Seasonal differences in the transcriptome profile of the Zhedong white goose (Anser cygnoides) pituitary gland.

In animals, the adaptation to breed at the time of greatest survival of the young is known as seasonal reproduction. This is mainly controlled by the photoperiod, which stimulates the hypothalamic-pituitary-gonadal axis and starts the breeding season. Herein, we have determined the seasonal changes in gene expression patterns of Zhedong white geese pituitary glands under a natural photoperiodism, conducted at autumn equinox (AE), winter solstice (WS), spring equinox (SE), and summer solstice (SS). Pairwise comparisons of WS vs. AE, SE vs. WS, SS vs. SE, and AE vs. SS resulted in 1,139, 33, 704, and 3,503 differently expressed genes, respectively. When compared with SS, AE showed downregulation of genes, such as vasoactive intestinal peptide receptor, prolactin receptor, and thyroid hormone receptor beta, whereas gonadotropin-releasing hormone II receptor was upregulated, indicating that these genes may be responsible for the transition from cessation to egg laying. In addition, the expression levels of 5 transcription factors (POU1F1, Pitx2, NR5A1, NR4A2, and SREBF2) and 6 circadian clock-associated genes (Clock, Per2, ARNTL2, Eya3, Dio2, and NPAS2) also changed seasonally. Gene Ontology term and Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that "response to oxidative stress" and steroid biosynthesis pathway also participate in regulating the reproduction seasonality of geese. Overall, these results contribute to the identification of genes involved in seasonal reproduction, enabling a better understanding of the molecular mechanism underlying seasonal reproduction of geese.

Porcine genome engineering for xenotransplantation.

The extreme shortage of human donor organs for treatment of patients with end-stage organ failures is well known. Xenotransplantation, which might provide unlimited organ supply, is a most promising strategy to solve this problem. Domestic pigs are regarded as ideal organ-source animals owing to similarity in anatomy, physiology and organ size to humans as well as high reproductive capacity and low maintenance cost. However, several barriers, which include immune rejection, inflammation and coagulative dysfunctions, as well as the cross-species transmission risk of porcine endogenous retrovirus, blocked the pig-to-human xenotransplantation. With the rapid development of genome engineering technologies and the potent immunosuppressive medications in recent years, these barriers could be eliminated through genetic modification of pig genome together with the administration of effective immunosuppressants. A number of candidate genes involved in the regulation of immune response, inflammation and coagulation have been explored to optimize porcine xenograft survival in non-human primate recipients. PERV inactivation in pigs has also been accomplished to firmly address the safety issue in pig-to-human xenotransplantation. Many encouraging preclinical milestones have been achieved with some organs surviving for years. Therefore, the clinical trials of some promising organs, such as islet, kidney and heart, are aimed to be launched in the near future.

Exploration of the temporal-spatial expression pattern and DNA methylation-related regulation of the duck telomerase reverse transcriptase gene.

Telomerase reverse transcriptase (TERT) is a catalytic subunit of telomerase that adds TTAGGG repeats to the 3'-overhang of telomeres. In the present study, we detected that the duck TERT (dTERT) gene was highly expressed in small intestine and kidney, followed by heart, leg muscle, spleen, pancreas, gonad, and liver at neonatal stage. From embryonic to neonatal stage, the highest dTERT mRNA in liver appeared at stage E19 (19 days at embryonic stage), while for the leg muscle the maximum expression occurred at E26. We also measured the relative telomerase activity (RTA) and relative telomere length (RTL) in the examined tissues and found that the changed tendency of RTA and RTL was not very consistent with that of TERT. In silico analysis revealed that there were three CpG islands (S1, S2, and S3) within the 5' regulatory region of the dTERT gene. Bisulfite sequencing PCR (BSP) assay showed that liver (D7, 7 days after birth) which expressed significantly lower dTERT mRNA had an obviously higher methylation level of S1 compared with small intestine (D7) or liver (E19). Quantitative real-time PCR analysis revealed that the expression of DNA methyltransferase DNMT1 in liver (D7) was significantly higher than that in small intestine (D7) or in liver (E19). In vitro, dTERT expression was upregulated and the methylation status of S1 decreased in both duck embryonic fibroblasts and small intestinal epithelial cells following treatment with the demethylation reagent, 5-aza-2'-deoxycytidine (5-aza-dC), further suggesting that dTERT is epigenetically regulated by DNA methylation. This work lays a solid foundation for further study of TERT function and regulation in avian species.

Phoenixin: A Newly Discovered Peptide with Multi-Functions.

Phoenixin (PNX), a newly identified peptide cleaved from the C-terminus of protein C4orf52, mainly exists in two active isoforms, phoenixin-14 (PNX-14) amide and phoenixin-20 (PNX-20) amide that were first isolated from the rat hypothalamus and bovine heart, respectively. Initial studies demonstrated that PNX is a reproductive peptide, which affects the hypothalamus pituitary genital (HPG) axis through regulating the expression of kisspeptin, GnRH, GnRH receptor, LH and oestrus process. However, further studies indicated that PNX might play a wide range of roles in additional physiological process such as inhibiting visceral pain and eliciting pruritus, inducing anxiety, improving memory retention. Recently, Gpr173, also designated as SREB3, was identified as the cognate receptor of PNX. Whereas, the regulatory mechanism of PNX has not been fully clarified. This review aims to provide the current knowledge of PNX and propose some study directions for future research.

Construction, expression and characterization of a single chain variable fragment antibody against human myostatin.

Myostatin plays negative roles in muscle development. To block the inhibitory effects of myostatin on myogenesis, a 759 bp single chain variable fragment antibody (scFv) against myostatin was constructed and expressed in Escherichia coli. ELISA detection showed that the scFv could bind to myostatin, and change of the scFv N-terminal peptides decreased its binding affinity. MTT assay and cell morphology demonstrated that the cell number and viability of the C2C12 myoblast were enhanced by the scFv. Meanwhile, the scFv significantly inhibited the myostatin-induced expression of cyclin-dependent kinase inhibitor p21 and Smad binding element-luciferase activity. H2O2 increased the expression of Muscle RING Finger 1 (MuRF1) and Muscle Atrophy F-box (MAFbx) in myoblasts as well as myostatin and MuRF1 in myotubes, and the scFv significantly decreased the H2O2-elevated expression of these genes. Conclusively, the scFv we developed could antagonize the inhibitory effects of myostatin on myogenesis through Smad pathway and regulation of p21, MuRF1 and MAFbx gene expression. The scFv may have application in the therapy of muscular dystrophy and improvement of animal meat production.

Effect of immunization with a recombinant cholera toxin B subunit/somatostatin fusion protein on immune response and growth hormone levels in mice.

Somatostatin (SS) is a hormone that inhibits growth hormone secretion. Cholera toxin B subunit (CTB) is a widely used adjuvant to improve the immunogenicity of co-administrated antigen. To block the growth hormone-inhibiting effect of SS, a fusion gene of CTB and SS was constructed and expressed in Escherichia coli. The purified CTB/SS fusion protein polymerized into a biologically active pentamer required for CTB binding to the GM1 ganglioside receptor. Immunization with the CTB/SS protein induced specific immunity against CTB and SS in mice. The serum growth hormone of the CTB/SS-treated mice increased by 29 % (P < 0.05) compared with the control. The results indicated that the CTB/SS fusion protein was effective in inducing immune response against SS as well as elevating the growth hormone level.

Effect of active immunization against a recombinant mouse granulocyte-macrophage colony-stimulating factor/somatostatin fusion protein on the growth of mice.

Female BALB/c mice were actively immunized subcutaneously with a recombinant protein of granulocyte-macrophage colony-stimulating factor (GM-CSF) fused with somatostatin (SS) (GM-CSF/SS). Fifty-four days after the primary immunization, the body weight of the immunized mice increased by 4.62% compared with the control (P < 0.05), together with the induction of detectable serum antibodies against SS. The level of serum growth hormone (GH) elevated by 44.54% (P < 0.05) and the mRNA expression of muscular IGF-1 increased by 94% for the GM-CSF/SS-treated mice. The results indicated that the recombinant protein GM-CSF/SS was efficient in inducing specific immunity against SS, subsequently leading to the increase of the GH level by SS neutralization, and ultimately improving the growth of mice.

Effect of the C-terminal domains and terminal residues of catalytic domain on enzymatic activity and thermostability of lichenase from Clostridium thermocellum.

To elucidate the effects of C-terminal domains of LicMB (mature lichenase from Clostridium thermocellum) and terminal residues of LicMB-CD (catalytic domain of LicMB) on the properties of lichenase, a series of truncated genes were constructed and expressed in E. coli. The Thr-Pro box had a positive effect while the dockerin domain had a negative impact on the properties of LicMB. The N-terminal 10-25th and C-terminal 1-9th residues of LicMB-CD were necessary to retain high thermostability while the N-terminal 1-7th and C-terminal 1-3rd residues were not necessary to maintain enzymatic activity.