title: Nucleosomes correlate with in vivo progression pattern of de novo methylation of p16 CpG islands in human gastric carcinogenesis.
abstract: BACKGROUND
  The exact relationship between nucleosome positioning and methylation of CpG islands in human pathogenesis is unknown.
  
  
  RESULTS
  In the present study, we characterized the nucleosome position within the p16 CpG island and established a seeding methylation-specific PCR (sMSP) assay based on bisulfite modification to enrich the p16 alleles containing methylated-CpG at the methylation "seeding" sites within its intron-1 in gastric carcinogenesis. The sMSP-positive rate in primary gastric carcinoma (GC) samples (36/40) was significantly higher than that observed in gastritis (19/45) or normal samples (7/13) (P<0.01). Extensive clone sequencing of these sMSP products showed that the density of methylated-CpGs in p16 CpG islands increased gradually along with the severity of pathological changes in gastric tissues. In gastritis lesions the methylation was frequently observed in the region corresponding to the exon-1 coding-nucleosome and the 5'UTR-nucleosome; the methylation was further extended to the region corresponding to the promoter-nucleosome in GC samples. Only few methylated-CpG sites were randomly detected within p16 CpG islands in normal tissues. The significantly inversed relationship between the p16 exon-1 methylation and its transcription was observed in GC samples. An exact p16 promoter-specific 83 bp-MSP assay confirms the result of sMSP (33/55 vs. 1/6, P<0.01). In addition, p16 methylation in chronic gastritis lesions significantly correlated with H. pylori infection; however, such correlation was not observed in GC specimens.
  
  
  CONCLUSIONS
  It was determined that de novo methylation was initiated in the coding region of p16 exon-1 in gastritis, then progressed to its 5'UTR, and ultimately to the proximal promoter in GCs. Nucleosomes may function as the basic extension/progression unit of de novo methylation of p16 CpG islands in vivo.
authors.affiliation: Key Laboratory of Carcinogenesis and Translational Research, Ministry of Education, Division of Cancer Etiology, Peking University Cancer Hospital and Institute, Beijing, China. zheminglu@163.com
authors.forename: Zhe-Ming
authors.initials: ZM
authors.lastname: Lu
authors.forename: Jing
authors.initials: J
authors.lastname: Zhou
authors.forename: Xiuhong
authors.initials: X
authors.lastname: Wang
authors.forename: Zhenpo
authors.initials: Z
authors.lastname: Guan
authors.forename: Hua
authors.initials: H
authors.lastname: Bai
authors.forename: Zhao-Jun
authors.initials: ZJ
authors.lastname: Liu
authors.forename: Na
authors.initials: N
authors.lastname: Su
authors.forename: Kaifeng
authors.initials: K
authors.lastname: Pan
authors.forename: Jiafu
authors.initials: J
authors.lastname: Ji
authors.forename: Dajun
authors.initials: D
authors.lastname: Deng
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D020029:Base Pairing / Q000235:genetics; D001706:Biopsy; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000473:pathology; D045584:Chromosome Positioning / Q000235:genetics; D018899:CpG Islands / Q000235:genetics*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics*; D019175:DNA Methylation / Q000235:genetics*; D018450:Disease Progression; D005260:Female; D005756:Gastritis / Q000235:genetics / Q000382:microbiology; D016480:Helicobacter pylori / Q000502:physiology; D006801:Humans; D008297:Male; D008875:Middle Aged; D009707:Nucleosomes / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D017422:Sequence Analysis, DNA; D013274:Stomach Neoplasms / Q000235:genetics* / Q000382:microbiology / Q000473:pathology; D013447:Sulfites
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D009707:Nucleosomes; D013447:Sulfites; C042345:hydrogen sulfite