期刊
JOURNAL OF CELLULAR AND MOLECULAR MEDICINE
卷 14, 期 11, 页码 2646-2654出版社
WILEY
DOI: 10.1111/j.1582-4934.2009.00792.x
关键词
DNA methylation; CpG islands; SINE; U87MG; bisulphite sequencing; genetic instability; long-term hypoxia
资金
- India Institute of Medical Sciences [F.6-1/2003-Acad]
- Indian Council of Medical Research, India [63/12/2002-BMS]
- Department of Science and Technology, India
Hypoxia is an integral part of tumorigenesis and contributes extensively to the neoplastic phenotype including drug resistance and genomic instability. It has also been reported that hypoxia results in global demethylation. Because a majority of the cytosine-phosphate-guanine (CpG) islands are found within the repeat elements of DNA, and are usually methylated under normoxic conditions, we suggested that retrotransposable Alu or short interspersed nuclear elements (SINEs) which show altered methylation and associated changes of gene expression during hypoxia, could be associated with genomic instability. U87MG glioblastoma cells were cultured in 0.1% O-2 for 6 weeks and compared with cells cultured in 21% O-2 for the same duration. Real-time PCR analysis showed a significant increase in SINE and reverse transcriptase coding long interspersed nuclear element (LINE) transcripts during hypoxia. Sequencing of bisulphite treated DNA as well as the Combined Bisulfite Restriction Analysis (COBRA) assay showed that the SINE loci studied underwent significant hypomethylation though there was patchy hypermethylation at a few sites. The inter-alu PCR profile of DNA from cells cultured under 6-week hypoxia, its 4-week revert back to normoxia and 6-week normoxia showed several changes in the band pattern indicating increased alu mediated genomic alteration. Our results show that aberrant methylation leading to increased transcription of SINE and reverse transcriptase associated LINE elements could lead to increased genomic instability in hypoxia. This might be a cause of genetic heterogeneity in tumours especially in variegated hypoxic environment and lead to a development of foci of more aggressive tumour cells.
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