Polymerase chain reaction-aided genomic sequencing of an X chromosome-linked CpG island: Methylation patterns suggest clonal inheritance, CpG site autonomy, and an explanation of activity state stability
- Beckman Research Inst. of the City of Hope, Duarte, CA (United States)
- Univ. of Washington, Seattle (United States)
The 5{prime} region of the gene encoding human X chromosome-linked phosphoglycerate kinase 1 (PGK1) is a promoter-containing CpG island known to be methylated at 119 of 121 CpG dinucleotides in a 450-base-pair region on the inactive human X chromosome in the hamster-human cell line X8-6T2. Here the authors report the use of polymerase chain reaction-aided genomic sequencing to determine the complete methylation pattern of this region in clones derived form X8-6T2 cells after treatment with the methylation inhibitor 5-azacytidine. They fine (i) a clone showing full expression of human phosphoglycerate kinase is fully unmethylated in this region; (ii) clones not expressing human phosphoglycerate kinase remain methylated at {approximately}50% of CpG sites, with a pattern of interspersed methylated (M) and unmethylated (U) sites different for each clone; (iii) singles, defined as M-U-M or U-M-U, are common; and (iv) a few CpG sites are partially methylated. The data are interpreted according to a model of multiple, autonomous CpG sites, and estimates are made for two key parameters, maintenance efficiency and de novo methylation efficiency. They also consider how the active region is kept free of methylation and suggest that transcription inhibits methylation by decreasing E{sub m} so that methylation cannot be maintained. Thus, multiple CpG sites, independent with respect to a dynamic methylation system, can stabilize two alternative states of methylation and transcription.
- OSTI ID:
- 5603430
- Journal Information:
- Proceedings of the National Academy of Sciences of the United States of America; (United States), Vol. 87:21; ISSN 0027-8424
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
DNA
METHYLATION
DNA POLYMERASES
GENE REGULATION
HUMAN X CHROMOSOME
DNA SEQUENCING
GLYCERIC ACID
OLIGONUCLEOTIDES
TRANSCRIPTION
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
CHROMOSOMES
ENZYMES
HETEROCHROMOSOMES
HUMAN CHROMOSOMES
HYDROXY ACIDS
NUCLEIC ACIDS
NUCLEOTIDYLTRANSFERASES
ORGANIC ACIDS
ORGANIC COMPOUNDS
PHOSPHORUS-GROUP TRANSFERASES
POLYMERASES
PROTEINS
STRUCTURAL CHEMICAL ANALYSIS
TRANSFERASES
X CHROMOSOME
550201* - Biochemistry- Tracer Techniques