Methyl picolinic acid plays a role in epigenetic modifications in Human HepG2 liver cells.
Loading...
Date
2019
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Abstract
Picolinic Acid (PA) is an endogenous catabolite synthesized from L-Tryptophan via the kynurenine pathway. PA has been reported to possess immunological, neuroprotective, and anti-proliferative properties; however, its most widely researched function is its efficient chelating properties. Despite these findings, the physiological function of PA is yet to be discovered. Many PA derivatives exist, with its methyl derivative being of interest in this study. Studies have shown that Picolinic Acid derivatives such as Fusaric Acid are involved in epigenetic regulation by inducing global DNA hypomethylation in HepG2 liver cells. Therefore, the aim of the study was to investigate the epigenetic properties of Picolinic Acid (parent molecule of Fusaric Acid) and Methyl Picolinic Acid in human HepG2 Liver cells.
DNA methylation and Histone modifications are epigenetic phenomena that modify accessibility to DNA and chromatin structure, which results in regulation of gene expressions. While DNA methylation results in long-term repression, Histone methylation leads to formation of heterochromatin. The methyl-CpG binding domain 2 (MBD2) binds to methylated CpG dinucleotides and negatively regulates DNA methylation. MBD2 also associates with histone lysine methyltransferases, such as SUV39H1, which is involved in pericentric heterochromatin silencing and is responsible for methylating Histone 3 on lysine 9 (H3K9).
The cytotoxicity of Picolinic Acid and Methyl Picolinic Acid in HepG2 liver cells was assessed by the WST-1 assay. The DNA methylation ELISA kit was used to quantify 5-methylcytosine in HepG2 cells. Gene expression for the DNA demethylase MBD2 was determined by qPCR. Protein expression for MBD2, trimethylated H3K9 (H3K9me3) and its methyltransferase, SUV39H1, was assessed by Western Blotting.
The IC50 values for Picolinic Acid (7.5 mM) and Methyl Picolinic Acid (13 mM) were obtained from the WST-1 assay. DNA methylation was significantly decreased in Methyl Picolinic Acid, Picolinic Acid and 5-aza-2-DC treated cells (p<0.0001). In Methyl Picolinic Acid and Picolinic Acid treated cells, MBD2 gene expression was downregulated (p<0.0001) followed by an increase in its protein expression (p<0.0001). MPA increased protein expression of SUV39H1 (p<0.0001) leading to an increase in H3K9me3 (p<0.0001), while Picolinic Acid induced a decrease in SUV39H1 and H3K9me3 protein expression (p<0.0001).
Methyl Picolinic Acid induced DNA hypomethylation as a result of increased MBD2 protein expression, despite the decrease in MBD2 gene expression. The upregulation of MBD2 protein expression promoted the activity of SUV39H1 which subsequently enhanced expression of H3K9me3. Since DNA hypomethylation can restore expression of aberrantly expressed genes, this study suggests a possible role for MPA in ameliorating carcinogenesis.
Description
Masters Degree. University of KwaZulu-Natal, Durban.