Genome-wide Screening And Analysis Of Responses To Lithium Hexafluorophosphate And Sodium Arsenite Cell Pathways

Environmental pollutants have significant influence on the production and cultivation of forest trees.As we all known,lithium and arsenic compounds are common environmental pollutants.Due to the extensive use and unreasonable discharge of them in industry and agriculture,high concentrations of lithium and arsenic accumulate in the soil and water,resulting in inhibition of plant growth and development.However,as the long growth cycle and complex genetic background,studies of the toxicity mechanism of lithium and arsenic in higher plants are difficult.Saccharomyces cerevisiae is a suitable model to the study of toxicity mechanism because of its clear genetic background and homologous genes with higher plants,as well as diverse genome collections including single gene deletion collection and yeast GFP collection.It is an ideal model organism for studying cell toxicity of higher organisms.In this study,high-throughput screens were performed to identify genes involved in the responsive to LiPF₆-mediated toxicity and proteins that could form aggregations under NaAsO₂treatment.Finally,the homologous genes were found by bioinformatics comparison with yeast-related genes of Populus trichocarpa,Eucalyptus grandis,Selaginella moellendorffii.This study provides insights into toxicity of lithium and arsenide compounds in forests and the knowledge obtained will promote the breeding and stress resistance research of forests and under-forest economic crops.First,we screened the gene deletion collection for changes in cell growth in solid plates containing LiPF₆ and got 75 strains that have increased sensitivity or tolerance to LiPF₆.Next,Gene Ontology(GO)enrichment analysis of genes deleted in the 76 strains was performed.We found that genes were enriched in several processes including mitochondrial respiratory chain complex III,proton-transporting two-sector ATPase complex assembly,cytochrome complex pathways.KEGG analysis of the sensitive strains was also performed,the enriched pathway was found:oxidative phosphorylation.We also found that LiPF₆ is more toxic to yeast than lithium chloride(LiCl)or sodium hexafluorophosphate(NaPF₆)or combined of them.This indicates that the cytotoxicity induced by LiPF₆ is not simply the addition of Li and PF₆^-toxicity,but specific,and specifically leads to the sensitivity of mutants related to the oxidative phosphorylation pathway.Physiological analysis showed that a high concentration of LiPF₆ caused mitochondrial damage,reactive oxygen species(ROS)accumulation,and ATP content changes.In these phosphorylation-related genes deletion mutants,LiPF₆ treatment resulted in higher ROS production and reduced ATP levels,suggesting that these genes were important for counteracting LiPF₆-induced toxicity.Through the screen of yeast GFP collection with NaAsO₂,there are 75 proteins that aggregated in response to NaAsO₂,proteins participated in chaperones-mediated protein folding,protein synthesis and degradation,and proteins in metabolic pathways were found to prone to aggregate.It is speculated that NaAsO₂ may inhibit the function of corresponding protein by inducing protein to form aggregates which can generate cytotoxicity.By searching the homologous genes of yeast genes deleted in the strains with increased sensitivity to LiPF6 in Populus trichocarpa,Eucalyptus grandis,Selaginella moellendorffii,total of 9 yeast genes could be compared to 28 homologous genes in Populus trichocarpa,Eucalyptus grandis,Selaginella moellendorffii.These genes may be involved in LiPF₆ tolerance in related trees.By searching the homologous genes of proteins prone to aggregation in trees,total of 6 yeast genes could be compared to32 homologous genes were found in Populus trichocarpa,Eucalyptus grandis,Selaginella moellendorffii,It was speculated that NaAsO₂ stress might induce homologous genes in these trees to form protein aggregates.In summary,this study identified that these genes specifically involved in regulating LiPF₆ toxicity and the cytotoxicity induced by LiPF₆ is corresponding to oxidative stress and ATP imbalance,oxidative phosphorylation related genes play an important role in the resistance to the cytotoxicity of LiPF₆.Through the screen of yeast GFP collection with NaAsO₂,75 aggregated proteins were obtained and analyzed.Finally,homologous genes were searched in Populus trichocarpa and related trees.This study provided a theoretical basis for the resistance of subsequent trees to the stress of lithium hexafluorophosphate and sodium arsenite.