Bel1, a transactivator of prototype foamy virus (PFV), plays pivotal roles in the replication of PFV. Previous studies have shown that Bel1 bears a nuclear localization signal (NLS), but its amino acid sequence remains unclear and the corresponding importins have not been identified. In this report, we inserted various fragments of Bel1 into an EGFP-GST fusion protein and investigated their subcellular localization by fluorescence microscopy. We found that the 215PRQKRPR221 fragment could direct nuclear localization, which accords with the consensus sequence K(K/R)X(K/R) of monopartite NLS. Point mutation experiments revealed that K218, R219, and R221 are essential for the nuclear localization of Bel1. The results of the GST-pulldown showed that the Bel1 fragment with residues 215-223, which bears the NLS, interacts with KPNA1, KPNA6, and KPNA7. This result suggests that KPNA1, KPNA6, and KPNA7 maybe involved in Bel1 nuclear translocation.
Cell Line ; Cell Nucleus ; genetics ; metabolism ; virology ; Humans ; Nuclear Localization Signals ; genetics ; metabolism ; Protein Binding ; Protein Transport ; Retroviridae Infections ; genetics ; metabolism ; virology ; Retroviridae Proteins ; chemistry ; genetics ; metabolism ; Spumavirus ; chemistry ; genetics ; physiology ; Trans-Activators ; chemistry ; genetics ; metabolism ; alpha Karyopherins ; genetics ; metabolism

Adult ; CD56 Antigen ; metabolism ; Combined Modality Therapy ; Diagnosis, Differential ; Humans ; Lymphoma, Extranodal NK-T-Cell ; metabolism ; pathology ; Lymphoma, Large B-Cell, Diffuse ; metabolism ; pathology ; therapy ; Lymphoma, Large-Cell, Anaplastic ; metabolism ; pathology ; Male ; Neprilysin ; metabolism ; Proto-Oncogene Proteins c-bcl-6 ; metabolism

Objective To investigate the mutation of hemagglutinin gene and amino acid variation of influenza B.Methods Influenza B virus was isolated from throat swab samples in sentinel surveillance of Yunnan province from 2009 to 2014.HA1 gene sequence analysis was applied to determine 12 randomly-selected strains of influenza B virus.The results were analyzed,MEGA software was used to do homology comparison and HA gene phylogenetic tree was established.Results Differences on the serotype and genotype identification of influenza strains were found and it might be caused by inadequate gene mutation accumulation.Amino acid variations were found in 3 important regions of antigenic determinants in HA1 protein:ring 120,ring 150 and ring 160.The amino acid variation of position 131 in ring 120 was N131K,and in position 137 was N137H.Two strains had P187S mutation in position 187.Conclusion There are some important variations in the hemagglutinin gene of influenza B strains in Yunnan Province,with some variations being the same as vaccine strains and some being not.

A research combined trickling filter system and active sludge aeration system was applied in the treatment of industrial wastewater from gas-generating with heavy oil. The wastewater contained both high contents of NH+4-N and mixed hydrocarbons including various PAHs. Its BOD5/COD ratio was less than 0.3 and belongs to recalcitrant, toxic wastewater. The results showed a touch-growth biofilms system was formed on the porous packing material and it played a key role in the decrease of toxicity of the influent. It could also improve the biodegradability of the wastewater.

Adrenal Gland Neoplasms ; pathology ; surgery ; Adrenal Glands ; pathology ; surgery ; Child, Preschool ; Diagnosis, Differential ; Female ; Follow-Up Studies ; Granuloma, Plasma Cell ; pathology ; surgery ; Histiocytoma, Malignant Fibrous ; pathology ; Humans ; Neoplasms, Muscle Tissue ; pathology ; surgery

Antigens, Neoplasm ; Brain ; metabolism ; pathology ; Humans ; Infant, Newborn ; Lung ; metabolism ; pathology ; Male ; Melanoma-Specific Antigens ; Melanosis ; complications ; congenital ; metabolism ; pathology ; Neoplasm Proteins ; metabolism ; Neurocutaneous Syndromes ; complications ; congenital ; metabolism ; pathology ; S100 Proteins ; metabolism ; Skin ; metabolism ; pathology

AIM To establish an HPLC method for the simultaneous content determination of four constituents in Kesuting Capsules (a fast cough suppressant,containing Ephedrae Herba,Papaveris Pericarpium,Platycodonis Radix,etc.).METHODS The analysis of trichloromethane-strong ammonia extract of Kesuting Capsules was performed on a 35 ℃ thermostatic Welch Ultimate(◎) XB-C18 column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.01 mol/L potassium dihydrogen phosphate buffer flowing at 1.0 mL/min in an isocratic elution manner,and the detection wavelength was set at 210 nm.RESULTS Morphine,ephedrine hydrochloride,pseudoephedrine hydrochloride and codeine phosphate showed good linear relationships within the ranges of 8.054-67.12 μg/mL (r =0.999 5),22.31-185.9 μg/mL (r =0.999 9),21.26-177.2 μg/mL (r =0.999 7) and 1.212-10.09 μg/mL (r =0.999 7),whose average recoveries (n =9) were 100.9％ (RSD =2.0％),101.4％ (RSD =3.6％),105.3％ (RSD =1.2％) and 106.2％ (RSD =1.2％),respectively.CONCLUSION This simple method can be used for the rapid quality control of Kesuting Capsules.

ATP sulfurylase (ATPS,EC 2.7.7.4) reversibly catalyzes the reaction between ATP and sulfate to produce APS and pyrophosphate (PPi), and has been used in pyrosequencing. The gene coding ATP sulfurylase was amplified from the genomic DNA of Saccharomyces cerevisias (CICC 1202), and cloned into prokaryotic expression plasmid pET28a( + ) to provide a recombinant expression plasmid pET28a( + )-ATPS. Upon IPTG induction, ATP sulfurylase was produced by E. coli BL21 (DE3) harboring the recombinant expression plasmid pET28a( + )-ATPS. The relative molecular weight of recombinant ATP sulfurylase with His tag was about 60 kD. The recombinant ATP sulfurylase with electrophoretic pure grade was obtained only by two purification steps: His * Bind Resin affinity chromatography and ultrafiltration. The specific activity of the purified recombinant ATP sulfurylase was as high as 5.1 x 10(4) u/mg. The successful application of the enzyme in pyrosequencing was also demostrated.
Escherichia coli ; genetics ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification ; Saccharomyces cerevisiae ; enzymology ; genetics ; Sequence Analysis, DNA ; methods ; Sulfate Adenylyltransferase ; biosynthesis ; genetics

This study was purposed to explore the mechanisms of preventive effect of tetrandrine (TTD) on doxorubicin (ADM)-induced multidrug resistance (MDR) in human leukemia cell line K562 from two aspects of the transcription control of MDR1 gene and cell apoptosis. The experiment was divided into 3 groups: group I-blank control; group II-ADM-induced drug-resistance; group III-ADM-induced drug-resistance after pretreatment with TTD. Reverse transcription-PCR (RT-PCR) was used to detect the mRNA expression levels of c-Jun, YB-1 and Survivin genes. Western blot was used to determine the nuclear protein expression levels of c-Jun and YB-1. Flow cytometry was used to assay the apoptosis of cells. The results showed that as compared with group I, the expression levels of c-Jun mRNA and nuclear protein decreased (p < 0.05), as well as the expression levels of YB-1 mRNA and nuclear protein increased in group II (p < 0.05). However, the expression of Survivin mRNA had no change (p > 0.05); the apoptosis rate of cells was 8.31%. As compared with group II, the expression levels of c-Jun mRNA and nuclear protein increased (p < 0.05), expression levels of YB-1 mRNA and nuclear protein as well as Survivin mRNA decreased in group III (p < 0.05). The apoptosis of cells was 97.2%. It is concluded that TTD can inhibit the expression of YB-1 and up-regulate the expression of c-Jun, thus inhibit the expression of MDR1 gene. TTD can also inhibit the expression of Survivin and increase the apoptosis of cells induced by ADM.
ATP-Binding Cassette, Sub-Family B, Member 1 ; metabolism ; Apoptosis ; drug effects ; genetics ; Benzylisoquinolines ; pharmacology ; Drug Resistance, Multiple ; drug effects ; genetics ; Drug Resistance, Neoplasm ; drug effects ; genetics ; Humans ; Inhibitor of Apoptosis Proteins ; metabolism ; K562 Cells ; Proto-Oncogene Proteins c-jun ; metabolism ; Y-Box-Binding Protein 1 ; metabolism

OBJECTIVETo investigate the effect of tetrandrine (TTD) on doxorubicin-induced mdr1 gene expression and its mechanism.

METHODSMTT assay was used to detect the cytotoxicity of TTD to K562 cells. K562 cells were treated with doxorubicin alone or 0.6 microg/ml doxorubicin combined with various concentrations of TTD. RT-PCR was used to detect the mRNA expression of mdr1 and NF-kappa B. Flow cytometry was used to assay the expression of P-glycoprotein (P-gp). Intracellular rhodamine 123 (Rho123) retention assay was applied to test the P-gp function.

RESULTSAfter treatment with 0.6 microg/ml doxorubicin for 24 hours, the expressions of mdr1 mRNA, NF-kappa B mRNA and P-gp in K562 cells were increased from 0.171 +/- 0.012, 0.783 +/- 0.090, 7.85 +/- 0.15 to 0.428 +/- 0.012, 1.075 +/- 0.047 and 73.68 +/- 1.84, respectively. The intracellular Rho123 retention was decreased from 711.9 +/- 63.6 to 347.8 +/- 60.6, indicating up-regulation of P-gp function (P<0.05). Pretreatment of K562 cells with 2.0 microg/ml TTD for 24 hours and then incubated for another 24 h with doxorubicin, the expressions of mdr1 mRNA, NF-kappa B mRNA, P-gp and up-regulation of P-gp function induced by doxorubicin were prevented in K562 cells (0.148 +/- 0.006, 0.627 +/- 0.098, 7.18 +/- 0.38 and 799.7 +/- 45.8, respectively P<0.05). But 0.5 microg/ml and 1.0 microg/ml TTD had little effect.

CONCLUSIONSTTD inhibits the expression of mdr1 mRNA, P-gp and up-regulated P-gp function induced by doxorubicin in a dose dependent manner. The mechanism of this effect may be down-regulation of NF-kappa B by TTD.

ATP Binding Cassette Transporter, Sub-Family B ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Benzylisoquinolines ; pharmacology ; Doxorubicin ; pharmacology ; Humans ; K562 Cells ; NF-kappa B ; metabolism ; RNA, Messenger ; genetics ; Up-Regulation ; drug effects