Differential promoter methylation may be a key molecular mechanism in regulating BubR1 expression in cancer cells.
The BubR1 mitotic-checkpoint protein monitors proper attachment of microtubules to kinetochores, and links regulation of chromosome-spindle attachment to mitotic-checkpoint signaling. Thus, disruption of BubR1 activity results in a loss of checkpoint control, chromosomal instability caused by a premature anaphase, and/or the early onset of tumorigenesis. The mechanisms by which deregulation and/or abnormalities of BubR1 expression operate, however, remain to be elucidated. In this study, we demonstrate that levels of BubR1 expression are significantly increased by demethylation. Bisulfite sequencing analysis revealed that the methylation status of two CpG sites in the essential BubR1 promoter appear to be associated with BubR1 expression levels. Associations of MBD2 and HDAC1 with the BubR1 promoter were significantly relieved by addition of 5-aza-2'-deoxycytidine, an irreversible DNA methyltransferase inhibitor. However, genomic DNA isolated from 31 patients with colorectal carcinomas exhibited a +84A/G polymorphic change in approximately 60% of patients, but this polymorphism had no effect on promoter activity. Our findings indicate that differential regulation of BubR1 expression is associated with changes in BubR1 promoter hypermethylation patterns, but not with promoter polymorphisms, thus providing a novel insight into the molecular regulation of BubR1 expression in human cancer cells.
Park
Hye-Young
HY
Department of Molecular Cell Biology, Center for Molecular Medicine, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746, Korea.
Jeon
Yoon-Kyung
YK
Shin
Hyun-Jin
HJ
Kim
Il-Jin
IJ
Kang
Hio-Chung
HC
Jeong
Sook-Jung
SJ
Chung
Doo-Hyun
DH
Lee
Chang-Woo
CW
D001374:Azacitidine / Q000494:pharmacology; D001483:Base Sequence; D045744:Cell Line, Tumor; D019175:DNA Methylation / Q000187:drug effects; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006367:HeLa Cells; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology*; D011110:Polymorphism, Genetic / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics*; D011485:Protein Binding / Q000187:drug effects; D011494:Protein Kinases / Q000235:genetics*; D017346:Protein Serine-Threonine Kinases; D014158:Transcription, Genetic / Q000187:drug effects
D004268:DNA-Binding Proteins; C115368:MBD2 protein; D011494:Protein Kinases; C584277:BUB1 protein, human; C090464:Bub1 spindle checkpoint protein; D017346:Protein Serine-Threonine Kinases; C110550:HDAC1 protein, human; D056284:Histone Deacetylase 1; D006655:Histone Deacetylases; D001374:Azacitidine