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METHYLATION STATUS OF BCL6 DISTINGUISHES DNA SAMPLES FROM BENIGN AND MALIGNANT LYMPHOPROLIFERATIVE DISORDERS
Author(s) -
JME Taylor,
PR Gould,
SM Cairns,
DV Spagnolo,
PH Kay
Publication year - 2001
Publication title -
pathology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.73
H-Index - 74
eISSN - 1440-1827
pISSN - 1320-5463
DOI - 10.1046/j.1440-1827.2001.00006.x
Subject(s) - bcl6 , microbiology and biotechnology , methylation , dna methylation , biology , chromosomal translocation , chromatin , dna , gene , b cell , cancer research , germinal center , genetics , gene expression , antibody
INTRODUCTION: Deregulation of DNA methylation occurs early in neoplastic transformation. Increased cytosine methylation leads to chromatin compaction with gene inactivation, and hypermethylated DNA in condensed chromatin may be less susceptible to mutation and translocation. Cytosine methylation sites in non‐malignant cells are highly regulated, in contrast to the regional hyper‐ and global hypomethylation frequently found in lymphoid and other neoplasms. This preliminary study assessed the methylation status of BCL6 upstream from exon 1. BCL6 encodes a transcription factor essential for normal germinal centre formation and for B cell development after antigen stimulation. It is a repressor of gene transcription but its target genes are ill‐defined. In some categories of B cell non‐Hodgkin lymphoma (B‐NHL), somatic mutation and translocation of BCL6 are frequent. METHODS: DNA was extracted from 60 lymphoproliferative disorders (LPD) comprising 24 benign LPD, 30 B‐NHL (low and high grade) and 6 T cell neoplasms. The level of BCL6 methylation was assessed by Southern hybridization: DNA was digested with Hpa II and Msp I (methylation sensitive and insensitive respectively), and restricted BCL6 fragments were detected with a 32 P‐dCTP radiolabelled 4 kb Sac I BCL6 probe that hybridized to exon 1 and 5′ flanking sequences. Following autoradiography, the size and relative density of hybridized fragments was used to assign methylation status. RESULTS: Hpa II digested DNA from benign LPDs was characterised by fragments of approximately 0.9 kb, 3.6 kb, 3.9 kb, 4.6 kb and 4.9 kb, and using GenBank sequence NT5962, 3 methylated Msp I restriction sites were predicted. Of 22/24 (92%) Hpa II digested DNA samples from benign LPDs, the 3.6 kb, 3.9 kb, 4.6 kb and 4.9 kb fragments all had either similar density, or the latter two fragment sizes had relatively greater density. 28/30 (93%) Hpa II digested DNA samples from B‐NHL had a decrease in relative density of the 4.6 kb and 4.9 kb fragments compared to the 3.6 kb and 3.9 kb fragments, and one sample had a 3.3 kb fragment. Of the 6 Hpa II digested DNA samples from T cell neoplasms, 3 samples had fragments larger than 4.9 kb, 1 sample had 4.6 kb and 4.9 kb fragments having less density than the 3.6 kb and 3.9 kb fragments, and 2 samples had findings indistinguishable from benign LPD samples. CONCLUSION: This study of the methylation status of the upstream flanking region of BCL6 indicates there may be alternative deregulatory mechanisms leading to hypomethylation in the majority of B‐NHL and hypermethylation in some T cell neoplasms. Whether hypomethylation of BCL6 in B‐NHL predisposes to somatic mutation and translocation of this gene remains to be investigated.