Expression profiling in vivo demonstrates rapid changes in liver microRNA levels of whitefish (Coregonus lavaretus) following microcystin-LR exposure

Abstract

At present, little is known about the role of miRNAs in liver response of fish to the cyanobacterial hepatotoxin microcystin-LR (MC-LR) treatment, despite the fact that the exposure is thought to underlie multiple acute and chronic effects. To address this question, we used the Real-Time PCR method to examine the differential expression of 6 miRNAs putatively playing roles in signal transduction (let-7c, miR-9b), apoptosis and cell cycle (miR-16a, miR-21a, miR-34a) and fatty acid metabolism (miR-122) in whitefish (Coregonus lavaretus) liver, during the first 48h after intraperitoneal injection of MC-LR (100μg/kg body weight). In addition, we analyzed expression levels of 8 mRNAs and p53 protein, known to be involved in the cell response on the exposure to environmental stressors. Following the challenge we observed a rapid and transient increase in the mean (n=5) levels of individual miRNA expression (from 2.7-fold for miR-122 to 6.8-fold for let-7c), compared to the respective levels in control fish, which mostly peaked at 24h of the experiment. This increase was correlated with a reduction in the expression of mRNAs of genes coding for ferritin H (frih) and HNK Ras –like protein (p-ras) and an overexpression of mRNAs of genes coding for bcl2-associated X protein (bax), cyclin dependent kinase inhibitor 1a (cdkn1a), dicer (dcr), histone 2A (h2a) and p53. Expression of the remaining caspase 6 (cas6) mRNA did not change over 48h of the treatment. Moreover, exposure to MC-LR did not alter whitefish p53 protein levels. Bearing in mind a variety of likely silencing targets for, and the onset of, the aberrant miRNA expression it may be concluded that they are involved in molecular pathways, such as liver cell metabolism, cell cycle regulation and apoptosis, and may contribute to the early phase of MC-LR induced hepatotoxicity.