The effect of histone deacetylase inhibitors on SRC and BCL2L1 gene expression and a potential role for phosphatases in their transcriptional repression
Histone Deacetylase Inhibitors (HDACi) are a new class of chemotherapeutics which have shown promise in pre-clinical and clinical settings. HDACi have been shown to act by re-programming gene expression, with the transcription of some genes such as p21WAF1 being activated, while others like SRC and BCL2L1 are repressed. The mechanism behind HDACi gene expression changes remains unknown; although it has been shown to involve a direct interaction with gene promoters. Using a quantitative qRT-PCR approach, the effect of various HDACi on the transcription of p21WAF1, SRC and BCL2L1 was examined. TSA and apicidin led to an up regulation of p21WAF1 mRNA levels while c-Src and Bcl-xL mRNA levels were downregulated. Short c-Src mRNA transcripts were unaffected following TSA and apicidin treatments, despite the full length transcripts being repressed. Repression of full length c-Src and Bcl-xL mRNA transcripts was not seen following treatment with MS-275 and MGCD0103, although p21WAF1 mRNA expression was induced. ChIP experiments revealed that following HDACi treatment, histone acetylation levels and RNA Polymerase II occupancy increased in the promoter regions of both the SRC and BCL2L1 genes. RNA Polymerase II occupancy lasted less than 15 minutes in the 3’ regions of the gene following treatment with apicidin and TSA, but was more long-term following MS-275 and MGCD0103 treatment. The protein phosphatase inhibitor Calyculin A completely blocked HDACi mediated repression of c-Src and Bcl-xL mRNA, suggesting a role for protein phosphatases in the mechanism behind HDACi. It is therefore hypothesized that HDACi work through at least two different mechanisms. Whether or not an HDACi leads to gene repression depends on its ability to disrupt an HDAC/protein phosphatase complex and not on their HDAC specificities. The disruption of the complex leads to the release of an active protein phosphatase. The released phosphatase can then presumably act on various factors changing a gene from an active to paused state, possibly through promoter proximal pausing. HDACi unable to disrupt this complex are unable to induce gene repression. Collectively, these studies highlight not only the complexity of HDACi mediated effects within the cell, but also present a new explanation behind HDACi mediated gene repression.
DegreeMaster of Science (M.Sc.)
CommitteeMoore, Stanley; Wilson, Heather; Misra, Vikram
Copyright DateAugust 2013
Histone Deacetylase Inhibitors