Genomic Phenotyping by Barcode Sequencing Broadly Distinguishes between Alkylating Agents, Oxidizing Agents, and Non-Genotoxic Agents, and Reveals a Role

Toxicity screening of compounds provides a means to identify compounds harmful for human health and the environment. Here, we further develop the technique of genomic phenotyping to improve throughput while maintaining specificity. We exposed cells to eight different compounds that rely on different modes of action: four genotoxic alkylating (methyl methanesulfonate (MMS), N-Methyl-N-nitrosourea (MNU), N,N9-bis(2-chloroethyl)-N-nitroso-urea (BCNU), N-ethylnitrosourea (ENU)), two oxidizing (2-methylnaphthalene-1,4-dione (menadione, MEN), benzene-1,4-diol (hydroquinone, HYQ)), and two non-genotoxic (methyl carbamate (MC) and dimethyl sulfoxide (DMSO)) compounds. A library of S. cerevisiae 4,852 deletion strains, each identifiable by a unique genetic ‘barcode’, were grown in competition; at different time points the ratio between the strains was assessed by quantitative high throughput ‘barcode’ sequencing. The method was validated by comparison to previous genomic phenotyping studies and 90% of the strains identified as MMS-sensitive here were also identified as MMS-sensitive in a much lower throughput solid agar screen. The data provide profiles of proteins and pathways needed for recovery after both genotoxic and non-genotoxic compounds. In addition, a novel role for aromatic amino acids in the recovery after treatment with oxidizing agents was suggested. The role of aromatic acids was further validated; the quinone subgroup of oxidizing agents were extremely toxic in cells where tryptophan biosynthesis was compromised. Citation: Svensson JP, Quirós Pesudo L, McRee SK, Adeleye Y, Carmichael P, et al. (2013) Genomic Phenotyping by Barcode Sequencing Broadly Distinguishes between Alkylating Agents, Oxidizing Agents, and Non-Genotoxic Agents, and Reveals a Role for Aromatic Amino Acids in Cellular Recovery after Quinone Exposure. PLoS ONE 8(9): e73736. doi:10.1371/journal.pone.0073736 Editor: Martin G. Marinus, University of Massachusetts Medical School, United States of America Received May 1, 2013; Accepted July 21, 2013; Published September 9, 2013 Copyright: 2013 Svensson et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This research was supported by Unilever and this work was supported by NCI R01-CA055042 (now R01-ES022872), the CEHS center grant NIEHS P30ES002109, and Unilever. LDS is an American Cancer Society Research Professor. LQP was supported by a Spanish Ministry of Science and Innovation Fellowship. Authors AY and PC are employees of Unilever and contributed with ideas and discussions. With the exception of Unilever, the funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors AY and PC are employed at Unilever. This does not alter the authors’ adherence to all the PLOS ONE policies on sharing data and materials. * E-mail: [email protected]