Global demethylation of rat chondrosarcoma cells after treatment with 5-aza-2'-deoxycytidine results in increased tumorigenicity.
Abnormal patterns of DNA methylation are observed in several types of human cancer. While localized DNA methylation of CpG islands has been associated with gene silencing, the effect that genome-wide loss of methylation has on tumorigenesis is not completely known. To examine its effect on tumorigenesis, we induced DNA demethylation in a rat model of human chondrosarcoma using 5-aza-2-deoxycytidine. Rat specific pyrosequencing assays were utilized to assess the methylation levels in both LINEs and satellite DNA sequences following 5-aza-2-deoxycytidine treatment. Loss of DNA methylation was accompanied by an increase in invasiveness of the rat chondrosarcoma cells, in vitro, as well as by an increase in tumor growth in vivo. Subsequent microarray analysis provided insight into the gene expression changes that result from 5-aza-2-deoxycytidine induced DNA demethylation. In particular, two genes that may function in tumorigenesis, sox-2 and midkine, were expressed at low levels in control cells but upon 5-aza-2-deoxycytidine treatment these genes became overexpressed. Promoter region DNA analysis revealed that these genes were methylated in control cells but became demethylated following 5-aza-2-deoxycytidine treatment. Following withdrawal of 5-aza-2-deoxycytidine, the rat chondrosarcoma cells reestablished global DNA methylation levels that were comparable to that of control cells. Concurrently, invasiveness of the rat chondrosarcoma cells, in vitro, decreased to a level indistinguishable to that of control cells. Taken together these experiments demonstrate that global DNA hypomethylation induced by 5-aza-2-deoxycytidine may promote specific aspects of tumorigenesis in rat chondrosarcoma cells.
Hamm
Christopher A
CA
Interdisciplinary Graduate Program in Genetics, University of Iowa, Iowa City, Iowa, United States of America.
Xie
Hehuang
H
Costa
Fabricio F
FF
Vanin
Elio F
EF
Seftor
Elisabeth A
EA
Sredni
Simone T
ST
Bischof
Jared
J
Wang
Deli
D
Bonaldo
Maria F
MF
Hendrix
Mary J C
MJ
Soares
Marcelo B
MB
D000818:Animals; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D045744:Cell Line, Tumor; D002813:Chondrosarcoma / Q000235:genetics* / Q000473:pathology*; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation / Q000187:drug effects*; D000077209:Decitabine; D044127:Epigenesis, Genetic / Q000187:drug effects; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020084:Long Interspersed Nucleotide Elements / Q000235:genetics; D051379:Mice; D018895:Microsatellite Repeats / Q000235:genetics; D000078792:Midkine; D009361:Neoplasm Invasiveness / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D051381:Rats; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D023041:Xenograft Model Antitumor Assays
D016207:Cytokines; D055748:SOXB1 Transcription Factors; C527784:Sox2 protein, rat; D000078792:Midkine; D000077209:Decitabine; D001374:Azacitidine