Disruption of polyubiquitin gene Ubc leads to attenuated resistance against arsenite-induced toxicity in mouse embryonic fibroblasts

Abstract

The polyubiquitin gene Ubc is upregulated under oxidative stress induced by arsenite [As(III)]. However, the detailed mechanism of Ubc upregulation and the exact role of ubiquitin (Ub) to protect cells against As(III)-induced toxicity remain unknown. Here, we found that Ubc-/- mouse embryonic fibroblasts (MEFs) exhibited reduced viability under As(III) exposure, although the Nrf2-Keap1 pathway was activated as a cytoprotective response. Intriguingly, due to the reduced polyubiquitination and delayed onset of degradation of Nrf2 in Ubc-/- MEFs, the basal expression levels of Nrf2 target genes were elevated. As(III)-induced accumulation of Ub conjugates occurred in an Nrf2-independent manner, probably due to cellular stress conditions, including reduced proteasomal activity. Increased cellular Ub levels were essential to polyubiquitinate misfolded proteins generated under As(III) exposure and to degrade them by the proteasome. However, when cellular Ub levels decreased, these misfolded proteins were not efficiently polyubiquitinated, but rather accumulated as large protein aggregates inside the cells, causing cytotoxicity. Furthermore, increased activity of the autophagic pathway to clear these aggregates was not observed in Ubc-/- MEFs. Therefore, reduced viability of Ubc-/- MEFs under As(III) exposure may not be due to dysregulation of the Nrf2-Keap1 pathway, but mostly to reduced efficacy to polyubiquitinate and degrade misfolded protein aggregates.

Proteomic Analysis Identifies Ribosome Reduction as an Effective Proteotoxic Stress Response

“…Increasing evidence suggests an alternate model for the toxicity of trivalent arsenic, which is that it induces widespread protein misfolding. Evidence in support of this model includes the following: 1) various mutants of the ubiquitin-pro- teasome pathway are sensitive to arsenic exposure (8,(11)(12)24), 2) arsenic induces accumulation of high molecular weight ubiquitin conjugates (36,37), and 3) arsenic induces the accumulation of protein aggregates associated with the protein disaggregase Hsp104 (24). Our data support the protein misfolding model.…”

Comparative mRNA analysis of behavioral and genetic mouse models of aggression

“…Existing literature suggests that UBC is extremely susceptible to oxidative stress, which may provide some insight into the neurobiology of aggression as the significance of oxidative stress in aggression has also been documented [Coccaro et al, 2016;Kim et al, 2015]. Research has found that reduced antioxidant function and increased levels of oxidative stress markers are associated with aggressive behavior.…”

Involvement of ubiquitin-coding genes in cadmium-induced protein ubiquitination in human proximal tubular cells

“…Hence, it is strongly suggested that polyUb is preferably engaged in Cd-induced accumulation of ubiquitinated proteins in HK-2 cells. Recently, Ubc was reported to be involved in arsenite-induced toxicity in mouse embryonic fibroblasts (MEFs) (Kim et al, 2015), suggesting that polyUb genes may be critical to the mechanism of metal(loid) toxicity. Cd causes the formation of protein inclusion bodies by promoting the accumulation of ubiquitinated proteins in aggresomes in HEK293 cells (Song et al, 2008), and it has been suggested that the Ub ligase FBXO6 reduces Cd toxicity by expediting degradation of misfolded proteins by the UPS in HEK293 cells (Du et al, 2014).…”

“…In addition to Cd, inorganic arsenic and inorganic mercury decrease gene expression of UPS-related proteins, such as Ube2d1, Ube2d2, and Ube2d4 . Kim et al (2015) demonstrated that inorganic arsenic upregulated Ubc in mouse embryonic fibroblasts. Recently, UPS-regulated proteins involved in defense against methylmercury toxicity were identified in yeast cells (Lee et al, 2015).…”