HYPOMETHYLATION OF LINE-1 RETROTRANSPOSON IN BREAST CANCER CELLS EXPOSED TO PESTICIDES. ROLE OF TGF-ß1 PATHWAY

NOELIA V MIRET ; AYELEN GÓMEZ; MARIANELA LASAGNA; LAURA KASS; GABRIELA ALTAMIRANO; CAROLINA ANDREA PONTILLO; CLAUDIA COCCA; ANDREA SILVANA RANDI; DANIEL ZAPPIA; LORENA ZÁRATE; FEDERICO MONCZOR

Mar Del Plata

Otro; LXIV Reunion Anual Sociedad Argentina de Investigación Clínica (SAIC); 2019

SAIC

Long interspersed nuclear element 1 (LINE-1) is the most common non-LTR retrotransposon in thehuman genome and comprises 17% of the genome. Reactivation of LINE-1 has been linked to breast cancer progression and metastasis. Strong ligands of aryl hydrocarbon receptor (AhR)activate LINE-1 through the transforming growth factor-ß1 (TGF- ß1) pathway in human liver cancer cells. We have observed that environmental doses of two weak AhR ligands: the organochlorine pesticide hexachlorobenzene (HCB, 0.005 μM) and the organophosphate Chocrpyrifos (CPF, 0.5 μM) activate the TGF-ß1 signaling and enhance LINE-1 mRNA expression in the human breast cancer cell line MDA-MB-231. Our aim was to investigate the role of the TGF-ß1 signaling in the mechanism of action and if the pesticides induce epigenetic changes. To study the involvement of the TGF-ß1 signaling on LINE-1 expression, cells were exposed to HCB or CPF in the presence of the TGF-ß1 receptor I inhibitor (SB431542, 2 μM). qPCR results showed that the enhancement exerted by the pesticides was prevented by the treatment with the inhibitor (p?0.05). Considering that LINE-1expression is regulated by methylation of its internal promoter, MDA-MB-231 cells were exposed to HCB or CPF and the methylation status of the LINE-1 5 ́-UTR was determined. Six sites (+38, +103, +167, +234, +306, +373) were analyzed by a combination of single digestions with methylation-sensitive restriction enzymes and qPCR. We found that the methylation was reduced by HCB at the sites +38, +103 and +167 (p?0.05) and by CPF at the sites +38 and +103 (p?0.05). In conclusion, HCB and CPF induce the demethylation of LINE-1 and enhance mRNA expression through TGF-ß1 pathway. Our findings support previous investigations showing that methylation of the first sitesin 5 ́-UTR are essential for the LINE-1 transcriptional regulation. Finally, environmental doses of pesticides promote epigenetic changes that could increase human breast cancer risk.