Differential expression of a novel gene <Emphasis Type="Italic">BRE</Emphasis> (TNFRSF1A modulator/BRCC45) in response to stress and biological signals

Stress-responsive genes play critical roles in many biological functions that includes apoptosis, survival, differentiation and regeneration. We have identified a novel stress-responsive gene called BRE which interacts with TNF-receptor-1 and blocks the apoptotic effect of TNF-α. BRE enhances tumor growth in vivo and is up-regulated in hepatocellular and esophageal carcinomas. BRE also regulates the ubiquitination of the DNA repair complex BRCC, and the synthesis of steroid hormones. Here, we examined BRE-mRNA in cells after treatments with UV and ionizing radiation (IR). UV and IR treatment alone suppressed BRE-mRNA levels by more than 90% at 24 h, while hydroxyurea, fluorodeoxyuridine, aphidicolin, known inhibitors of S-phase DNA synthesis, had no significant effect. BRE protein expression was unaltered in cells treated with TNF-α, Interleukin-1 and Dexamethasone, while a threefold increase was observed following chorionic gonadotropin exposure. Although BRE plays a regulatory role in many different pathways, yet its expression is apparently under very stringent control.     

Lack of apoE causes alteration of cytokines expression in young mice liver

To investigate the effect of apolipoprotein E (apoE) on cytokine expression profile of the liver of young mice, quantitative RT-PCR (qRT-PCR) assay and cytokine antibody array for multiplex analysis of 62 cytokines have been used to analyze characteristics of expression of cytokines in the liver of 6-week-old apoE-null (apoE^−/−) mice. The levels of plasma cytokines were also analyzed. The mRNA level of IL-1β, IL-2, IL-6, ICAM-1, VCAM-1, MCP-1, NF-κB (p65), IFN-γ and IκB-α were increased significantly in apoE^−/− mice comparative to wild-type (WT) mice. IL-4, IL-10 and GM-CSF, however, were slightly decreased. Compared with WT, levels of 21 cytokines altered twofold or more in apoE^−/− mice, including 10 cytokines increased and 11 decreased. Expression patterns of IL-1β, IL-2, IL-4, IL-6, IL-10, GM-CSF, IFN-γ and VCAM-1 showed identical trend between cytokine antibody array and qRT-PCR analysis. Moreover, levels of IL-1β, IFN-γ and IL-6 in the plasma were elevated, while IL-4 was lightly decreased in apoE^−/− mice compared to those in WT mice. These results implied that promotion of type I immune response in the liver of young apoE^−/− mice due to alteration of these cytokines, and the phenotypes may be caused by the regulation of NF-κB. The inflammation and lipid metabolism dysfunction in the liver cooperated in dysfunction of the liver in young apoE^−/− mice.     

Molecular cloning and functional expression analysis of a new gene encoding geranylgeranyl diphosphate synthase from hazel (<Emphasis Type="Italic">Corylus avellana</Emphasis> L. Gasaway)

Geranylgeranyl diphosphate synthase (GGPPS) [EC 2.5.1.29] catalyzes the biosynthesis of geranylgeranyl diphosphate (GGPP), which is a key precursor for diterpenes such as taxol. Herein, a full-length cDNA encoding GGPPS (designated as CgGGPPS) was cloned and characterized from hazel (Corylus avellana L. Gasaway), a taxol-producing angiosperms. The full-length cDNA of CgGGPPS was 1515 bp with a 1122 bp open reading frame (ORF) encoding a 373 amino acid polypeptide. The CgGGPPS genomic DNA sequence was also obtained, revealing CgGGPPS gene was not interrupted by an intron. Southern blot analysis indicated that CgGGPPS belonged to a small gene family. Tissue expression pattern analysis indicated that CgGGPPS expressed the highest in leaves. RT–PCR analysis indicated that CgGGPPS expression could be induced by exogenous methyl jasmonate acid. Furthermore, carotenoid accumulation was observed in Escherichia coli carrying pACCAR25ΔcrtE plasmid carrying CgGGPPS. The result revealed that cDNA encoded a functional GGPP synthase.     

Purification of a bioactive recombinant human Reg IV expressed in Escherichia coli

Regenerating gene (Reg) IV is a newly discovered member of the regenerating gene family belonging to the calcium (C-type) dependent lectin superfamily. Reg IV is highly expressed in the gastrointestinal tract and markedly up-regulated in colon adenocarcinoma, pancreatic cancer, gastric adenocarcinoma, and inflammatory bowel disease. However, the physiological and pathological functions of Reg IV are largely unknown, partly due to the limited access of the bioactive protein. We report here the first expression and purification of Reg IV proteins using a prokaryotic system. Human Reg IV was expressed in Escherichia coli as an insoluble protein which was identified in the fraction of inclusion body after ultrasonication of the bacteria. After the protein aggregate was solubilized by guanidine–HCl, it was refolded by sucrose and arginine-assisted procedures and purified using cation-exchange chromatography. The protein identity and purity of the final preparation were confirmed by analysis of the protein mass and immune specificity in SDS–PAGE, Western blotting, and HPLC assay. The biological activity of the protein was determined by the HCT116 and HT29 cell proliferation assays. The highly purified bioactive human Reg IV should aid in further characterization of its physiological and pathological functions.     

A novel testis-specific GTPase serves as a link to proteasome biogenesis: functional characterization of RhoS/RSA-14-44 in spermatogenesis

Most Rho family GTPases serve as key molecular switches in a wide spectrum of biological processes. An increasing number of studies have expanded their roles to the spermatogenesis. Several members of Rho family have been confirmed to be essential for mammalian spermatogenesis, but the precise roles of this family in male reproduction have not been well studied yet. Here we report a surprising function of an atypical and testis-specific Rho GTPase, RSA-14-44 in spermatogenesis. Featured by unique structural and expressional patterns, RSA-14-44 is distinguished from three canonical members of Rho cluster. Thus, we define RSA-14-44 as a new member of Rho GTPases family and rename it RhoS (Rho in spermatogenic cells). RhoS associates with PSMB5, a catalytic subunit of the proteasome, in a series of stage-specific spermatogenic cells. More importantly, RhoS does not directly modulate the cellular proteasome activity, but participates in regulating the stability of "unincorporated" PSMB5 precursors. Meanwhile, our data demonstrate that the activation of RhoS is prerequisite for negatively regulating the stability of PSMB5 precursors. Therefore, our finding uncovers a direct and functional connection between the Rho GTPase family and the pathway of proteasome biogenesis and provide new clues for deciphering the secrets of spermatogenesis.     

Studies on the structure–activity relationship of 1,3,3,4-tetra-substituted pyrrolidine embodied CCR5 receptor antagonists. Part 1: Tuning the N-substituents

A novel series of CCR5 antagonists has been identified, utilizing the lead, nifeviroc, which were further modified based on bioisosteric principles. Lead optimization was pursued by balancing potential toxicity and potency. Potent analogues with low toxic properties were successfully developed by formation of urea and amide bonds at the nitrogen at position 4- of the pyrrolidine ring.     

Methane production from rice straw pretreated by a mixture of acetic–propionic acid

Rice straw was treated with a mixed solution of acetic acid and propionic acid to enhance its biodegradability. The effect of acid concentration, pretreatment time, and the ratio of solid to liquid on the delignification performance of rice straw were investigated. It was found that the optimal conditions for hydrolysis were 0.75 mol/L acid concentration, 2 h pretreatment time and 1:20 solid to liquid ratio. Batch methane fermentation of untreated rice straw, pretreated rice straw, and the hydrolysates (the liquid fraction) of pretreatment were conducted at 35 °C for 30 days, and the results indicated that methane production of rice straw can be enhanced by dilute organic acid pretreatment. Moreover, most of the acid in hydrolysates can also be converted into methane gas.     

Fatty acid synthase inhibitors of phenolic constituents isolated from Garcinia mangostana

Natural inhibitors of fatty acid synthase (FAS) are emerging as potential therapeutic agents to treat cancer and obesity. The bioassay-guided chemical investigation of the hulls of Garcinia mangostana led to the isolation of 13 phenolic compounds (1–13) mainly including xanthone and benzophenone, in which compounds 7, 8, 9, 10, and 11 were isolated from this plant for the first time and compound 9 was a new natural product. These isolates possess strong inhibitory activity of FAS with the IC₅₀ values ranging from 1.24 to 91.07 μM. The study indicates that two types of natural products, xanthones and benzophenones, could be considered as promising FAS inhibitors.     

Synthesis and structure–activity relationships of 2-aryl-4-oxazolylmethoxy benzylglycines and 2-aryl-4-thiazolylmethoxy benzylglycines as novel,potent PPARα selective activators- PPARα and PPARγ selectivity modulation

The synthesis and follow-up SAR studies of our development candidate 1 by incorporating 2-aryl-4-oxazolylmethoxy and 2-aryl-4-thiazolylmethoxy moieties into the oxybenzylglycine framework of the PPARα/γ dual agonist muraglitazar is described. SAR studies indicate that different substituents on the aryloxazole/thiazole moieties as well as the choice of carbamate substituent on the glycine moiety can significantly modulate the selectivity of PPARα versus PPARγ. Potent, highly selective PPARα activators 2a and 2l, as well as PPARα activators with significant PPARγ activity, such as 2s, were identified. The in vivo pharmacology of these compounds in preclinical animal models as well as their ADME profiles are discussed.