Identification and Genomic Analysis of a Pathogenic Strain of Mycoplasma hyopneumoniae (TB1) Isolated from Tibetan Pigs.

The present study aims to identify the species and strains of Mycoplasma hyopneumoniae isolated from Tibetan pigs (Mh TB1) at the genetic level for understanding the basis of its pathogenicity. Mh TB1 was isolated from the consolidated lungs of Tibetan pigs by liquid culture and agar plate colony method. Polymerase chain reaction (PCR) amplification of the 16S recombinant DNA (rDNA) conservative sequence and a species-specific gene (P36) of Mh provided species confirmation. PCR products were imaged on gels and shotgun sequencing was performed. DNA sequences were compared for assessing genetic similarity between Mh TB1 and Mh reference strains in the GenBank database. The isolated strains were >98% similar to the Mh reference strains. Genomic analysis revealed significant sequence conservation between Mh TB1 and the reference strains; however, differential genes were more prevalent in Mh TB1 than in other reported strains. Therefore, we concluded that Mh is a major pathogen of Tibetan pigs that cause enzootic pneumonia. The Mh TB1 strain harbors more genes and specific virulence factors, consistent with its plateau-related adaptability to hypoxia and virulence. Differential gene analysis revealed gene variations in the inclement plateau environment, enriched gene pool, and plateau adaptability of the Mh TB1 strain, which will be important for vaccine development.

Ferritin light chain and squamous cell carcinoma antigen 1 are coreceptors for cellular attachment and entry of hepatitis B virus.

Overexpression of squamous cell carcinoma antigen 1 (SCCA1) in hepatitis G2 (HepG2) and Chinese hamster ovary cells can increase hepatitis B virus (HBV) binding capacity by interacting with the preS1 domain of the HBV surface antigen. However, the magnitude of increase in binding capacity was higher by several orders in the former, indicating the existence of additional factor(s) produced by HepG2 cells, which facilitates HBV attachment. Ferritin light chain (FTL) was identified as the sole high hit candidate by screening human liver cDNA library using a bacterial two-hybrid system with either preS or SCCA1 as the bait. Subsequent in vitro protein-protein interaction assays confirmed the binding activity of FTL to both preS and SCCA1, as well as the formation of triple complex preS-FTL-SCCA1, and narrowed down the binding sites on FTL. In vitro overexpression of FTL could further enhance HBV attachment in both HepG2 and Chinese hamster ovary cells, which were already overexpressing SCCA1. Importantly, in vivo co-expression of human FTL and SCCA1 in mouse liver by means of tailvein hydrodynamic injection increased serum levels of HBV surface antigen transiently 24 hours post challenge with HBV-positive human sera, and a large amount of HBV core antigen-positive hepatocytes around blood vessels could be identified by immunohistochemical staining 48 hours post challenge. The data strongly suggest that FTL and SCCA1 may serve as coreceptors in HBV cellular attachment and virus entry into hepatocytes.

Hydrophobic cavity in C-terminus is essential for hTNF-α trimer conformation.

A variety of tumour necrosis factor α (TNF-α) derivatives have been bioengineered to improve antitumour activity and reduce toxicity. The expression of TNF-α in Escherichia coli usually yields a mixture of homotrimers and monomers; however, only the trimer shows antitumour activity. TNF-αD10, a bioengineered hTNF-α derivative, demonstrated 10-fold higher cytotoxicity against tumour cells compared to hTNF-α, but the trimer to monomer ratio was 58:42. In the present study, we investigated the structural differences between the trimer and the monomer of TNF-αD10. We found that the chemical shifts of the C-terminal Trp(114) in the trimer were significantly different from those in the monomer and that the replacement of Trp(114) with different amino acids remarkably reduced the trimer production. Further analysis of the publicly available X-ray crystallographic data for trimeric and monomeric hTNF-α revealed that the conformation of the U-shaped region formed by the fragment Cys(101)-Trp(114) was different between the two forms: a hydrophilic cavity in the monomer and a hydrophobic cavity in the trimer. These findings suggested the potential approaches of molecular and structural modification for future improvement of hTNF-α trimer production.

Involvement of PU.1 in mouse adar-1 gene transcription induced by high-dose esiRNA.

Adar-1 gene plays an important role in the negative regulation of RNA interference. We previously showed that increased adar-1 mRNA level was associated with the rebound of gene expression after RNAi suppression. In this study, we identified a PU.1 binding site upstream from transcription start point of adar-1 gene and is essential for the promoter activity. Knockdown and over-expression of the PU.1 gene resulted in decreased and increased activity of adar-1 promoter, respectively. Our results suggest that transcription factor PU.1, could bind to the adar-1 promoter and play a key role in activating transcription of gene induced by high-dose esiRNAs.

[Genetic polymorphism of mitochondrial DNA in coding region in 16 ethnic populations of Yunnan].

The genetic polymorphism of mitochondrial DNA in the coding region in 16 ethnic populations of Yunnan was analyzed using PCR-RFLP in a total of 654 samples. Seventeen haplogroups were found, four of which were undefined haplogroups. Haplogroup distribution and Principal Component (PC) analysis showed that the ethnic groups descended from Bai-Yue tribe have B, F and M7 as the predominant haplogroups,which indicated their origination from southern China. Haplogroup A, D and N9 were predominant in the Mongolian ethnic group, reflecting their north-originated characteristics. The groups descended from Di-Qiang tribe shared the predominant haplogroups with both the south and north originated groups,which demonstrated that they inherit the maternal characteristics from both southern and northern populations. There is genetic difference among the populations in the same ethnic group and is usually smaller than that among the ethnic groups from different ancient tribes, but not necessarily smaller than that among the groups from the same tribe.

[The frequency distribution of flavin-containing monooxygenase 3 mutant alleles in 28 populations from Yunnan].

OBJECTIVE: To study the frequency distribution of flavin-containing monooxygenase 3 (FMO3) mutant alleles in 28 populations originating from 24 ethnic minorities in Yunnan of China. METHODS: FMO3 genotypes were analyzed by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: The average frequencies of FMO3/Stop(148), FMO3/Lys(158) and FMO3/Gly(308) were 0.395(0.174-0.803), 0.208 (0.056-0.414), 0.046(0-0.217), respectively. The frequencies of FMO3/Gly(308) in Blang, Huayaodai, Shuidai, Zhuang, De'ang, Jingpo, Nu and Hui populations were null. CONCLUSION: It was found that the frequencies of FMO3 mutant alleles varied not only in different ethnic groups, but also in different populations that stemmed from the same ethnic group.