DNA methylation in HeLa cells after treatment with valproic acid

Abstract

Valproic Acid (VPA) is a well known Histone Deacetylase (HDAC) inhibitor. Currently, in addition to HDAC inhibition, DNA demethylation is being considered as part of the VPA action on certain cell types. In the present work, the process of DNA demethylation following VPA treatment will be studied in detail in HeLa cells, in which chromatin remodeling attributed to HDAC inhibition has been reported by our group. Previously obtained image analysis data will be expanded to establish patterns of chromatin remodeling as affected by DNA demethylation. Immunocytochemical assays and flow cytometry will be carried out to quantify the DNA methylation abundance. In addition, the Fourier Transform-Infrared (FT-IR) spectrum will be obtained by microspectroscopy to detect spectral changes that may occur as a function of vibrational changes in the DNA molecule due to DNA demethylation. Because the VPA action may be associated to the recently described enzymes named Ten-Eleven Translocation Methylcytosine Dioxygenases (TET), the levels of gene expression and the abundance of these enzymes as well as the levels of DNA hydroxymethylation will also be investigated. It is expected that a better knowledge on the dynamic action of VPA on DNA methylation/demethylation mechanisms could be revealed. (AU)