Relationship between O6-methylguanine-DNA methyltransferase gene regulated by H3K36me3 and DNA damage induced by arsenic in HaCaT Cells / 中华地方病学杂志

ABSTRACT

Objective To observe the influences of NaAsO2 on H3K36me3 modifications,mRNA transcription of O6-methylguanine-DNA methyltransferase gene (MGMT) in HaCaT cells,and to explore the relationship between the transcription of MGMT gene regulated by H3K36me3 and DNA damage induced by arsenic,in order to provide new ideas and scientific basis for prevention and intervention of arsenism.Methods HaCaT cells were treated with 1.25,2.50,5.00 and 10.00 μmol/L NaAsO2 for 24 h,and were also treated with 10.00 μmol/L NaAsO2 for 6,12 and 24 h.HaCaT cells that treated with 0.00 pmol/L NaAsO2 and 0 h were used as blank control group.The degree of DNA damage in peripheral blood cells was detected by single cell gel electrophoresis.The level of H3K36me3 modifications was detected using Western blotting.Quantitative real-time polymerase chain reaction was used to detect the mRNA levels of MGMT gene.Quantitative chromatin immuno-precipitation was used to detect the level of H3K36me3 modifications in the coding regions (ChIP1 and ChIP2) of MGMT gene.Results ①Among the groups of HaCaT cells treated with 2.50,5.00 and 10.00 μmol/L NaAsO2,the levels of tail DNA％ (11.83 ± 1.15,16.85 ± 2.52,24.23 ± 2.75) and olive tail moment (OTM,10.90 ± 1.13,16.19 ± 2.26,23.83 ± 2.79)were significantly increased compared with those of the control group (0.00 μmol/L,2.40 ± 0.51,2.26 ± 0.40,all P ＜ 0.05).After treated with 10.00 μmoFL NaAsO2 for 12 and 24 h,compared with the control group (0 h,3.66 ± 1.02,3.38 ± 1.00),the degrees of tail DNA％ (15.51 ± 1.92,24.18 ± 2.42) and OTM (13.58 ± 2.04,23.14 ± 2.11)were significantly increased (all P ＜ 0.05).②Compared with the control group (0.00 μmol/L,100.00 ± 0.00),the levels of H3K36me3 modifications (60.59 ± 9.75,57.82 ± 11.28,39.45 ± 7.09) were lower at the dosages of 2.50,5.00 and 10.00 μmol/L NaAsO2 (all P ＜ 0.05).Compared with the control group (0 h,100.00 ± 0.00),the levels of H3K36me3 modifications (48.47 ± 9.67,47.75 ± 6.98) were lower after treated with 10.00 μ mol/L NaAsO2 for 12 and 24 h (all P ＜ 0.05).③The levels of H3K36me3 modifications in HaCaT cells exposed to different doses of NaAsO2 were negatively associated with the tail DNA％ and OTM (r =-0.897,-0.903,all P ＜ 0.05).④Compared with the control group (0.00 μmol/L,100.00 ± 0.00),the mRNA levels of MGMT gene were lower at the dosages of 2.50,5.00 and 10.00 pmol/L NaAsO2 (78.20 ± 3.50,61.40 ± 2.60,49.15 ± 4.70,all P ＜ 0.05).⑤There was no observed H3K36me3 enrichmem regularity in the gene encoding ChIP1 and ChIP2 regions of MGMT gene in all doses of NaAsO2 groups (all P ＞ 0.05).Conclusions H3K36me3 may be involved in the regulation of arsenicinduced DNA damage in HaCaT cell.Amenic could inhibit the mRNA transcription of MGMT gene in HaCaT cells,but the transcription of MGMT gene regulate by H3K36me3 is not closely related to DNA damage induced by arsenic.