The inhibiting effect of the transcription factor p53 on dengue virus infection by activating the type I interferon.

To investigate the role of the transcription factor p53 in the course of the dengue virus (DV) infection. The human hepatocellular carcinoma cell strain HepG2 with a low expression level of p53 was built by using the retroviral-mediated RNA interference technology, and was detected by Western blot. The wild group and the interference group were respectively infected by the type 2 DV. The viral titration was detected by the Vero plaque assay, the viral multiplication was detected by the immunofluorescence, the cell apoptosis after virus infection was detected by FCM and the level of IFN-ß was analyzed by ELISA. Compared to the wild group, the expression level of p53 in the interference group decreased significantly, which indicated that the HepG2 cell strain with the low expression level of p53 was successfully built. 24h after DV infection, the virus titration in the interference group was 100 times higher than that in the wild group. The result of the immunofluorescence showed that, the amount of green fluorescent cells in the interference group was significant higher than that in the wild group. It was indicated that the DV infection was inhibited by p53. However, 24h after DV infection, there was no significant difference in the amount of apoptotic cells in both groups. And the amount of IFN-ß in the wild group increased 6 times. The DV infection was inhibited by the transcription factor p53 by activating type I interferon pathway other than promoting the cell apoptosis.

Dax1 and Nanog act in parallel to stabilize mouse embryonic stem cells and induced pluripotency.

Nanog expression is heterogeneous and dynamic in embryonic stem cells (ESCs). However, the mechanism for stabilizing pluripotency during the transitions between Nanog(high) and Nanog(low) states is not well understood. Here we report that Dax1 acts in parallel with Nanog to regulate mouse ESC (mESCs) identity. Dax1 stable knockdown mESCs are predisposed towards differentiation but do not lose pluripotency, whereas Dax1 overexpression supports LIF-independent self-renewal. Although partially complementary, Dax1 and Nanog function independently and cannot replace one another. They are both required for full reprogramming to induce pluripotency. Importantly, Dax1 is indispensable for self-renewal of Nanog(low) mESCs. Moreover, we report that Dax1 prevents extra-embryonic endoderm (ExEn) commitment by directly repressing Gata6 transcription. Dax1 may also mediate inhibition of trophectoderm differentiation independent or as a downstream effector of Oct4. These findings establish a basal role of Dax1 in maintaining pluripotency during the state transition of mESCs and somatic cell reprogramming.

[Expression profile analysis of host HeLa cells invasived by Shigella flexneri 2a].

The changes of genes expression in HeLa cell during the invasion with Shigella species for 1h and 3h were analyzed by cDNA microarrays. The data showed that the expression levels of 752 genes were altered twice or greater as compared with the control 509 of them were up-regulated, and 306 were down-regulated. It was supposed that some signal pathways in HeLa cell were activated, then many genes were induced, and at last comprehensive cell responses were produced, so that HeLa cell could prevent against Shigella species infection. Two striking difference cDNA fragments TNFR 1B and ERBB2, which were up-regulated in the host epithelial cell during Shigella infection, analyzed expression by real time RT-PCR as determined by cDNA arrays. We suggested they play important roles in response to the invasive S. flexneri 2457T. Based on the results of gene expression alterations, the molecular pathogenic mechanism of Shigella species could be greatly and deeply understood, and the strategy for prevention against and treatment for shigellosis would be formed.

[Study on expression of Streptococcus suis serotype 2 sly gene, purification and activity of its product].

OBJECTIVE: To clone and express Streptococcus suis serotype 2 (S. suis 2) sly gene for constructing an foundation on identification of S. suis 2 protective antigen. METHODS: The sly gene was amplified from S. suis 2 clinical isolate strain 05ZYH33 genome DNA by PCR. The gene fragment was inserted into the expression vector pET-30b(+) to build pET30b-sly. When recombinant vector pET30b-sly was identified by restriction enzyme cutting and DNA sequencing as a correct one, subsequently it was transformed to E. coli Rosetta for expression under IPTG induction. The obtained fusion protein was purified by Ni-NTA affinity chromatography. The immunologic and hemolysis activity of the purified protein was proved through Western blot and hemolysis assay respectively. RESULTS: The PCR product was around 1500 bp. The gene segment inserted into the recombinant vector was proven to be completely identical with the sly gene sequence in the total genome sequence of S. suis 2. The target protein expressed was up to 30% of the total somatic protein under IPTG induction. The protein purity reached above 80% after purification. The protein could be recognized by human serum infected with S. suis 2 and could dissolve swine erythrocytes with the Hemolytic titer as 256. CONCLUSION: The expression vector pET30b-sly was successfully constructed. The target protein could be over-expressed in E. coli and possessed its biological activity after purification.