Investigation of miRNA responses against heavy metal stress in aquatic plant Lemna minör

Heavy metals are among the most common environmental pollutants we encounter today. All ecosystems on earth, especially water ecosystems, are negatively affected by the toxic effects of heavy metals. Lemna minor is a monocotyl aquatic plant that belongs to the Lemnaceae family and is generally used in wastewater treatment to remove mineral and organic contamination. MicroRNAs (miRNAs) are known to be important modulators of responses to heavy metal stress for detoxification of metal. In this study, we aimed to determine expression levels of 5 different miRNA (miR156, miR159, miR171, miR395 and miR396) and their target genes (SPL, MYB, GRAS, SULTR and GRF) in L. minor exposed to cadmium stress. L. minor cultured in Hoagland solution was incubated in Hoagland solution containing 100 µM CdCl2 for 0, 6, 12, 24hours. After incubation, miRNA and mRNA isolation were performed from plant samples powdered in liquid nitrogen. After that cDNAs were obtained by RT-PCR and expression levels were determined by qRT-PCR. The wet burning method was used to determine the amount of cadmium in the same L. minor samples. Measurements were accomplished by using the ICP-OES instrument. As a result, it was determined that the amount of heavy metal increased depending on time in L. minor samples exposed to cadmium. Compared to control, the miRNA expression levels showed that expression increased at 6th and 12th hours for each studied miRNA, whereas expression decreased at 24th hour. While the expression level of the SPL gene did not change, the expression levels of the MYB and SULTR2 genes decreased, the expression level of the GRAS gene increased. On the other hand, the expression in the GRF gene increased at the 6th and 12th hours, but decreased at the 24th hour. This study showed that the expression profiles of miRNAs studied in L. minör samples exposed to cadmium stress and that of their target genes were arranged differently from the expression profiles compared to the plants studied in the literature. This is thought to be due to the hyperacumulator characteristic of Lemna minor. Although L. minor is a highly preferred organism to study heavy metal stress, to date, no study examples related to gene expression profiles have been found in the literature. The information obtained from the study is important in terms of guiding further studies on this plant.