BACKGROUNDDNA hypomethylation of long interspersed nuclear elements-1 (LINEs-1) occurs during carcinogenesis, whereas information addressing LINE-1 methylation in Wilms tumor (WT) is limited. The main purpose of our study was to quantify LINE-1 methylation levels and evaluate their relationship with relative telomere length (TL) in WT.

METHODSWe investigated LINE-1 methylation and relative TL using bisulfite-polymerase chain reaction (PCR) pyrosequencing and quantitative PCR, respectively, in 20 WT tissues, 10 normal kidney tissues and a WT cell line. Significant changes were analyzed by t-tests.

RESULTSLINE-1 methylation levels were significantly lower (P < 0.05) and relative TLs were significantly shorter (P < 0.05) in WT compared with normal kidney. There was a significant positive relationship between LINE-1 methylation and relative TL in WT (r = 0.671, P = 0.001). LINE-1 Methylation levels were significantly associated with global DNA methylation (r = 0.332, P < 0.01). In addition, relative TL was shortened and LINE-1 methylation was decreased in a WT cell line treated with the hypomethylating agent 5-aza-2'-deoxycytidine compared with untreated WT cell line.

CONCLUSIONThese results suggest that LINE-1 hypomethylation is common and may be linked to telomere shortening in WT.

Cell Line, Tumor ; Child ; Child, Preschool ; DNA Methylation ; genetics ; Female ; Humans ; Long Interspersed Nucleotide Elements ; genetics ; Male ; Polymerase Chain Reaction ; Telomere ; genetics ; Wilms Tumor ; genetics

OBJECTIVETo explore the impact of Line-1 methylation on clinical features of non-small cell lung cancer and its connection with smoking and other living habits.

METHODSPyrosequencing was used to determine the extent of Line-1 methylation in cancer and adjacent tissues derived from 197 patients with primary non-small cell lung cancer. Non-conditional logistic regression analysis was performed to correlate the level of Line-1 methylation with clinical features and living habits of the patients.

RESULTSLine-1 methylation for cancer tissue and adjacent tissue has measured 68.20±11.63 and 78.90±2.09, respectively (P < 0.01), and has been associated with TNM staging, smoking history and histopathological types.

CONCLUSIONLung cancer tissue Line-1 methylation level is closely related with clinical features and smoking. There is also a correlation between histopathological types of lung cancer and relative hypomethylation of Line-1.

Aged ; Aged, 80 and over ; Carcinoma, Non-Small-Cell Lung ; genetics ; metabolism ; pathology ; DNA Methylation ; Female ; Humans ; Long Interspersed Nucleotide Elements ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged

Transitional-CpG methylation between unmethylated promoters and nearby methylated retroelements plays a role in the establishment of tissue-specific transcription. This study examined whether chromosomal losses reducing the active genes in cancers can change transitional-CpG methylation and the transcription activity in a cancer-type-dependent manner. The transitional-CpG sites at the CpG-island margins of nine genes and the non-island-CpG sites round the transcription start sites of six genes lacking CpG islands were examined by methylation-specific polymerase chain reaction (PCR) analysis. The number of active genes in normal and cancerous tissues of the stomach, colon, breast, and nasopharynx were analyzed using the public data in silico. The CpG-island margins and non-island CpG sites tended to be hypermethylated and hypomethylated in all cancer types, respectively. The CpG-island margins were hypermethylated and a low number of genes were active in the normal stomach compared with other normal tissues. In gastric cancers, the CpG-island margins and non-island-CpG sites were hypomethylated in association with high-level chromosomal losses, and the number of active genes increased. Colon, breast, and nasopharyngeal cancers showed no significant association between the chromosomal losses and methylation changes. These findings suggest that chromosomal losses in gastric cancers are associated with the hypomethylation of the gene-control regions and the increased number of active genes.
Alu Elements/genetics ; *Chromosome Deletion ; CpG Islands/*genetics ; *DNA Methylation ; DNA, Neoplasm/chemistry/isolation & purification ; Gene Expression Profiling ; *Genes, Neoplasm ; Humans ; Long Interspersed Nucleotide Elements/genetics ; Polymerase Chain Reaction ; *Promoter Regions, Genetic ; Stomach Neoplasms/*genetics

We exploited the serial analysis of gene expression (SAGE) libraries and human genome database in silico to correlate the breadth of expression (BOE; housekeep-ing versus tissue-specific genes) and peak rate of expression (PRE; high versus low expressed genes) with the density distribution of the retroelements. The BOE status is linearly associated with the density of the sense Alus along the 100 kb nucleotides region upstream of a gene, whereas the PRE status is inversely correlated with the density of antisense L1s within a gene and in the up- and downstream regions of the 0-10 kb nucleotides. The radial distance of intranuclear position, which is known to serve as the global domain for transcription regulation, is reciprocally correlated with the fractions of Alu (toward the nuclear center) and L1 (toward the nuclear edge) elements in each chromosome. We propose that the BOE and PRE statuses are related to the reciprocal distribution of Alu and L1 elements that formulate local and global expression domains.
Alu Elements/*genetics ; Chromosome Mapping/*methods ; Comparative Study ; Databases, Genetic ; Gene Expression Profiling/*methods ; Gene Expression Regulation/*genetics ; Genome, Human ; Humans ; Long Interspersed Nucleotide Elements/*genetics ; Retroelements/genetics ; Sequence Analysis, DNA/*methods ; Statistics ; Tissue Distribution

OBJECTIVETo finish the work of sequencing the full sequence of intron 51 of dystrophin gene and understand its characteristic of sequence.

METHODSThe whole intron 51 was sequenced by primer walking. The sequencing results were analyzed by repeat sequences, matrix attachment region (MAR) and topoisomerase II cleavage sites. The residue sequences, after removal of the repetitive sequences, were subjected to the analysis of CpG islands, promoter, open reading frame (ORF) and unidentified low copy repeat sequence.

RESULTSThe acquired intron 51 sequence was composed of 38725 bp. Repetitive sequences constituted 37.53% of total intron sequence. The overall G+C content of intron 51 was 36.34%. There are four potential MARs in intron 51. Three of them are clustered in the 12 kb region near exon 51. Numerous ORFs were found on both strands, but no homologues proteins were found in Genbank CDS transcriptional peptide, PDB, SwissProt, PIR and PRF databases.

CONCLUSIONThe expansion of intron 7 over the last 120 million years was mainly the result of L1 insertion into intron 7, and not all of repetitive sequences are associated with chromosomal rearrangement. No sequence of functional significance was found in intron 51. The results suggest that the cluster of MARs may be associated with the instability of intron 51.

Base Sequence ; CpG Islands ; genetics ; Databases, Nucleic Acid ; Dystrophin ; genetics ; Gene Deletion ; Genes ; Humans ; Introns ; genetics ; Long Interspersed Nucleotide Elements ; genetics ; Mutagenesis, Insertional ; Open Reading Frames ; Promoter Regions, Genetic ; Repetitive Sequences, Nucleic Acid ; genetics ; Sequence Analysis, DNA ; methods

OBJECTIVETo prepare the LINE1-ORF1p polyclonal antibody, and to study the effect of LINE1-ORF1p on the proliferation of nephroblastoma WT_CLS1 cells.

METHODSA genetic engineering method was used to achieve prokaryotic expression of LINE1-ORF1p, and rabbits were immunized with LINE1-ORF1p to prepare polyclonal antibody. Indirect ELISA was used to evaluate antibody titer, and Western blot and immunohistochemistry were used to evaluate the specific ability of antibody to recognize LINE1-ORF1p. The eukaryotic expression vector pEGFP-N1-LINE1-ORF1 was constructed and used to transfect WT_CLS1 cells. Western blot and qRT-PCR were used to measure the protein and mRNA expression of LINE1-ORF1, respectively, and cell proliferation assay and colony-forming assay were used to evaluate the effect of LINE1-ORF1p on the proliferation of WT_CLS1 cells and the formation of tumor cell clone.

RESULTSThe LINE1-ORF1p antibody prepared had a titer of >1:16 000 and could specifically recognize LINE1-ORF1p in cells and tumor tissue. WT_CLS1 cells transfected with pEGFP-N1-LINE1-ORF1 had significant increases in the mRNA and protein expression of LINE1-ORF1 and significantly enhanced cell proliferation ability and colony formation ability (P<0.05).

CONCLUSIONSLINE1-ORF1p can promote the growth of nephroblastoma cells and the formation of tumor cell clone, and may be involved in the pathogenesis of nephroblastoma.

Animals ; Antibodies ; analysis ; Blotting, Western ; Cell Line, Tumor ; Cell Proliferation ; Deoxyribonuclease I ; analysis ; genetics ; metabolism ; Humans ; Long Interspersed Nucleotide Elements ; RNA, Messenger ; genetics ; metabolism ; Rabbits ; Transfection ; Wilms Tumor ; genetics ; metabolism ; physiopathology

LINEs (long interspersed nuclear elements or long interspersed repeated DNA elements) contains two open reading frames (ORFs), ORF1 and ORF2. We analysed the ORF2 located in the 5' region to the first exon of oncogene c-myc in canine transmissible venereal tumor (TVT) cell. We also showed the transcription activation was induced by this TVT-LINE sequence using CAT assay. To identify the mutation of tumor suppressor gene, sequence analysis of p53 from TVT cell was performed. We identified the point mutation of 964 nucleotide (T-->C) resulting in the change of amino acid (Phe-->Ser) of p53 tumor suppressor protein.
Amino Acid Sequence ; Animals ; Base Sequence ; Cells, Cultured ; Cricetinae ; DNA, Neoplasm/chemistry/genetics ; Dog Diseases/*genetics ; Dogs ; Long Interspersed Nucleotide Elements/*genetics ; Molecular Sequence Data ; Point Mutation ; Polymerase Chain Reaction ; Sequence Alignment ; Transcription, Genetic ; Tumor Suppressor Protein p53/chemistry/*genetics ; Venereal Tumors, Veterinary/chemistry/*genetics