Pharmacophore-based discovery of triaryl-substituted imidazole as new telomeric G-quadruplex ligand

Discovery of potent and selective ligands for telomeric G-quadruplex DNA is a challenging work. Through a combination approach of pharmacophore model construction, model validation, database virtual screening, chemical synthesis and interaction evaluation, we discovered and confirmed triaryl-substituted imidazole TSIZ01 to be a new telomeric G-quadruplex ligand with potent binding and stabilizing activity to G-quadruplex DNA, as well as a 8.7-fold selectivity towards telomeric G-quadruplex DNA over duplex DNA.     

Abscisic acid plays an important role in the regulation of strawberry fruit ripening

The plant hormone abscisic acid (ABA) has been suggested to play a role in fruit development, but supporting genetic evidence has been lacking. Here, we report that ABA promotes strawberry (Fragaria ananassa) fruit ripening. Using a newly established Tobacco rattle virus-induced gene silencing technique in strawberry fruit, the expression of a 9-cis-epoxycarotenoid dioxygenase gene (FaNCED1), which is key to ABA biosynthesis, was down-regulated, resulting in a significant decrease in ABA levels and uncolored fruits. Interestingly, a similar uncolored phenotype was observed in the transgenic RNA interference (RNAi) fruits, in which the expression of a putative ABA receptor gene encoding the magnesium chelatase H subunit (FaCHLH/ABAR) was down-regulated by virus-induced gene silencing. More importantly, the uncolored phenotype of the FaNCED1-down-regulated RNAi fruits could be rescued by exogenous ABA, but the ABA treatment could not reverse the uncolored phenotype of the FaCHLH/ABAR-down-regulated RNAi fruits. We observed that down-regulation of the FaCHLH/ABAR gene in the RNAi fruit altered both ABA levels and sugar content as well as a set of ABA- and/or sugar-responsive genes. Additionally, we showed that exogenous sugars, particularly sucrose, can significantly promote ripening while stimulating ABA accumulation. These data provide evidence that ABA is a signal molecule that promotes strawberry ripening and that the putative ABA receptor, FaCHLH/ABAR, is a positive regulator of ripening in response to ABA.     

Metabolic profiling in validation of plasma biomarkers for green tea polyphenols

Green tea polyphenols (GTP) effectively protect against chronic diseases in various animal models but human studies have been inconclusive. GTP components and metabolites in body fluids have been suggested as potential biomarkers, but validation of these biomarkers has rarely been done in human populations. A randomized, double-blinded, and placebo-controlled phase IIa chemoprevention study with GTP was conducted in 120 human subjects for 3 months. To validate GTP biomarker profiles, plasma samples were collected at baseline, 1-month, and 3-month and were analyzed by HPLC-Coularray electrochemical detection (ECD) for specific GTP components as well as for non-targeted metabolites. The levels of 2 GTP components, epigallocatechin-3-gallate (EGCG) and epicatechin-3-gallate (ECG), were homogenous at baseline (p > 0.45) but were significantly elevated (p < 0.01) by GTP treatment. Metabolic profiling identified 106 metabolites, and 56 of them were chosen to construct discriminant functions (DFs) based on the data at 3 months. The DFs clearly separated the placebo, 500 mg GTP, and 1000 mg GTP groups with an accuracy rate of 97.3%. When the DFs were applied to the combined baseline and 1-month data, the accuracy rate was 62.9% in classifying subjects into the 3 intervention groups. DFs derived from 1-month data showed similar results. Overall, this study validated plasma EGCG and ECG as reliable biomarkers for GTP consumption, and found metabolic profiles effective in discriminating different GTP dosages.     

Age-related changes of structures in cerebellar cortex of cat

We studied the structures of the cerebellar cortex of young adult and old cats for age-related changes, which were statistically analysed. Nissl staining was used to visualize the cortical neurons. The immunohistochemical method was used to display glial fibrillary acidic protein (GFAP)-immunoreactive (IR) astrocytes and neurofilament-immunoreactive (NF-IR) neurons. Under the microscope, the thickness of the cerebellar cortex was measured; and the density of neurons in all the layers as well as that of GFAP-IR cells in the granular layer was analysed. Compared with young adult cats, the thickness of the molecular layer and total cerebellar cortex was significantly decreased in old cats, and that of the granular layer increased. The density of neurons in each layer was significantly lower in old cats than in young adult ones. Astrocytes in old cats were significantly denser than in young adult ones, and accompanied by evident hypertrophy of the cell bodies and enhanced immunoreaction of GFAP substance. Purkinje cells (PCs) in old cats showed much fewer NF-IR dendrites than those in young adults. The above findings indicate a loss of neurons and decrease in the number of dendrites of the PCs in the aged cerebellar cortex, which might underlie the functional decline of afferent efficacy and information integration in the senescent cerebellum. An age-dependent enhancement of activity of the astrocytes may exert a protective effect on neurons in the aged cerebellum     

Small molecules blocking the entry of severe acute respiratory syndrome coronavirus into host cells

Severe acute respiratory syndrome coronavirus (SARS-CoV) is the pathogen of SARS, which caused a global panic in 2003. We describe here the screening of Chinese herbal medicine-based, novel small molecules that bind avidly with the surface spike protein of SARS-CoV and thus can interfere with the entry of the virus to its host cells. We achieved this by using a two-step screening method consisting of frontal affinity chromatography-mass spectrometry coupled with a viral infection assay based on a human immunodeficiency virus (HIV)-luc/SARS pseudotyped virus. Two small molecules, tetra-O-galloyl-beta-D-glucose (TGG) and luteolin, were identified, whose anti-SARS-CoV activities were confirmed by using a wild-type SARS-CoV infection system. TGG exhibits prominent anti-SARS-CoV activity with a 50% effective concentration of 4.5 microM and a selective index of 240.0. The two-step screening method described here yielded several small molecules that can be used for developing new classes of anti-SARS-CoV drugs and is potentially useful for the high-throughput screening of drugs inhibiting the entry of HIV, hepatitis C virus, and other insidious viruses into their host cells.     

Activation of the Imprinted Dlk1-Dio3 Region Correlates with Pluripotency Levels of Mouse Stem Cells

Low reprogramming efficiency and reduced pluripotency have been the two major obstacles in induced pluripotent stem (iPS) cell research. An effective and quick method to assess the pluripotency levels of iPS cells at early stages would significantly increase the success rate of iPS cell generation and promote its applications. We have identified a conserved imprinted region of the mouse genome, the Dlk1-Dio3 region, which was activated in fully pluripotent mouse stem cells but repressed in partially pluripotent cells. The degree of activation of this region was positively correlated with the pluripotency levels of stem cells. A mammalian conserved cluster of microRNAs encoded by this region exhibited significant expression differences between full and partial pluripotent stem cells. Several microRNAs from this cluster potentially target components of the polycomb repressive complex 2 (PRC2) and may form a feedback regulatory loop resulting in the expression of all genes and non-coding RNAs encoded by this region in full pluripotent stem cells. No other genomic regions were found to exhibit such clear expression changes between cell lines with different pluripotency levels; therefore, the Dlk1-Dio3 region may serve as a marker to identify fully pluripotent iPS or embryonic stem cells from partial pluripotent cells. These findings also provide a step forward toward understanding the operating mechanisms during reprogramming to produce iPS cells and can potentially promote the application of iPS cells in regenerative medicine and cancer therapy.     

Colocalization of amanitin and a candidate toxin-processing prolyl oligopeptidase in Amanita basidiocarps

Fungi in the basidiomycetous genus Amanita owe their high mammalian toxicity to the bicyclic octapeptide amatoxins such as α-amanitin. Amatoxins and the related phallotoxins (such as the heptapeptide phalloidin) are encoded by members of the "MSDIN" gene family and are synthesized on ribosomes as short (34- to 35-amino-acid) proproteins. Antiamanitin antibodies and confocal microscopy were used to determine the cellular and subcellular localizations of amanitin accumulation in basidiocarps (mushrooms) of the Eastern North American destroying angel (Amanita bisporigera). Consistent with previous studies, amanitin is present throughout the basidiocarp (stipe, pileus, lamellae, trama, and universal veil), but it is present in only a subset of cells within these tissues. Restriction of amanitin to certain cells is especially marked in the hymenium. Several lines of evidence implicate a specific prolyl oligopeptidase, A. bisporigera POPB (AbPOPB), in the initial processing of the amanitin and phallotoxin proproteins. The gene for AbPOPB is restricted taxonomically to the amatoxin-producing species of Amanita and is clustered in the genome with at least one expressed member of the MSDIN gene family. Immunologically, amanitin and AbPOPB show a high degree of colocalization, indicating that toxin biosynthesis and accumulation occur in the same cells and possibly in the same subcellular compartments.