Assessment of the Biological Pathways Targeted by Isocyanate Using N-Succinimidyl N-Methylcarbamate in Budding Yeast Saccharomyces cerevisiae

Gajendra Kumar Azad,Vikash Singh,Raghuvir S Tomar,Stefan Wölfl

doi:10.1371/journal.pone.0092993

Gajendra Kumar Azad, Vikash Singh + Show 2 more

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https://doi.org/10.1371/journal.pone.0092993

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Abstract

Isocyanates, a group of low molecular weight aromatic and aliphatic compounds possesses the functional isocyanate group. They are highly toxic in nature hence; we used N-succinimidyl N-methylcarbamate (NSNM), a surrogate chemical containing a functional isocyanate group to understand the mode of action of this class of compounds. We employed budding yeast Saccharomyces cerevisiae as a model organism to study the pathways targeted by NSNM. Our screening with yeast mutants revealed that it affects chromatin, DNA damage response, protein-ubiquitylation and chaperones, oxidative stress, TOR pathway and DNA repair processes. We also show that NSNM acts as an epigenetic modifier as its treatment causes reduction in global histone acetylation and formation of histone adducts. Cells treated with NSNM exhibited increase in mitochondrial membrane potential as well as intracellular ROS levels and the effects were rescued by addition of reduced glutathione to the medium. We also report that deletion of SOD1 and SOD2, the superoxide dismutase in Saccharomyces cerevisiae displayed hypersensitivity to NSNM. Furthermore, NSNM treatment causes rapid depletion of total glutathione and reduced glutathione. We also demonstrated that NSNM induces degradation of Sml1, a ribonucleotide reductase inhibitor involved in regulating dNTPs production. In summary, we define the various biological pathways targeted by isocyanates.

Highlights

In recent decades, a key center of attention has been on understanding exposure to hazardous agents in the natural environment including water, soil and air
Significant growth inhibition was observed at very low dose of 5.0 mM N-succinimidyl N-methylcarbamate (NSNM) and the growth was completely inhibited at 100 mM concentration (Fig. 1A)
We screened some of important pathways including epigenetics, histone modifications, DNA damage response, protein-ubiquitylation and chaperones, oxidative stress, TOR pathway and DNA repair pathways

Summary

Introduction

A key center of attention has been on understanding exposure to hazardous agents in the natural environment including water, soil and air. Elucidation of modes of action through studying effects of occupational exposure to environmental contaminants on human health is of foremost concern. Such exposures are often associated with a number of diseases, including cancer, diabetes, and infertility, etc. Evaluation of exposure to natural, environmental hazards, has not been properly elucidated in most human health risk assessments. Isocyanates are shown to be highly reactive to biological macromolecules leading to the formation of carbamylated products. These products include DNA cross links/adducts, which in turn contribute to cytotoxicity and harmful effects [7,8]. It is demonstrated that isocyanates are capable of undergoing bio-transformation reactions, such as DNA damage, apoptosis, oxidative stress, and inflammation in cultured human lymphocytes and lung fibroblasts [9,10]

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