Mild folate deficiency induces genetic and epigenetic instability and phenotype changes in prostate cancer cells

Gaia Bistulfi,Erika Vandette,Dominic J Smiraglia,Sei-Ichi Matsui

doi:10.1186/1741-7007-8-6

Gaia Bistulfi, Erika Vandette + Show 2 more

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https://doi.org/10.1186/1741-7007-8-6

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Abstract

BackgroundFolate (vitamin B9) is essential for cellular proliferation as it is involved in the biosynthesis of deoxythymidine monophosphate (dTMP) and s-adenosylmethionine (AdoMet). The link between folate depletion and the genesis and progression of cancers of epithelial origin is of high clinical relevance, but still unclear. We recently demonstrated that sensitivity to low folate availability is affected by the rate of polyamine biosynthesis, which is prominent in prostate cells. We, therefore, hypothesized that prostate cells might be highly susceptible to genetic, epigenetic and phenotypic changes consequent to folate restriction.ResultsWe studied the consequences of long-term, mild folate depletion in a model comprised of three syngenic cell lines derived from the transgenic adenoma of the mouse prostate (TRAMP) model, recapitulating different stages of prostate cancer; benign, transformed and metastatic. High-performance liquid chromatography analysis demonstrated that mild folate depletion (100 nM) sufficed to induce imbalance in both the nucleotide and AdoMet pools in all prostate cell lines. Random oligonucleotide-primed synthesis (ROPS) revealed a significant increase in uracil misincorporation and DNA single strand breaks, while spectral karyotype analysis (SKY) identified five novel chromosomal rearrangements in cells grown with mild folate depletion. Using global approaches, we identified an increase in CpG island and histone methylation upon folate depletion despite unchanged levels of total 5-methylcytosine, indicating a broad effect of folate depletion on epigenetic regulation. These genomic changes coincided with phenotype changes in the prostate cells including increased anchorage-independent growth and reduced sensitivity to folate depletion.ConclusionsThis study demonstrates that prostate cells are highly susceptible to genetic and epigenetic changes consequent to mild folate depletion as compared to cells grown with supraphysiological amounts of folate (2 μM) routinely used in tissue culture. In addition, we elucidate for the first time the contribution of these aspects to consequent phenotype changes in epithelial cells. These results provide a strong rationale for studying the effects of folate manipulation on the prostate in vivo, where cells might be more sensitive to changes in folate status resulting from folate supplementation or antifolate therapeutic approaches.

Highlights

Folate is essential for cellular proliferation as it is involved in the biosynthesis of deoxythymidine monophosphate and s-adenosylmethionine (AdoMet)
We present a study that, for the first time, simultaneously investigates several aspects characteristic of the molecular, genetic, epigenetic and cellular response to folate depletion in a model comprised of three syngenic prostate cell lines derived from the transgenic adenoma of the mouse prostate (TRAMP) mouse model recapitulating the different stages of prostate cancer - benign, transformed and metastatic - in order to address the possibility that different stages of transformation might alter the response to folate depletion
Chronic mild folate depletion affects prostate cells’ phenotype We grew three syngenic, but phenotypically distinct, clonal cell lines; C-2D, C-2G and C-2H, which were generated from a primary prostate tumour in a TRAMP mouse [14], in folaterestricted (100 nM) medium for 20 population doublings (PD) and refer to these cells as D100, G100, and H100, respectively

Summary

Introduction

Folate (vitamin B9) is essential for cellular proliferation as it is involved in the biosynthesis of deoxythymidine monophosphate (dTMP) and s-adenosylmethionine (AdoMet). Folate (vitamin B9) is an essential nutrient required for the de novo synthesis of deoxythymidine monophosphate (dTMP) and s-adenosylmethionine (AdoMet) through one-carbon metabolism and methionine cycle, respectively (Figure 1). Epidemiological studies associating dietary intake of folate with the incidence of different cancer types are often conflicting likely reflecting this apparent paradox [3]. Molecular studies, both in vitro and in vivo, suggest that the effects of folate depletion might depend on the cell type [4] and on the cellular stage of transformation [5]. Dietary folate depletion, alone or combined with a low methyl diet, induced tumorigenesis of liver and colon in rodents [6,7]

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