Epigenetic changes induced by exposure to ligands of aryl hydrocarbon receptor and breast cancer

MIRET NV; ALTAMIRANO GA; ZAPPIA D; GOMEZ AL; PONTILLO CA; ZARATE LV; LASAGNA M; COCCA C; MONCZOR F; KASS L; RANDI A

Mar del Plata

Congreso; Reunión Anual de Sociedades de Biociencias-LXIV Reunión de la Sociedad Argentina de Investigación Clínica; 2019

Sociedad Argentina de Investigación Clínica

Breast cancer is the main cause of cancer deathamong women and epigenetic changes contribute to the disease. These alterationsinclude the demethylation and activation of the long interspersed nuclearelement 1 (LINE-1) retrotransposon, which has been linked to tumor progression.Strong ligands of aryl hydrocarbon receptor (AhR) activate LINE-1, throughtransforming growth factor-β1 (TGF-β1)/Smad pathway. This study analyzed theeffect of two weak ligands of the AhR on LINE-1 expression and the role of AhRand TGF-β1 in their mechanism of action. The human breast cancer cell lineMDA-MB-231 was exposed to the organochlorine pesticide hexachlorobenzene (HCB) orthe organophosphate chlorpyrifos (CPF). Both activated the AhR/c-Src/Smad axisand enhanced LINE-1 mRNA levels (0.05-50 μM CPF, 0.005 μM HCB, p<0.01). Thisaction on LINE-1 expression was prevented when cells were pretreated with the TGF-β1receptor I inhibitor (2 μM SB431542, p<0.05). Considering that LINE-1 transcriptionis regulated by methylation, 3 sites in the 5´-UTR LINE-1 sequence (+167, +234,+373) were studied by digestion with methylation-sensitive restriction enzymesand qPCR. Only HCB reduced the methylation at the +167 site (p<0.05). LINE-1encodes two proteins, ORF1p and ORF2p, which associate with their own mRNA andallow retrotransposition. ORF1p expression and localization were analyzed by subcellularfractionation and western blot, showing that the pesticides modulated only itslocalization. HCB promoted ORF1p translocation to the nucleus at 0.005 μM (p<0.05)and its cytosolic retention at 0.5-5 μM (p<0.05). In contrast, 0.5-5 μM CPFincreased ORF1p cytosolic localization (p<0.01), but induced translocationto the nucleus at 50 μM (p<0.05). In conclusion, HCB and CPF reactivate LINE-1,enhancing its expression through the AhR/TGF-β1 axis and regulating itslocalization in MDA-MB-231. Furthermore, HCB promotes LINE-1 demethylation, whichcould contribute to increase LINE-1 transcription.