Reactivating aberrantly hypermethylated p15 gene in leukemic T cells by a phenylhexyl isothiocyanate mediated inter-active mechanism on DNA and chromatin.
BACKGROUND
We have previously demonstrated that phenylhexyl isothiocyanate (PHI), a synthetic isothiocyanate, inhibits histone deacetylases and remodels chromatins to induce growth arrest in HL-60 myeloid leukemia cells in a concentration-dependent manner.


METHODS
To investigate the effect of PHI, a novel histone deacetylases inhibitor (HDACi), on demethylation and activation of transcription of p15 in acute lymphoid leukemia cell line Molt-4, and to further decipher the potential mechanism of demethylation, DNA sequencing and modified methylation specific PCR (MSP) were used to screen p15-M and p15-U mRNA after Molt-4 cells were treated with PHI, 5-Aza and TSA. DNA methyltransferase 1 (DNMT1), 3A (DNMT3A), 3B (DNMT3B) and p15 mRNA were measured by RT-PCR. P15 protein, acetylated histone H3 and histone H4 were detected by Western Blot.


RESULTS
The gene p15 in Molt-4 cells was hypermethylated and inactive. Hypermethylation of gene p15 was attenuated and p15 gene was activated de novo after 5 days exposure to PHI in a concentration-dependent manner. DNMT1 and DNMT3B were inhibited by PHI (P < 0.05). Alteration of DNMT3A was not significant at those concentrations. Acetylated histone H3 and histone H4 were accumulated markedly after exposure to PHI.


CONCLUSIONS
PHI could induce both DNA demethylation and acetylated H3 and H4 accumulation in Molt-4 cells. Hypermethylation of gene p15 was reversed and p15 transcription could be reactivated de novo by PHI.
Zhangzhou Affiliated Hospital of Fujian Medical University, Zhangzhou, Fujian Province, China.
Shaohong
S
Jiang
Xudong
X
Ma
Yiqun
Y
Huang
Yunlu
Y
Xu
Ruiji
R
Zheng
Jen-Wei
JW
Chiao
D000107:Acetylation / Q000187:drug effects; D045744:Cell Line, Tumor; D002843:Chromatin / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D050762:Cyclin-Dependent Kinase Inhibitor p15 / Q000235:genetics* / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation / Q000187:drug effects*; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology*; D015458:Leukemia, T-Cell / Q000235:genetics* / Q000378:metabolism; D013601:T-Lymphocytes / Q000473:pathology
D002843:Chromatin; D050762:Cyclin-Dependent Kinase Inhibitor p15; D017879:Isothiocyanates; C073279:6-phenylhexyl isothiocyanate; D004247:DNA