A Mechanistic Study On Arsenic Trioxide-induced Downregulation Of Human Histone H4 Acetylation At Lysine 16

Epigenetic mechanisms such as histone post-translational modification heritably regulate gene expression involved in most cellular biological processes. Experimental studies have clarified that the alteration of histone modifications affects the gene expression by changing the chromatin structure, and thereby leads to various cellular responses to environmental influences. Arsenic(As2O3), one of the major pollutants in drinking water, has become a worldwide environmental concern as an already established human carcinogen. Even so, arsenic, especially As2O3 as one of the most effective drugs for treatment of acute APL(acute promyelocytic leukemia), has been used as medical agents for many years. Recently, increasing evidence suggests that As-mediated epigenetic mechanisms may be involved in its toxicity and carcinogenicity, but how this occurs is unclear. It is reported that arsenic trioxide reduced acetylation of histone 4 lysine 16(H4K16ac) in UROtsa cells. However, the effect of arsenic trioxide on histone H4K16 acetyltransferase h MOF(MYST1/KAT8, human male absent on first) in this process is unknown. Here we present evidence from a series of biochemical experiments and knock down/over expression of h MOF approaches arguing that arsenic-induced global histone H4K16 acetylation(H4K16ac) is partly due to the direct interaction between arsenic and the histone acetyltransferase(HAT) h MOF, leading to loss of h MOF HAT activity.Based on our experiments, decreased global H4K16 ac were observed in He La or HEK293 T cells exposed by arsenic for 24, 48 and 72 hours. And at the same time, the transcription and expression of h MOF did not change. In in vitro HAT assay, arsenic directly inhibited the enzymatic activity of h MOF. We synthesized As-immobilized agarose by p-Aminophenyl Arsenoxide(PAPAO) and NHS-activated agarose. Using As-immobilized agarose, we confirmed that arsenic binds directly to h MOF, and that this interaction was competitively inhibited by free As2O3. Also, the direct interaction of arsenic and C2 CH zinc finger peptide was verified by MAIDI-TOF MS and UV absorption detection. Furthermore, over-expression h MOF not only increased resistance to arsenic and caused less toxicity but also effectively reversed the reduction of H4K16 ac caused by arsenic exposure to cells. And knockdown h MOF increased arsenic-induced cell necrosis and apoptosis in He La cells. In addition, arsenic exposure increased expression of deacetyltransferase HDAC4. However, depletion of HDAC4 did not affect global H4K16 ac, and it could not raise H4K16 ac in cells exposed to arsenic, suggesting that HDAC4 might not directly be involved in histone H4K16 acetylation. Chromatin immunoprecipitation(CHIP) assay shows that h MOF enriched at-1890,-476 and +588 kb of the HDAC4 transcriptional start site on HDAC4 was blocked in arsenic-exposed cells.Taken together, As-induced global histone H4K16 ac is partly due to the fact that As binds directly to the histone acetyltransferase h MOF protein and decreases HAT activity of h MOF, and acetylation of histone H4K16 catalyzed by h MOF improved the resistance of human cells to arsenic trioxide, These data suggest a theoretical basis for elucidating the mechanism of arsenic toxicity on the bidirectional functions as a carcinogen and an anti-cancer agent in human cancer cells.