Targeting aberrant colon cancer-specific DNA methylation with lipoteichoic acid-deficient Lactobacillus acidophilus

Pathogenic autoinflammatory responses triggered by dysregulated microbial interactions may lead to intestinal disorders and malignancies. Previously, we demonstrated that a lipoteichoic acid (LTA)-deficient Lactobacillus acidophilus strain, NCK2025, ameliorated inflammation-induced colitis, significantly reduced the number of polyps in a colonic polyposis cancer model and restored physiological homeostasis in both cases. Nonetheless, the regulatory signals delivered by NCK2025 to reprogram the gastrointestinal microenvironment, and thus resist colonic cancer progression, remain unknown. Accumulating evidence suggest that epigenetic changes, in the presence and absence of pathogenic inflammation, can result in colorectal cancer (CRC). To test possible epigenetic modifications induced by NCK2025, the expression of epigenetically regulated, CRC-associated genes was measured with and without bacterial treatment. In vivo and in vitro, NCK2025 enhanced the expression of tumor suppressor genes that may regulate CRC development. Therefore, differential epigenetic regulation of CRC-related genes by NCK2025 represents a potential therapy against colitis-associated and sporadic CRC.     

Antisilencing role of the RNA-directed DNA methylation pathway and a histone acetyltransferase in Arabidopsis

REPRESSOR OF SILENCING 1 (ROS1) is a DNA demethylation enzyme that was previously identified during a genetic screen for the silencing of both RD29A-LUC and 35S-NPTII transgenes on a T-DNA construct. Here we performed a genetic screen to identify additional mutants in which the 35S-NPTII transgene is silenced. We identified several alleles of ros1 and of the following components of the RNA-directed DNA methylation (RdDM) pathway: NRPD1 (the largest subunit of polymerase IV), RDR2, NRPE1 (the largest subunit of polymerase V), NRPD2, AGO4, and DMS3. Our results show that the silencing of 35S-NPTII in the RdDM pathway mutants is due to the reduced expression of ROS1 in the mutants. We also identified a putative histone acetyltransferase (ROS4) from the genetic screen. The acetyltransferase contains a PHD-finger domain that binds to unmethylated histone H3K4. The mutation in ROS4 led to reduction of H3K18 and H3K23 acetylation levels. We show that the silencing of 35S-NPTII and some transposable element genes was released by the ddm1 mutation but that this also required ROS4. Our study identifies a unique antisilencing factor, and reveals that the RdDM pathway has an antisilencing function due to its role in maintaining ROS1 expression.     

应用MS-RDA筛选胃癌甲基化差异片段

目的:应用甲基化敏感性代表性差异分析法(methylation-sensitive representational difference analysis,MS-RDA)筛选胃癌和正常胃组织间甲基化差异DNA片段．方法:通过MS-RDA筛选胃癌和正常胃组织间DNA甲基化差异片段,经克隆、测序后进行生物信息学分析．结果:获得3个甲基化差异片段,分别为 CRS1308,CRS1309,CRS1310k列,其中 CRS1309和CRS1310已被GenBank收录,登陆亏分别为AY887106和AY887107,CRS1309 序列与LOC440683基因第11外显子、 LOC440887基因的3’端,DRD5基因启动子和外显子区域均有很高的相似性(分别为98％,99％, 94％),CRS1310序列与1999年Minoru Toyota在人类结肠癌中分离出的核糖体RNA上的甲基化差异性CpG岛有很高的相似性(98％)．结论:胃癌和正常胃组织间DNA甲基化存在差异,MS-RDA可有效分析这两种不同组织间甲基化的差异,筛选出有意义的甲基化差异片段.     

Association of tissue-specific differentially methylated regions (TDMs) with differential gene expression

Early studies proposed that DNA methylation could have a role in regulating gene expression during development [Riggs, A.D. (1975) Cytogenet. Cell Genet. 14, 9-25]. However, some studies of DNA methylation in known tissue-specific genes during development do not support a major role for DNA methylation. In the results presented here, tissue-specific differentially methylated regions (TDMs) were first identified, and then expression of genes associated with these regions correlated with methylation status. Restriction landmark genomic scanning (RLGS) was used in conjunction with virtual RLGS to identify 150 TDMs [Matsuyama, T., Kimura, M.T., Koike, K., Abe, T., Nakao, T., Asami, T., Ebisuzaki, T., Held, W.A., Yoshida, S. & Nagase, H. (2003) Nucleic Acids Res. 31, 4490-4496]. Analysis of 14 TDMs by methylation-specific PCR and by bisulfite genomic sequencing confirms that the regions identified by RLGS are differentially methylated in a tissue-specific manner. The results indicate that 5% or more of the CpG islands are TDMs, disputing the general notion that all CpG islands are unmethylated. Some of the TDMs are within 5' promoter CpG islands of genes, which exhibit a tissue-specific expression pattern that is consistent with methylation status and a role in tissue differentiation.     

DNA甲基化与胃癌的关系研究

基因的表观遗传学修饰是一种不依赖DNA序列改变的可逆的基因表达调控过程。主要包括DNA甲基化与去甲基化,这些可逆的表观遗传学修饰可以方便地关闭和开放某些特定基因,有选择性表达基因组的信息,在维持染色体结构的稳定、基因组的完整、组织特异性基因的表达调节、细胞分化和个体成长等过程中发挥着至关重要的作用,而DNA的甲基化与胃癌的关系日益受到重视,为胃癌的诊断和治疗提供了新的途径。这些分子病理基础研究的不断深入使我们更好地理解发病机制,进而能够为临床治疗提供更好的借鉴价值。     

Function of the DEMETER DNA glycosylase in the Arabidopsis thaliana male gametophyte

In double fertilization, the vegetative cell of the male gametophyte (pollen) germinates and forms a pollen tube that brings to the female gametophyte two sperm cells that fertilize the egg and central cell to form the embryo and endosperm, respectively. The 5-methylcytosine DNA glycosylase DEMETER (DME), expressed in the central cell, is required for maternal allele demethylation and gene imprinting in the endosperm. By contrast, little is known about the function of DME in the male gametophyte. Here we show that reduced transmission of the paternal mutant dme allele in certain ecotypes reflects, at least in part, defective pollen germination. DME RNA is detected in pollen, but not in isolated sperm cells, suggesting that DME is expressed in the vegetative cell. Bisulfite sequencing experiments show that imprinted genes (MEA and FWA) and a repetitive element (Mu1a) are hypomethylated in the vegetative cell genome compared with the sperm genome, which is a process that requires DME. Moreover, we show that MEA and FWA RNA are detectable in pollen, but not in isolated sperm cells, suggesting that their expression occurs primarily in the vegetative cell. These results suggest that DME is active and demethylates similar genes and transposons in the genomes of the vegetative and central cells in the male and female gametophytes, respectively. Although the genome of the vegetative cell does not participate in double fertilization, its DME-mediated demethylation is important for male fertility and may contribute to the reconfiguration of the methylation landscape that occurs in the vegetative cell genome.     

Isolation and bioinformatics analysis of differentially methylated genomic fragments in human gastric cancer

AIM：To isolate and analyze the DNA sequences which are methylated differentially between gastric cancer and normal gastric mucosa.
METHODS： The differentially methylated DNA sequences between gastric cancer and normal gastric mucosa were isolated by methylation-sensitive representational difference analysis （MS-RDA）. Similarities between the separated fragments and the human genomic DNA were analyzed with Basic Local Alignment Search Tool （BLAST）.
RESULTS： Three differentially methylated DNA sequences were obtained, two of which have been accepted by GenBank. The accession numbers are AY887106 and AY887107. AY887107 was highly similar to the 11th exon of LOC440683 （98%）, 3＇ end of LOC440887 （99%）, and promoter and exon regions of DRD5 （94%）. AY887106 was consistent （98%） with a CpG island in ribosomal RNA isolated from colorectal cancer by Minoru Toyota in 1999.
CONCLUSION： The methylation degree is different between gastric cancer and normal gastric mucosa. The differentially methylated DNA sequences can be isolated effectively by MS-RDA.     

MicroRNA和DNA甲基化在肺癌诊断中的研究进展

小分子核糖核酸（MicroRNA）作为肺癌早期的一种新的生物标志物,为肿癌的诊断提供了一种新的方法。MicroRNA在肿瘤发生发展过程中发挥癌基因和抑癌基因的作用。MicroRNA与肺癌增殖、侵袭、治疗、预后和复发有着密切关系。DNA甲基化是目前最主要的一种表观遗传修饰形式,抑癌基因的高甲基化使DNA染色质构象发生改变导致转录失活,引起肿瘤发生。DNA甲基化和MicroRNA在功能上相互调控,对肺癌的发生、发展同样重要。     

The role of cell-free DNA in predicting colorectal cancer prognosis

Introduction: Colorectal cancer is a cancer of the digestive system with poor prognosis. Cell-free DNA has received much attention with its unique predominance, especially in colorectal cancer.

Areas covered: This study has summarized recent advancements and challenges regarding cell-free DNA in predicting CRC prognosis. Furthermore, the authors make predictions on the potential developments concerning cell-free DNA in future prognosis prediction techniques.

Expert commentary: Cell-free DNA has the value of predicting CRC prognosis as an important biomarke. Further clinical trials should be performed to promote translating cell-free DNA into clinical applications.     

DNA甲基转移酶3A基因在胃癌中的表达及其临床意义

目的 研究DNA甲基转移酶3A（DNMT3A）在天津汉族人群胃癌（GC）组织中的表达,并分析DNMT3A的表达与胃癌临床病理因素之间的关系,探讨其在胃癌发生发展中的作用.方法 用免疫组织化学法检测DNMT3A在130例胃癌组织及256例胃炎组织中的表达,将两者进行对比并分析其与胃癌临床病理之间的关系.结果 DNMT3A蛋白在胃癌组织中的阳性细胞表达率（78.4％）明显高于胃炎组织的表达率（64.4％）,两者间比较差异有统计学意义（P＜0.05）.DNMT3A蛋白的异常表达与胃癌患者年龄、性别、肿瘤大小、浸润深度无明显相关,与组织分化程度、淋巴结转移及TNM分期密切相关（P＜0.05）.结论 在胃癌组织中,DNMT3A蛋白呈高表达状态,并与肿瘤组织分化、淋巴结转移及TNM分期有关,DNMT3A的高表达可能与胃癌的发生、发展有关,且可能是胃癌发生的早期诊断指标.