Evaluating the effects of oxidative stress on microRNA accumulation in Drosophila melanogaster and their regulatory roles in cellular protection and degeneration
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Date
2017
Authors
Doughty, Natalie Joan
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Abstract
Oxidative stress is considered to be an underlying mechanism in the pathogenesis of many cellular degenerative processes. In this study, microRNAs (miRNAs) are examined as potential epigenetic regulators of the oxidative stress response. In an effort to identify miRNAs up- or down-regulated during oxidative stress, we conducted a microarray analysis to evaluate miRNA accumulation changes in Drosophila melanogaster exposed to hyperoxic versus normoxic conditions. Several miRNAs were further evaluated using qRT-PCR to determine their accumulation in whole bodies and heads as well as changes over extended hyperoxia exposures. Dme-miR-8, -11, and -970 were found to be up-regulated in both whole bodies and heads after 5 days hyperoxia exposure. Jaguar (jag), castor (cas), and derailed (drl) were identified as putative targets of these three miRNAs using miRNA target prediction algorithms. Reporter gene-based assays were used to examine the interaction of the miRNAs with the target mRNAs, and confirmed functional suppressive relationships between miR-11:cas and miR-970:drl, but not miR-8:jag. Cell-based assays were also used to assess the ability of candidate microRNAs to suppress expression of several different predicted target genes with known antioxidant activities: superoxide dismutase (Sod), heat shock protein cognate70-4 (Hsc70-4), sniffer (sni), thioredoxin-2 (trx-2), and catalase (cat). Sod was not significantly down-regulated by any miRNA, but mir-927:Hsc70-4, mir-964:Hsc70-4, mir-277:sni, mir-1013:trx-2, and mir-1012:cat interactions were all functionally verified in addition to some pairings having correlational accumulation/expression profiles under hyperoxic stress conditions.
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Keywords
Oxidative stress, Drosophila melanogaster, microRNA, Antioxidant genes