Global Fitness Profiling Identifies Arsenic and Cadmium Tolerance Mechanisms in Fission Yeast

Abstract

Heavy metals and metalloids such as cadmium [Cd(II)] and arsenic [As(III)] are widespread environmental toxicants responsible for multiple adverse health effects in humans. However, the molecular mechanisms underlying metal-induced cytotoxicity and carcinogenesis, as well as the detoxification and tolerance pathways, are incompletely understood. Here, we use global fitness profiling by barcode sequencing to quantitatively survey the Schizosaccharomyces pombe haploid deletome for genes that confer tolerance of cadmium or arsenic. We identified 106 genes required for cadmium resistance and 110 genes required for arsenic resistance, with a highly significant overlap of 36 genes. A subset of these 36 genes account for almost all proteins required for incorporating sulfur into the cysteine-rich glutathione and phytochelatin peptides that chelate cadmium and arsenic. A requirement for Mms19 is explained by its role in directing iron–sulfur cluster assembly into sulfite reductase as opposed to promoting DNA repair, as DNA damage response genes were not enriched among those required for cadmium or arsenic tolerance. Ubiquinone, siroheme, and pyridoxal 5′-phosphate biosynthesis were also identified as critical for Cd/As tolerance. Arsenic-specific pathways included prefoldin-mediated assembly of unfolded proteins and protein targeting to the peroxisome, whereas cadmium-specific pathways included plasma membrane and vacuolar transporters, as well as Spt–Ada–Gcn5-acetyltransferase (SAGA) transcriptional coactivator that controls expression of key genes required for cadmium tolerance. Notable differences are apparent with corresponding screens in the budding yeast Saccharomyces cerevisiae, underscoring the utility of analyzing toxic metal defense mechanisms in both organisms.

Tdp1 processes chromate-induced single-strand DNA breaks that collapse replication forks

“…Parallel mutant phenotyping by barcode sequencing (Bar-seq) [ 26 , 27 ] was used to identify haploid deletion mutants that are sensitive to 10 μM potassium dichromate (K 2 Cr 2 O 7 ), which slightly inhibits the growth of wild type (described in Materials and Methods). Growth inhibition (GI) scores were obtained for 2,887 mutants, leading to the identification of 68 genes required for chromate resistance ( S1 Table ).…”

“…Sensitivity to DNA damaging agents, such as camptothecin (CPT), hydroxyurea (HU), ionizing radiation (IR), and ultraviolet (UV) light, were amongst the most highly enriched phenotypes annotated to these genes ( Table 2 ). Indeed, hierarchical cluster analysis revealed that the chromium profile more closely resembles DNA damaging agents (CPT, HU and UV) than the toxic metal cadmium or metalloid arsenic ( Fig 1 ) [ 26 , 27 ].…”

“…Toxin concentrations for each screen are indicated at the top of each column. GI values were obtained from the chromium screens reported in this study or published screens using the same functional profiling platform [ 26 , 27 ]. Zoom view (right panel, upper half) indicates mutants shared in cadmium and arsenic screens but not chromium.…”

Expression of mammalian Onzin and Fungal Cadmium Resistance 1 in S. cerevisiae suggests ancestral functions of PLAC8 proteins in regulating mitochondrial metabolism and DNA damage repair

“…Therefore, we could speculate that, by up-regulating ISC biosynthesis, yeast cells expressing the two PLAC8 proteins may increase their survival because they provide essential cofactors to enzymes involved in DNA damage repair, thus reducing the high DNA mutation rate observed in cadmium-exposed yeast cells (Table 1). MM19, a late-acting CIA component likely implicated in the delivery of ISC into nuclear and cytosolic apoproteins [56], was found to be necessary to Schizosaccharomyces pombe to grow on cadmium [60].…”

New Strategies and Tools in Quantitative Genetics: How to Go from the Phenotype to the Genotype

“…The mapping resolution of QTL-seq can be high if phenotyping is not a bottleneck and a large number of plants can be bulked, as in the study by Rishmawi et al (102), which led to the direct identification of a candidate gene for a major root hair branching locus. Although the number of individuals per bulk is important for QTL resolution, it is noteworthy that the power of BSA also depends on the sequencing depth and the heritability of the trait studied (23,37,73,111). It is possible to dissect traits in populations with a more complex genetic architecture and to identify major and minor QTLs using this approach, as illustrated by a study of cold tolerance in rice (138) and a study of germination speed in Arabidopsis (143).…”