs were synthesized in the presence of chitosan (CTS) and then embedded into chitosan/citrate gel, generating selenium nanoparticles-loaded chitosan/citrate complex (SeNPs-C/C). Additionally, the clinical potential of SeNPs-C/C was evaluated by using d-galactose (d-gal)-induced aging mice model.

Results

SeNPs in high uniform with an average diameter of around 50 nm were synthesized in the presence of chitosan, and reversible ionic gelation between chitosan and citrate was utilized to load SeNPs. Subsphaeroidal SeNPs-C/C microspheres of 1–30 μm were obtained by spay-drying. Single SeNPs were physically separated and embedded inside SeNPs-C/C microparticles, with excellent stability and acceptable release. Acute fetal test showed SeNPs-C/C was safer than selenite, with a median lethal dose (LD₅₀) of approximately 4-fold to 11-fold of that of selenite. Oral administration of SeNPs-C/C remarkably retarded the oxidative stress of d-gal in Kunming mice by enhancing the activity of antioxidase, as evidenced by its significant protection of the growth, liver, Se retention and antioxidant bio-markers of mice against d-gal.

Conclusions

The design of SeNPs-C/C opens a new path for oral delivery of SeNPs with excellent stability, energy-conservation and environment-friendliness. SeNPs-C/C, as a novel supplement of Se, could be further developed to defend the aging process induced by d-gal.

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Morphological changes of Pseudomonas pseudoalcaligenes in response to temperature selection

Adaptation to novel environments usually entails morphological changes. The cell morphology of six experimental populations of Pseudomonas pseudoalcaligenes and their common ancestor were examined with scanning electron microscopy (SEM). The six experimental populations were propagated under different temperatures for 10 months: three of them cultured at constant normal temperature (35 degrees C) forming the control group, and the other three cultured at incremental higher temperatures (from 41 degrees to 47 degrees C) as the HT group. SEM showed the deformed and elongated cells in the 6-h cultures of both ancestral and control populations at 45 degrees C, indicating that 45 degrees C is stressful for the ancestral and the control populations. In contrast, the HT populations retained normal cell shape in the 6-h cultures at both 35 degrees C and 45 degrees C. The mean cell volumes of control and HT populations increased 29% and 34%, respectively, relative to the ancestor at their respective thermal regimens, suggestion that the culturing conditions might favor larger cells.     

R2R3 MYB‐dependent auxin signalling regulates trichome formation,and increased trichome density confers spider mite tolerance on tomato

Unicellular and multicellular tomato trichomes function as mechanical and chemical barriers against herbivores. Auxin treatment increased the formation of II, V and VI type trichomes in tomato leaves. The auxin response factor gene SlARF4, which was highly expressed in II, V and VI type trichomes, positively regulated the auxin‐induced formation of II, V and VI type trichomes in the tomato leaves. SlARF4 overexpression plants with high densities of these trichomes exhibited tolerance to spider mites. Two R2R3 MYB genes, SlTHM1 and SlMYB52, were directly targeted and inhibited by SlARF4. SlTHM1 was specifically expressed in II and VI type trichomes and negatively regulated the auxin‐induced formation of II and VI type trichomes in the tomato leaves. SlTHM1 down‐regulation plants with high densities of II and VI type trichomes also showed tolerance to spider mites. SlMYB52 was specifically expressed in V type trichomes and negatively regulated the auxin‐induced formation of V type trichome in the tomato leaves. The regulation of SlARF4 on the formation of II, V and VI type trichomes depended on SlTHM1 and SlMYB52, which directly targeted cyclin gene SlCycB2 and increased its expression. In conclusion, our data indicates that the R2R3 MYB‐dependent auxin signalling pathway regulates the formation of II, V and VI type trichomes in tomato leaves. Our study provides an effective method for improving the tolerance of tomato to spider mites.     

Structure of the autocatalytic cysteine protease domain of potyvirus helper-component proteinase

The helper-component proteinase (HC-Pro) of potyvirus is involved in polyprotein processing, aphid transmission, and suppression of antiviral RNA silencing. There is no high resolution structure reported for any part of HC-Pro, hindering mechanistic understanding of its multiple functions. We have determined the crystal structure of the cysteine protease domain of HC-Pro from turnip mosaic virus at 2.0 Å resolution. As a protease, HC-Pro only cleaves a Gly-Gly dipeptide at its own C terminus. The structure represents a postcleavage state in which the cleaved C terminus remains tightly bound at the active site cleft to prevent trans activity. The structure adopts a compact α/β-fold, which differs from papain-like cysteine proteases and shows weak similarity to nsP2 protease from Venezuelan equine encephalitis alphavirus. Nevertheless, the catalytic cysteine and histidine residues constitute an active site that is highly similar to these in papain-like and nsP2 proteases. HC-Pro recognizes a consensus sequence YXVGG around the cleavage site between the two glycine residues. The structure delineates the sequence specificity at sites P1–P4. Structural modeling and covariation analysis across the Potyviridae family suggest a tryptophan residue accounting for the glycine specificity at site P1′. Moreover, a surface of the protease domain is conserved in potyvirus but not in other genera of the Potyviridae family, likely due to extra functional constrain. The structure provides insight into the catalysis mechanism, cis-acting mode, cleavage site specificity, and other functions of the HC-Pro protease domain.