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Publication Detail

Title: Arsenite Exposure Displaces Zinc from ZRANB2 Leading to Altered Splicing.

Authors: Banerjee, Mayukh; Ferragut Cardoso, Ana P; Lykoudi, Angeliki; Wilkey, Daniel W; Pan, Jianmin; Watson, Walter H; Garbett, Nichola C; Rai, Shesh N; Merchant, Michael L; States, J Christopher

Published In Chem Res Toxicol, (2020 06 15)

Abstract: Exposure to arsenic, a class I carcinogen, affects 200 million people globally. Skin is the major target organ, but the molecular etiology of arsenic-induced skin carcinogenesis remains unclear. Arsenite (As3+)-induced disruption of alternative splicing could be involved, but the mechanism is unknown. Zinc finger proteins play key roles in alternative splicing. As3+ can displace zinc (Zn2+) from C3H1 and C4 zinc finger motifs (zfm's), affecting protein function. ZRANB2, an alternative splicing regulator with two C4 zfm's integral to its structure and splicing function, was chosen as a candidate for this study. We hypothesized that As3+ could displace Zn2+ from ZRANB2, altering its structure, expression, and splicing function. As3+/Zn2+ binding and mutual displacement experiments were performed with synthetic apo-peptides corresponding to each ZRANB2 zfm, employing a combination of intrinsic fluorescence, ultraviolet spectrophotometry, zinc colorimetric assay, and liquid chromatography-tandem mass spectrometry. ZRANB2 expression in HaCaT cells acutely exposed to As3+ (0 or 5 μM, 0-72 h; or 0-5 μM, 6 h) was examined by RT-qPCR and immunoblotting. ZRANB2-dependent splicing of TRA2B mRNA, a known ZRANB2 target, was monitored by reverse transcription-polymerase chain reaction. As3+ bound to, as well as displaced Zn2+ from, each zfm. Also, Zn2+ displaced As3+ from As3+-bound zfm's acutely, albeit transiently. As3+ exposure induced ZRANB2 protein expression between 3 and 24 h and at all exposures tested but not ZRANB2 mRNA expression. ZRANB2-directed TRA2B splicing was impaired between 3 and 24 h post-exposure. Furthermore, ZRANB2 splicing function was also compromised at all As3+ exposures, starting at 100 nm. We conclude that As3+ exposure displaces Zn2+ from ZRANB2 zfm's, changing its structure and compromising splicing of its targets, and increases ZRANB2 protein expression as a homeostatic response both at environmental/toxicological exposures and therapeutically relevant doses.

PubMed ID: 32274925 Exiting the NIEHS site

MeSH Terms: Alternative Splicing/drug effects; Arsenites/toxicity*; Cell Line; Cell Survival/drug effects; Environmental Pollutants/toxicity*; Humans; RNA-Binding Proteins/genetics; RNA-Binding Proteins/metabolism*; Zinc/metabolism*

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