RNA metabolism is the primary target of formamide in vivo

Abstract

The synthesis, processing and function of coding and non-coding RNA molecules and their interacting proteins has been the focus of a great deal of research that has boosted our understanding of key molecular pathways that underlie higher order events such as cell cycle control, development, innate immune response and the occurrence of genetic diseases. In this study, we have found that formamide preferentially weakens RNA related processes in vivo. Using a non-essential Schizosaccharomyces pombe gene deletion collection, we identify deleted loci that make cells sensitive to formamide. Sensitive deletions are significantly enriched in genes involved in RNA metabolism. Accordingly, we find that previously known temperature-sensitive splicing mutants become lethal in the presence of the drug under permissive temperature. Furthermore, in a wild type background, splicing efficiency is decreased and R-loop formation is increased in the presence of formamide. In addition, we have also isolated 35 formamide-sensitive mutants, many of which display remarkable morphology and cell cycle defects potentially unveiling new players in the regulation of these processes. We conclude that formamide preferentially targets RNA related processes in vivo, probably by relaxing RNA secondary structures and/or RNA-protein interactions, and can be used as an effective tool to characterize these processes.

A Synthetic Approach to Reconstruct the Evolutionary and Functional Innovations of the Plant Histone Variant H2A.W

“…Rapid and robust response to environmental stress is critical for cell survival, and growth rates are affected by temperature 21 or the denaturing agent formamide when combined with other deficiencies. 22 We thus performed a serial dilution spot assay to assess the impact of each chimeric H2A on cell survival rates in response to environmental stressors. In contrast to WT, expression of spH2A.W At , which harbors the KSPK motif, caused a severe growth defect in synthetic yeast strains growing at high temperature or on formamide-containing media (Figure 2C; 37 C and 2% formamide).…”

The telomeric Cdc13-Stn1-Ten1 complex regulates RNA polymerase II transcription

“…S4B ) (Prelich and Winston, 1993). This drug has been also used to detect mRNA biogenesis defects (Hoyos-Manchado et al 2017).…”

Mutational analysis of N-ethyl-N-nitrosourea (ENU) in the fission yeast Schizosaccharomyces pombe.

“…In a previous trial for a random mutagenesis study [3] we had roughly adjusted ENU concentration to allow a comparable survival rate to a standard EMS mutagenesis protocol for yeast [18]. Therefore, in the first place, we aimed to finely assess cell viability after ENU treatment in our experimental conditions as compared to well characterised EMS.…”

“…For theses alleles, the switch to destroy function, activity or structure when desired can be turned off simply modulating the culture conditions (temperature, media, presence of chemicals, etc.) [3–5]. These mutations enable elegant genetic studies -otherwise impossible- of essential genes applied to structure-function relationships, biochemical regulation, protein-protein and protein to other partners interactions, etc.…”

“…Another alkylating agent, N-ethyl-N-nitrosourea (ENU), has also been extensively used for a long time for random mutagenesis screens, especially in mice [9]. However, as far as we know, we are the only ones to use it as a mutagen in fission yeast [3]. As other alkylating counterparts, ENU does not need to be metabolised to alkylate DNA proactively in vivo [10–12].…”