The Effect of the Cyp19a1 Gene Methylation Modification on Temperature-dependent Sex Determination of Reeves' Turtle(Mauremys reevesii)

Temperature-dependent sex determination（TSD） is a type of environmental sex determination in which the sex of the embryos depends on the ambient temperature; however,the molecular mechanisms governing this process remain unknown.Aromatase,encoded by the cyp19a1 gene,which converts androgens into estrogens in animals,was considered to be the key gene for TSD.In this study,the 5＇-flanking region of the cyp19a1 gene in Reeves＇ turtle（Mauremys reevesii） was cloned,and the promoter region was identified using the luciferase reporter assay.Then the eggs of Reeves＇ turtle were incubated at different temperatures（26°C： male-biased temperature; 29°C： non-sex-biased temperature and 32°C： female-biased temperature）.During the thermosensitive period,the adrenal kidney gonad complexes（AKG） were sampled.DNA methylation analysis of the AKG samples showed that the promoter region of the cyp19a1 gene was significantly de-methylated in the female-biased temperature regime（P＆lt;0.01）.Quantitative analysis of the cyp19a1 gene and estrogen by q PCR and Elisa assay showed that the expression level of the cyp19a1 gene and estrogen content were both upregulated significantly at the female-biased temperature（P＆lt;0.01）.These results indicated that the de-methylation response of the cyp19a1 gene to incubation temperature,especially at the female-biased temperature,could lead to temperature-specific expression of aromatase and increased estrogen levels,which may further determine gonadal development in Reeves＇ turtle.These findings provide insights into the genetic mechanisms underlying TSD.

DNA methylation in poultry:a review

As an important epigenetic modification,DNA methylation is involved in many biological processes such as animal cell differentiation,embryonic development,genomic imprinting and sex chromosome inactivation.As DNA methylation sequencing becomes more sophisticated,it becomes possible to use it to solve more zoological problems.This paper reviews the characteristics of DNA methylation,with emphasis on the research and application of DNA methylation in poultry.

Genic C-Methylation in Soybean Is Associated with Gene Paralogs Relocated to Transposable Element-Rich Pericentromeres

Most plants are polyploid due to whole-genome duplications （WGD） and can thus have duplicated genes. Following a WGD, paralogs are often fractionated （lost） and few duplicate pairs remain. Little attention has been paid to the role of DNA methylation in the functional divergence of paralogous genes. Using high- resolution methylation maps of accessions of domesticated and wild soybean, we show that in soybean, a recent paleopolyploid with many paralogs, DNA methylation likely contributed to the elimination of ge- netic redundancy of polyploidy-derived gene paralogs. Transcriptionally silenced paralogs exhibit partic- ular genomic features as they are often associated with proximal transposable elements （TEs） and are pref- erentially located in pericentromeres, likely due to gene movement during evolution. Additionally, we provide evidence that gene methylation associated with proximal TEs is implicated in the divergence of expression profiles between orthologous genes of wild and domesticated soybean, and within populations.

5-aza-2'-deoxycitydine induces demethylation and up-regulates transcription of p16^(INK4A) gene in human gastric cancer cell lines

Background To investigate the effects of DNA methylation on the expression of tumor-associated genes and the cell cycle in human gastric cancer cells. Methods Two gastric cancer cell lines (MKN-45 and HGC-27) were treated with DNA methyltransferase (DNMT) inhibitor,5-aza-2’-deoxycytidine (5-aza-dC). The expressions of p16 INK4A,p21 WAF1,p53 , p73 ,c-Ha-ras and c-myc genes mRNA were detected by using reverse transcription PCR (RT-PCR). DNA methylation status of p16 INK4A gene promoter was assayed by bisulfite modification and sequencing. The cell cycle was analyzed by using flow cytometry (FCM). Results 5-aza-dC induced the demethylation of p16 INK4A gene promoter. The expression of p16 INK4A mRNA was obviously up-regulated by treatment with 10 μmol/L (MKN-45 cells) or 5 μmol/L (HGC-27 cells) of 5-aza-dC for 24 hours. However,5-aza-dC treatment failed to regulate the expressions of p21 WAF1,p53 , p73 ,c-Ha-ras and c-myc genes in MKN-45 and HGC-27 cells. Furthermore,5-aza-dC induced the cell cycle arrest in G1 phase in HGC-27 cell,but not in MKN-45 cell. Conclusions DNA methylation regulates the transcription of p16 INK4A but not p21 WAF1 and proto-oncogenes in human gastric cancer cell lines MKN-45 and HGC-27.

The exploration of the correlation between the risk of obesity and the promoter methylation of PRDM16 gene

Objective To explore the association between the Cp G methylation level of positive regulatory domain containing 16(PRDM16)gene promoter and obesity or body mass index(BMI).Methods A total of 116 patients(91female adults and 25 male adults)with abdominal operation in a municipal hospital of Henan province were enrolled in this study and they were divided into

Efficacy of Yisui granule(益髓颗粒)on myelodysplastic syndromes in SKM-1 mouse xenograft model through suppressing Wnt/β-catenin signaling pathway

OBJECTIVE:To unmask the underlying mechanisms of Yisui granule(益髓颗粒,YSG)for the treatment of Myelodysplastic syndromes(MDS).METHODS:Our study used an SKM-1 mouse xenograft model of MDS to explore the anti-tumor potential of YSG and its safety,assess its effect on overall survival(OS),and evaluate whether its mechanism is associated with the demethylation of the secreted frizzled related protein 5(s FRP5)gene and suppressing Wnt/β-catenin pathway.Bisulfite amplicon sequencing was applied to detect the level of methylation of the s FRP5 gene;western blotting,immunofluorescence staining,and real-time Polymerase Chain Reaction were performed to detect DNA methyltransferase 1(DNMT1),s FRP5,and other Wnt/β-catenin pathway-related m RNA and protein expression.RESULTS:The results showed that high-dosage YSG exerted an anti-tumor effect similar to that of decitabine,improved OS,and reduced long-term adverse effects in the long term.Mechanically,YSG reduced the expression of DNMT1 methyltransferase,decreased the methylation,and increased the expression of the Wnt/β-catenin pathway antagonist-s FRP5.Furthermore,components of the Wnt/β-catenin pathway,including Wnt3a,β-catenin,c-Myc,and cyclin D1,were down-regulated in response to YSG,suggesting that YSG could treat MDS by demethylating the s FRP5 gene and suppressing the Wnt/β-catenin pathway.CONCLUSIONS:Our findings demonstrated that YSG could be used alone or in combination with decitabine to improve outcomes in the MDS animal model,providing an alternative solution for treating MDS.

Specifications of Targeting Heterochromatin Modifications in Plants

Plants encode a diverse repertoire of DNA methyltransferases that have specialized to target cytosines for methylation in specific sequence contexts. These include the de novo methyltransferase, DOMAINS REARRANGED METHYLTRANSFERASE 2 （DRM2）, which methylates cytosines in all sequence contexts through an RNA-guided process, the CHROMOMETHYLASES （CMTs）, which methylate CHH and CHG cytosines （where H is A, T, or C）, and METHYLTRANSFERASE 1 （MET1）, which maintains methylation of symmetrical CG contexts. In this review, we discuss the sequence specificities and targeting of each of these pathways. In particular, we highlight recent studies that indicate CMTs preferentially target CWG or CWA/CAW motifs （where W is A or T）, and discuss how self-reinforcing feedback loops between DNA methyltransferases and histone modifications characteristic of heterochromatin specify targeting. Finally, the initiating events that lead to gene body methylation are discussed as a model illustrating how interde- pendent targeting of different silencing pathways can potentiate the establishment of off-target epialleles.