A convenient model of severe, high incidence autoimmune gastritis caused by polyclonal effector T cells and without perturbation of regulatory T cells

Autoimmune gastritis results from the breakdown of T cell tolerance to the gastric H(+)/K(+) ATPase. The gastric H(+)/K(+) ATPase is responsible for the acidification of gastric juice and consists of an α subunit (H/Kα) and a β subunit (H/Kβ). Here we show that CD4(+) T cells from H/Kα-deficient mice (H/Kα(-/-)) are highly pathogenic and autoimmune gastritis can be induced in sublethally irradiated wildtype mice by adoptive transfer of unfractionated CD4(+) T cells from H/Kα(-/-) mice. All recipient mice consistently developed the most severe form of autoimmune gastritis 8 weeks after the transfer, featuring hypertrophy of the gastric mucosa, complete depletion of the parietal and zymogenic cells, and presence of autoantibodies to H(+)/K(+) ATPase in the serum. Furthermore, we demonstrated that the disease significantly affected stomach weight and stomach pH of recipient mice. Depletion of parietal cells in this disease model required the presence of both H/Kα and H/Kβ since transfer of H/Kα(-/-) CD4(+) T cells did not result in depletion of parietal cells in H/Kα(-/-) or H/Kβ(-/-) recipient mice. The consistency of disease severity, the use of polyclonal T cells and a specific T cell response to the gastric autoantigen make this an ideal disease model for the study of many aspects of organ-specific autoimmunity including prevention and treatment of the disease.     

Effects of the C-terminal truncation in NS1 protein of the 2009 pandemic H1N1 influenza virus on host gene expression

The 2009 pandemic H1N1 influenza virus encodes an NS1 protein with 11 amino acids (aa) truncation at the C-terminus. The C-terminal tail of influenza virus NS1 protein constitutes a nucleolar localization signal (NoLS) and is the binding domain of the cellular pre-mRNA processing protein, poly(A)-binding protein II (PABII). Here, our studies showed that the C-terminal-truncated NS1 of the 2009 pandemic virus was inefficient at blocking host gene expression, extension of the truncated NS1 to its full length increased the inhibition of host gene expression. Mechanistically, this increased inhibition of host gene expression by the full-length NS1 was not associated with nucleolar localization, but was due to the restoration of NS1's binding capacity to PABII. Furthermore, in vitro and in vivo characterization of two recombinant viruses encoding either the C-terminal 11-aa truncated or full-length NS1 of the 2009 pandemic virus showed that the C-terminal 11-aa truncation in NS1 did not significantly alter virus replication, but increased virus pathogenicity in mice.     

Combined Genomics and Experimental Analyses of Respiratory Characteristics of Shewanella putrefaciens W3-18-1

It has previously been shown that the Shewanella putrefaciens W3-18-1 strain produces remarkably high current in microbial fuel cells (MFCs) and can form magnetite at 0°C. To explore the underlying mechanisms, we developed a genetic manipulation method by deleting the restriction-modification system genes of the SGI1 (Salmonella genome island 1)-like prophage and analyzed the key genes involved in bacterial respiration. W3-18-1 has less respiratory flexibility than the well-characterized S. oneidensis MR-1 strain, as it possesses fewer cytochrome c genes and lacks the ability to oxidize sulfite or reduce dimethyl sulfoxide (DMSO) and timethylamine oxide (TMAO). W3-18-1 lacks the hydrogen-producing Fe-only hydrogenase, and the hydrogen-oxidizing Ni-Fe hydrogenase genes were split into two separate clusters. Two periplasmic nitrate reductases (NapDAGHB and NapDABC) were functionally redundant in anaerobic growth of W3-18-1 with nitrate as the electron acceptor, though napDABC was not regulated by Crp. Moreover, nitrate respiration started earlier in W3-18-1 than in MR-1 (with NapDAGHB only) under microoxic conditions. These results indicate that Shewanella putrefaciens W3-18-1 is well adapted to habitats with higher oxygen levels. Taken together, the results of this study provide valuable insights into bacterial genome evolution.     

Directed evolution of cytochrome P450 for sterol epoxidation

16,17-Epoxysterol plays an important role in pharmaceutical steroid synthesis. To investigate the potential application of cytochrome P450 for epoxysterol synthesis, an approach to the epoxidation of 16,17-epoxysterol, based on directed evolution of cytochrome P450 BM-3, was developed. This comprised random gene mutagenesis for optimizing the activity of P450 BM-3 for epoxidation of hydrophobic sterol, followed by the 7-ethoxycoumarin de-ethylation assay for general enzyme activity detection and the modified picric acid assay for epoxidation activity screening. By the two-step screening, one mutant from 792 clones showed specific substrate activity of converting progesterone to 16,17-epoxysterol, which validated the possibility to evolve the cytochrome P450 for the synthesis of steroidal epoxides.     

Heat stress-induced BBX18 negatively regulates the thermotolerance in Arabidopsis

There is increasing evidence for considerable interlinking between the responses to heat stress (HS) and light signaling. In the present work, we provide molecular evidence that BBX18, a negative regulator in photomorphogenesis belonging to the B-box zinc finger protein family in Arabidopsis thaliana, is involved in the regulation of thermotolerance. Using quantitative RT-PCR, GUS staining and immunoblot analysis, our results indicate that the expression of BBX18 was induced by HS. BBX18-RNAi and 35S::BBX18 transgenic Arabidopsis plants were obtained for functional analysis of BBX18. Under-expression of BBX18 displayed increased both basal and acquired thermotolerance in the transgenic plants, while over-expression of BBX18 reduced tolerance to HS in transgenic lines. Moreover, when wild-type, BBX18-RNAi and 35S::BBX18 transgenic plants were treated with HS, HR-related digalactosyldiacylglycerol synthase 1 (DGD1) was down-regulated by BBX18 in both normal and heat shock conditions. Besides, the expression levels of Hsp70, Hsp101 and APX2 were increased in BBX18-RNAi transgenic plants, but lower in 35S::BBX18 transgenic plants. However, the expression of HsfA2 was lower in BBX18-RNAi transgenic plants and higher in the 35S::BBX18 after high-temperature treatment. These results suggesting that, by modulated expression of a set of HS-responsive genes, BBX18 weakened tolerance to HS in Arabidopsis. So our data indicate that BBX18 plays a negative role in thermotolerance.     

卵巢癌细胞与巨噬细胞共培养对B7-H1表达的影响及机制

为了探讨卵巢癌细胞与巨噬细胞共培养后对B7．H1表达的影响及其可能机制,利用佛波酯（PMA）诱导THP-1或外周血单核细胞分化为巨噬细胞后,与人卵巢癌细胞株SKOV3体外非接触共培乔24h,qRT-PCR、Western blot以及流式细胞术分别检测SKOV3与巨噬细胞B7-H1的表达：进一步利用NF-KB、JAK2／STAT3、p38MAPK信号通路的抑制剂作用于共培养体系,检测B7-H1表达的变化,以探讨其机制。结果显示,共培养24h后,SKOV37L巨噬细胞B7-H1mRNA和蛋白的表达较非共培养组均显著升高（P〈0．05）,而阻断NF-κB、JAK2／STAT3、p38MAPK信号通路后,B7-H1的上调均明显被抑制（P〈0．05）。SKOV3与巨噬细胞共培养后B7-H1的表达升高伊〈0．05）,其机制可能涉及到NF—κB、JAK2／STAT3、p38MAPK信号通路的激活。     

Large cargo transport by nuclear pores: implications for the spatial organization of FG‐nucleoporins

Nuclear pore complexes (NPCs) mediate cargo traffic between the nucleus and the cytoplasm of eukaryotic cells. Nuclear transport receptors (NTRs) carry cargos through NPCs by transiently binding to phenylalanine‐glycine (FG) repeats on intrinsically disordered polypeptides decorating the NPCs. Major impediments to understand the transport mechanism are the thousands of FG binding sites on each NPC, whose spatial distribution is unknown, and multiple binding sites per NTR, which leads to multivalent interactions. Using single molecule fluorescence microscopy, we show that multiple NTR molecules are required for efficient transport of a large cargo, while a single NTR promotes binding to the NPC but not transport. Particle trajectories and theoretical modelling reveal a crucial role for multivalent NTR interactions with the FG network and indicate a non‐uniform FG repeat distribution. A quantitative model is developed wherein the cytoplasmic side of the pore is characterized by a low effective concentration of free FG repeats and a weak FG‐NTR affinity, and the centrally located dense permeability barrier is overcome by multivalent interactions, which provide the affinity necessary to permeate the barrier.     

First report of HGD mutations in a Chinese with alkaptonuria

Alkaptonuria (AKU) is one of the first prototypic inborn errors in metabolism and the first human disease found to be transmitted via Mendelian autosomal recessive inheritance. It is caused by HGD mutations, which leads to a deficiency in homogentisate 1,2-dioxygenase (HGD) activity.     

Effects of site-specific guanine C8-modifications on an intramolecular DNA G-quadruplex

Understanding the fundamentals of G-quadruplex formation is important both for targeting G-quadruplexes formed by natural sequences and for engineering new G-quadruplexes with desired properties. Using a combination of experimental and computational techniques, we have investigated the effects of site-specific substitution of a guanine with C8-modified guanine derivatives, including 8-bromo-guanine, 8-O-methyl-guanine, 8-amino-guanine, and 8-oxo-guanine, within a well-defined (3 + 1) human telomeric G-quadruplex platform. The effects of substitutions on the stability of the G-quadruplex were found to depend on the type and position of the modification among different guanines in the structure. An interesting modification-dependent NMR chemical-shift effect was observed across basepairing within a guanine tetrad. This effect was reproduced by ab initio quantum mechanical computations, which showed that the observed variation in imino proton chemical shift is largely influenced by changes in hydrogen-bond geometry within the guanine tetrad.