Transgenerational changes in Arabidopsis tsRNA expression and chloroplast genomic methylation following exposure to CeO2 nanoparticles
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Crops and wild plants are being exposed to increasing amounts of engineered cerium oxide nanoparticles (CeO2-NPs) through soil, water, and air. Recent studies showed that exposure to CeO2-NPs can alter the transcriptome profiles of plants. To further identify transgenerational molecular impacts from exposure, experimental groups of Arabidopsis thaliana plants were grown for three generations. Treatments with 15 mL of 500 mg/L CeO2-NP +/- treatments occurred in the 1st and 2nd generations (C1C2 = control both generations; T1C2 = treated 1st generation; C1T2 = treated 2nd generation; T1T2 = treated both generations). No treatments were applied during the 3rd generation. Leaves from 28-day-old 2nd and 3rd generation plants were sampled (N = 5) for small RNA sequencing and reduced representation bisulfite sequencing, respectively. Differentially expressed transcripts were predominantly tRNA-derived small RNAs (tsRNA) that were cleaved within the anti-codon loop, and which are capable of regulating translation. For T1C2 plants there was significant up-regulation of 5′ tRH-Asp-GTCs, whereas C1T2 had up-regulation of 5′ tRH-Gly-GCCs. Interestingly, for T1T2 plants there was a switch to down-regulation of 5′ tRH-Gly-GCCs. We used 124,208 100-CpG probes to quantitate differential methylation at genes across the genomes of 3rd generation plants. Notably, 84% of the genes with differentially methylated CpGs were in the chloroplast genome, while only 3% of all 100-CpG probes were generated there. The most wide-spread enrichments of gene ontologies across treatments were those related to ATP synthesis, electron transport and photosystem II. The most frequently impacted biochemical pathway was also ATP synthesis. Overall, these results point to development of epigenetic memory of particular CeO2-NP exposures that occurred in prior generations. The tsRNA expression was more sensitive to exposures in just the 1st or 2nd generations (T1C2 and C1T2). By contrast, 2nd generation exposures (C1T2 and T1T2) had the largest effect on genomic DNA methylation of 3rd generation plants. Results from these experiments suggest that CeO2 exposure during the 1st generation attenuated the responses to a follow-on exposure in the 2nd generation.