<Emphasis Type="Italic">FAT4</Emphasis> hypermethylation and grade dependent downregulation in gastric adenocarcinoma

Gastric cancer is one of the major causes of death due to cancer in the world. It is a multi-factorial disease with epigenetic factors being also involved in its development. FAT4 is a tumor suppressor gene exerting an important role in cell adhesion. This study aimed at analyzing FAT4 expression and promoter methylation in gastric cancer. FAT4 expression was studied in 30 tumoral tissues and their non-tumoral counterparts using Taqman real time PCR method. Promoter methylation was assessed using bisulfite conversion method followed by sequencing. Tumor tissues showed reduced FAT4 expression (P = 0.04). FAT4 downregulation was associated with tumor grade, with higher repression at advanced grades. Significant increase of promoter methylation was observed in tumoral tissues. Reduced expression of FAT4 and increased methylation of its promoter may be one of the effective processes in turning a healthy stomach tissue into a tumor tissue.     

Prognostic potential of <Emphasis Type="Italic">KLOTHO</Emphasis> and <Emphasis Type="Italic">SFRP1</Emphasis> promoter methylation in head and neck squamous cell carcinoma

Hypermethylation in the CpG island promoter regions of tumor suppressors is known to play a significant role in the development of HNSCC and the detection of which can aid the classification and prognosis of HNSCC. This study aims to profile the methylation patterns in a panel of key genes including CDKN2A, CDKN2B, KLOTHO (KL), RASSF1A, RARB, SLIT2, and SFRP1, in a group of HNSCC samples from Saudi Arabia. The extent of methylation in these genes is determined using the MethyLight assay and correlated with known clinicopathological parameters in our samples of 156 formalin-fixed and paraffin-embedded HNSCC tissues. SLIT2 methylation had the highest frequency (64.6%), followed by RASSF1A (41.3%), RARB (40.7%), SFRP1 (34.9), KL (30.7%), CKDN2B (29.6%), and CKDN2A (29.1%). KL and SFRP1 methylation were more predominant in nasopharyngeal tumors (P = 0.001 and P = 0.031 respectively). Kaplan Meier analysis showed that patients with moderately differentiated tumors who display SFRP1 methylation have significantly worse overall survival in comparison with other samples. In contrast, better clinical outcomes were seen in patients with KL methylation. In conclusion, our findings suggest that the detection of frequent methylation in SFRP1 and KL genes’ promoters could serve as prognostic biomarkers for HNSCC.     

<Emphasis Type="Italic">Braf</Emphasis>, <Emphasis Type="Italic">Kras</Emphasis> and <Emphasis Type="Italic">Helicobacter pylori</Emphasis> epigenetic changes-associated chronic gastritis in Egyptian patients with and without gastric cancer

We aimed to study MLH1 and MGMT methylation status in Helicobacter pylori-associated chronic gastritis in Egyptian patients with and without gastric cancer. 39 patients were included in our study. They were divided into 2 groups; patients without (group I) and with gastric adenocarcinoma (group II). Patients were subjected to clinical examination, abdominal ultrasound and upper endoscopy for gastric biopsy. Biopsies were subjected to urease test, histological examination, and DNA purification. H. pylori, Braf, Kras, MLH1 and MGMT methylation were assessed by quantitative PCR. DNA sequencing was performed to assess Braf and Kras genes mutation. qPCR of H. pylori was significantly higher in patients with adenocarcinoma (group II) than those without adenocarcinoma (group I); with a p < 0.001 as well as in patients with age above 50 years with a p value = 0.008. By applying logistic regression analysis it was reported that the H. pylori qPCR is a significant predictor to the adenocarcinoma with OR = 1.025 (95 % CI: 1. 002–1.048), with sensitivity of 90 % and specificity of 100 %. Adenocarcinoma patients had a significantly higher mean age and levels of H. Pylori, Braf, K-ras, methylated MGMT and methylated MLH1 than those of gastritis patients. DNA sequence analysis of Braf (codon 12) and Kras (codon 600) had genes mutation in gastric adenocarcinoma versus chronic gastritis. Conclusion: H. pylori may cause epigenetic changes predisposing the patients to cancer stomach. Estimation of H. pylori by qPCR can be a good predictor to adenocarcinoma. Braf and Kras genes mutation were reveled in gastritis and adenocarcinoma patients.     

Opioid Exposure is Associated with Aberrant DNA Methylation of <Emphasis Type="Italic">OPRM1</Emphasis> Promoter Region in a Chinese Han Population

The μ-opioid receptor (OPRM1) plays an important role in opiate addiction. The OPRM1 gene promoter showed hypermethylation in lymphocytes of opiate addicts as well as opioid medications users, while the methylation status displayed ethnic diversity. The purpose of the study was to investigate the methylation pattern of OPRM1 promoter in the Han Chinese population. We analyzed 22 CpG sites located in OPRM1 promoter in 186 former opiate addicts (94 males and 92 females) and 184 healthy controls (102 males and 82 females). The +?126 CpG site was significantly hypermethylated in the former heroin addicts compared with controls (13.67% versus 8.39%, \(P = 3.78 \times 10^{ - 9}\), corrected for 36 tests). Six CpG sites were significantly associated with opioid exposure, including the most significant +126 CpG site (opiate addicts 13.57%, control 8.39%, \(P = 9.19 \times 10^{ - 12}\), corrected for 36 tests), while the +23 GpG site was the only hypomethylated one in former opiate addicts compared with controls (P?=?0.0023 after Bonferroni correction). Our results supported that opioid exposure was associated with methylation status of OPRM1 promoter and showed ethnic dependence.     

Identification and Functional Analysis of Two Cotton Orthologs of <Emphasis Type="Italic">MAX2</Emphasis> Which Control Shoot Lateral Branching

Cotton (Gossypium spp.), as the most important fiber and oilseed crop in the world, is extremely important for the industry. However, due to its indeterminate growth habit and complex branching system, massive labor costs are needed for shoot apex removal and branch pruning during cotton production. Therefore, it is very important to explore branch-controlling genes and genetically modify the branch architecture of cotton. Strigolactones (SLs) are a novel class of plant hormone that inhibit the outgrowth of lateral branches. To elucidate the role of SLs in branch development of cotton, we cloned and characterized GhMAX2a and GhMAX2b from tetraploid upland cotton (Gossypium hirsutum), the orthologs of Arabidopsis MAX2, rice D3, and petunia RMS4. GhMAX2a/2b was ubiquitously expressed in all tested tissues of cotton, with relatively higher expression levels in leaves and lateral buds. Subcellular localization assay showed that the GhMAX2-GFP fusion protein localized to the nucleus. Both GhMAX2a and GhMAX2b can fully rescue the dwarfed and highly branched phenotypes of the Arabidopsis max2-1 mutant, indicating that GhMAX2s have conserved functions with that of AtMAX2. The cotton GhMAX2b interacted with Arabidopsis Skp1-like 1 (ASK1) proteins in vitro which was further confirmed in the Arabidopsis protoplasts using the co-immunoprecipitation assay, indicating that GhMAX2b probably functions through forming an SCF E3 complex with Skp and other proteins in the Arabidopsis. These results suggest that the cotton GhMAX2s encode functional MAX2 that can inhibit the shoot lateral branching. Further functional analysis of GhMAX2s in determining cotton branch architecture and yield is underway.     

Inhibition of breast cancer cell proliferation and tumorigenesis by long non-coding RNA <Emphasis Type="Italic">RPPH1</Emphasis> down-regulation of miR-122 expression

Background

Recent studies showed that long non-coding RNA (lncRNA) plays an important role in many life activities. RPPH1 is one of the lncRNA genes that are expressed differently between breast cancer and normal tissues by the lncRNA gene chip. Our study was conducted to examine the regulation of lncRNA RPPH1 in breast cancer.

Methods

Two cell lines, MCF-7 and MDA-MB-231, were selected to be the research objects in this study; RPPH1 overexpression and knockdown models were established by transforming vectors. Real-time polymerase chain reaction, MTT assay, clone formation and cell flow cytometer assay were used to test the function of RPPH1. Dual-luciferase assay was used to detect a target relationship between RPPH1 and miR-122.

Results

RPPH1 overexpression promoted cell cycle and proliferation and increased colony formation. In the RPPH1 overexpression model, there was a target relationship between RPPH1 and miR-122, and some of the downstream genes of miR-122, including ADAM10, PKM2, NOD2 and IGF1R, were increased. Moreover, we found that lentivirus-mediated interference of lncRNA RPPH1 inhibited tumour growth in nude mice.

Conclusion

Breast cancer progression can be promoted by directly targeting miR-122 through lncRNA RPPH1. This study provided evidence that can serve as the molecular basis for improving treatment options for patients.

     

Polymorphic Imprinting of <Emphasis Type="Italic">SLC38A4</Emphasis> Gene in Bovine Placenta

Imprinted genes are characterized by monoallelic expression that is dependent on parental origin. Comparative analysis of imprinted genes between species is a powerful tool for understanding the biological significance of genomic imprinting. The slc38a4 gene encodes a neutral amino acid transporter and is identified as imprinted in mice. In this study, the imprinting status of SLC38A4 was assessed in bovine adult tissues and placenta using a polymorphism-based approach. Results indicate that SLC38A4 is not imprinted in eight adult bovine tissues including heart, liver, spleen, lung, kidney, muscle, fat, and brain. It was interesting to note that SLC38A4 showed polymorphic status in five heterogeneous placentas, with three exhibiting paternal monoallelic expression and two exhibiting biallelic expression. Monoallelic expression of imprinted genes is generally associated with allele-specific differentially methylation regions (DMRs) of CpG islands (CGIs)-encompassed promoter; therefore, the DNA methylation statuses of three CGIs in the SLC38A4 promoter and exon 1 region were tested in three placentas (two exhibiting paternal monoallelic and one showing biallelic expression of SLC38A4) and their corresponding paternal sperms. Unexpectedly, extreme hypomethylation (<?3%) of the DNA was observed in all the three detected placentas and their corresponding paternal sperms. The absence of DMR in bovine SLC38A4 promoter region implied that DNA methylation of these three CGIs does not directly or indirectly affect the polymorphic imprinting of SLC38A4 in bovine placenta. This suggested other epigenetic features other than DNA methylation are needed in regulating the imprinting of bovine SLC38A4, which is different from that of mouse with respect to a DMR existence at the mouse’s slc38a4 promoter region. Although further work is needed, this first characterization of polymorphic imprinting status of SLC38A4 in cattle placenta provides valuable information on investigating the genomic imprinting phenomenon itself.     

Epigenetic and genetic variation in <Emphasis Type="Italic">GATA5</Emphasis> is associated with gastric disease risk

Gastric cancer incidence varies considerably among populations, even those with comparable rates of Helicobacter pylori infection. To test the hypothesis that genetic variation plays a role in gastric disease, we assessed the relationship between genotypes and gastric histopathology in a Colombian study population, using a genotyping array of immune-related single nucleotide polymorphisms (SNPs). Two synonymous SNPs (rs6061243 and rs6587239) were associated with progression of premalignant gastric lesions in a dominant-effects model after correction for multiple comparisons (p = 2.63E?07 and p = 7.97E?07, respectively); effect sizes were β = ?0.863 and β = ?0.815, respectively, where β is an estimate of effect on histopathology scores, which ranged from 1 (normal) to 5 (dysplasia). In our replication cohort, a second Colombian population, both SNPs were associated with histopathology when additively modeled (β = ?0.256, 95 % CI = ?0.47, ?0.039; and β = ?0.239, 95 % CI = ?0.45, ?0.024), and rs6587239 was significantly associated in a dominant-effects model (β = ?0.330, 95 % CI = ?0.66, 0.00). Because promoter methylation of GATA5 has previously been associated with gastric cancer, we also tested for the association of methylation status with more advanced histopathology scores in our samples and found a significant relationship (p = 0.001). A multivariate regression model revealed that the effects of both the promoter methylation and the exonic SNPs in GATA5 were independent. A SNP-by-methylation interaction term was also significant. This interaction between GATA5 variants and GATA5 promoter methylation indicates that the association of either factor with gastric disease progression is modified by the other.