The effects of 5‐azacytidine and cadmium on global 5‐methylcytosine content and secondary metabolites in the freshwater microalgae Chlamydomonas reinhardtii and Scenedesmus quadricauda

Abstract

Epigenetic changes are important mechanisms in the regulation of chromatin structure and gene expression. Cytosine methylation is one of the major epigenetic modifications, mediated by DNA methyltransferases, which transfer methyl groups from S‐adenosyl‐L‐methionine (SAM) to the fifth carbon of cytosine. Various external environmental conditions can change the global hypo/hypermethylation pattern of DNA. These alterations may affect the organism's response to stress conditions. In this study, for the first time, we investigated the effects of 5‐azacytidine, a DNA methyltransferase inhibitor, and cadmium, a toxic metal and environmental pollutant, on the growth, biosynthesis of secondary metabolites (phenols, flavonoids, carotenoids), SAM, S‐adenosylhomocysteine, 5′‐methylthioadenosine and global 5‐methylcytosine (5‐mC) in the green microalgae Chlamydomonas reinhardtii and Scenedesmus quadricauda. The studied species showed major differences in 5‐mC content, secondary metabolite content, and antioxidant activity. Cadmium increased GSH (glutathione) content in C. reinhardtii by 60% whereas 5‐azacytidine did not affect GSH. The biosynthesis of GSH in S. quadricauda in response to the stressors was the opposite. Global 5‐mC content of C. reinhardtii was 1%–1.5%, and the content in S. quadricauda was 3.5%. Amount of some investigated methionine cycle metabolites (SAM, S‐adenosyl homocysteine [SAH], methionine) in S. quadricauda distinctly exceeded C. reinhardtii as well. However, chlorophylls a and b, carotenoids, total phenolic content, total flavonoid content and, antioxidant activity were significantly higher in C. reinhardtii than S. quadricauda. Therefore, in further studies it would be advisable to verify whether methylation of cytosine affects the expression of genes encoding certain secondary metabolites.

Effects of 5-azaC on Iridoid Glycoside Accumulation and DNA Methylation in Rehmannia glutinosa

“…The SmDMLs (DEMETER-like DNA glycosylases) expression was downregulated by 5-azaC treatment in Salvia miltiorrhiza, in which the tanshinone content was increased significantly (Jiang et al, 2018;Li et al, 2018), suggesting that the tanshinone accumulation was pronounced by 5-azaC induced DNA demethylation. The effect of 5-azaC on secondary metabolites accumulation and genes expression was also confirmed in Chlamydomonas reinhardtii (Bacova et al, 2019), Dendrobium (Ni et al, 2014), and Pogostemon cablin (Luo et al, 2021). In summary, we speculated that the upregulation of DXS, DXR, GPPS, G10H, and 10HGO induced by 5-azaC treatment was involved in the iridoid glycoside accumulation in R. glutinosa.…”

“…DNA methylation could affect plant growth and development (Candaele et al, 2014 ; Yamamuro et al, 2014 ; Zhou W. et al, 2016 ), SMs accumulation, and the expression of synthetase gene associated with SMs (Qi et al, 2016 ; Wang B. et al, 2018 ; Wang et al, 2019 ). Studies have shown that the gene expression of the secondary metabolic biosynthesis pathway was upregulated under 5-azaC (a DNA methylation inhibitor) treatment (Ni et al, 2014 ; Jiang et al, 2018 ; Li et al, 2018 ; Bacova et al, 2019 ; Luo et al, 2021 ). In our research, under 15 and 50 μM 5-azaC treatment, the expression of DXS, DXR, GPPS, G10H , and 10HGO was upregulated in root and leaf, especially 50 μM ( Figure 5 ).…”

“…In our research, under 15 and 50 μM 5-azaC treatment, the expression of DXS, DXR, GPPS, G10H , and 10HGO was upregulated in root and leaf, especially 50 μM ( Figure 5 ). In addition, the accumulation of secondary metabolites was also increased under 5-azaC treatment (Jiang et al, 2018 ; Li et al, 2018 ; Bacova et al, 2019 ; Luo et al, 2021 ). Ample studies revealed that the expression level of DXS, DXR, GPPS, G10H , and 10HGO could affect the accumulation of the corresponding secondary metabolites.…”

DNA Methylation in Algae and Its Impact on Abiotic Stress Responses

“…Heavy metals (HMs) are significant environmental pollutants, and their toxicity is a problem of increasing significance for ecological, evolutionary, nutritional and environmental reasons [ 97 , 98 ]. Despite low concentrations of iron (Fe), vanadium (V), zinc (Zn) and molybdenum (Mo) are essential to carry out cellular functions of algae [ 99 , 100 ]; the same metals at high concentrations, also including the unessential HMs, such as arsenic (As), cadmium (Cd), chromium (Cr), lead (Pb) and mercury (Hg), cause blockages of the cell division, reduction in photosynthesis and inhibition of various thiol-group-containing enzyme activities [ 101 , 102 , 103 ].…”

“…However, the minimal data in the literature about algal DNA methylation in response to HMs stress, concern only Cd and Cr [ 82 , 102 , 103 , 107 ].…”