Data Mining Suggests That CXCL14 Gene Silencing in Colon Cancer Is Due to Promoter Methylation
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PROVIDER: S-EPMC10671198 | biostudies-literature | 2023 Nov
REPOSITORIES: biostudies-literature
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Project description:Recent transgenic studies on L1 retrotransposons have afforded exciting insights into L1 biology, and a unique opportunity to model their function and regulation in vivo. Thus far, the majority of the transgenic L1 mouse lines are constructed via pronuclear microinjection, a procedure that typically results in the integration of tandem arrayed transgenes. Transgene arrays are susceptible to repeat-induced gene silencing (RIGS) in both plants and animals. In order to examine the potential impact of RIGS on L1 retrotransposition, we derived a cohort of animals carrying reduced copies of ORFeus transgene at the same genomic locus by Cre-mediated recombination. The copy number reduction of ORFeus transgenes did not decrease the overall retrotransposition activity. Using a sensitive and reproducible quantitative PCR assay, an average frequency of 0.45 insertions per cell was observed for animals carrying the donor transgene at a single copy, representing a 9-fold increase of retrotransposition frequency on a per-copy basis. DNA methylation analyses revealed that the observed retrotransposition activity was correlated with differential CpG methylation at the heterologous promoter: the promoter region was largely methylated in animals with the high-copy array but significantly hypomethylated in animals with the single-copy counterpart. In contrast, the ORF2 region, which represents the body of the ORFeus transgene, and the 3' end of the transgene showed high level of methylation in both high-copy and single-copy samples. The observed methylation patterns were metastable across generations. In summary, our data suggest that tandem arrayed L1 transgenes are subject to RIGS, and transgenes present at a single copy in the genome are thus recommended for modeling L1 in animals.
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Project description:The present study was planned to explore the correlation between the methylation of APC (adenomatous polyposis coli) and colon carcinogenesis. Colon cancer tissues and tumor-adjacent normal tissues of 60 colon cancer patients (who received surgical operation in our hospital from January 2012 to December 2014) were collected. SW1116 cells in human colon cancer tissues were selected for culturing. 5-aza-2c-deoxycytidine (5-aza-dC) was utilized as an inhibitor of the methylation for APC gene. Methylation specific PCR (MSP) was utilized for detection of APC methylation in SW1116 cells. The MTT and Transwell assays were performed to detect the effect of the methylation of APC gene on the proliferation and invasive abilities of SW1116 cells. The correlation between the methylation of APC gene and pathological parameters of colon cancer patients was analyzed. MSP results revealed that 41 cases (68.33%) showed methylation of APC gene in colon cancer tissues. No methylation of APC gene was found in tumor-adjacent normal tissues. 5-aza-dC was able to inhibit the methylation of APC gene in SW1116 cells. APC gene methylation was correlated with tumor size, differentiation degree, lymph node metastasis and Dukes staging. In conclusion, the levels of the methylation of APC in colon cancer tissues and SW1116 cells are relatively high. The methylation of APC promoted the proliferation and invasion abilities of SW1116 cells. Furthermore, methylation is correlated with a variety of clinicopathological features of colon cancer patients.
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Project description:Targeted mutations in mouse disrupt local chromatin structure and may lead to unanticipated local effects. We evaluated targeted gene promoter silencing in a group of six mutants carrying the tm1a Knockout Mouse Project allele containing both a LacZ reporter gene driven by the native promoter and a neo selection cassette. Messenger RNA levels of the reporter gene and targeted gene were assessed by qRT-PCR, and methylation of the promoter CpG islands and LacZ coding sequence were evaluated by sequencing of bisulfite-treated DNA. Mutants were stratified by LacZ staining into presumed Silenced and Expressed reporter genes. Silenced mutants had reduced relative quantities LacZ mRNA and greater CpG Island methylation compared with the Expressed mutant group. Within the silenced group, LacZ coding sequence methylation was significantly and positively correlated with CpG Island methylation, while promoter CpG methylation was only weakly correlated with LacZ gene mRNA. The results support the conclusion that there is promoter silencing in a subset of mutants carrying the tm1a allele. The features of targeted genes which promote local silencing when targeted remain unknown.
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Project description:BackgroundExpanded GGGGCC hexanucleotide repeats located in the noncoding region of the chromosome 9 open reading frame 72 (C9orf72) gene represent the most common genetic abnormality for familial and sporadic amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Formation of nuclear RNA foci, accumulation of repeat-associated non-ATG-translated dipeptide-repeat proteins, and haploinsufficiency of C9orf72 are proposed for pathological mechanisms of C9ALS/FTD. However, at present, the physiological function of C9orf72 remains largely unknown.MethodsBy searching on a bioinformatics database named COXPRESdb composed of the comprehensive gene coexpression data, we studied potential C9orf72 interactors.ResultsWe identified the ATP/GTP binding protein 1 (AGTPBP1) gene alternatively named NNA1 encoding a cytosolic carboxypeptidase whose mutation is causative of the degeneration of Purkinje cells and motor neurons as the most significant gene coexpressed with C9orf72. We verified coexpression and interaction of AGTPBP1 and C9orf72 in transfected cells by immunoprecipitation and in neurons of the human brain by double-labeling immunohistochemistry. Furthermore, we found a positive correlation between AGTPBP1 and C9orf72 mRNA expression levels in the set of 21 human brains examined.ConclusionsThese results suggest that AGTPBP1 serves as a C9orf72 interacting partner that plays a role in the regulation of neuronal function in a coordinated manner within the central nervous system.
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Project description:Loss of 3p11-p14 is a frequent event in epithelial cancer and a candidate prognostic biomarker in cervical cancer. In addition to loss, promoter methylation can participate in gene silencing and promote tumor aggressiveness. We have performed a complete mapping of promoter methylation at 3p11-p14 in two independent cohorts of cervical cancer patients (n = 149, n = 121), using Illumina 450K methylation arrays. The aim was to investigate whether hyperm-ethylation was frequent and could contribute to gene silencing and disease aggressiveness either alone or combined with loss. By comparing the methylation level of individual CpG sites with corresponding data of normal cervical tissue, 26 out of 41 genes were found to be hypermethylated in both cohorts. The frequency of patients with hypermethylation of these genes was found to be higher at tumor stages of 3 and 4 than in stage 1 tumors. Seventeen of the 26 genes were transcriptionally downregulated in cancer compared to normal tissue, whereof 6 genes showed a significant correlation between methylation and expression. Integrated analysis of methylation, gene dosage, and expression of the 26 hypermethylated genes identified 3 regulation patterns encompassing 8 hypermethylated genes; a methylation driven pattern (C3orf14, GPR27, ZNF717), a gene dosage driven pattern (THOC7, PSMD6), and a combined methylation and gene dosage driven pattern (FHIT, ADAMTS9, LRIG1). In survival analysis, patients with both hypermethylation and loss of LRIG1 had a worse outcome compared to those harboring only hypermethylation or none of the events. C3orf14 emerged as a novel methylation regulated suppressor gene, for which knockdown was found to promote invasive growth in human papilloma virus (HPV)-transformed keratinocytes. In conclusion, hypermethylation at 3p11-p14 is common in cervical cancer and may exert a selection pressure during carcinogenesis alone or combined with loss. Information on both events could lead to improved prognostic markers.
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