Effects of Nicotine on Caenorhabditis elegans survival reproduction and gene expressions : Development of an Invertebrate Animal Model for Drugs of Abuse

ECU Author/Contributor (non-ECU co-authors, if there are any, appear on document)

Michael A. Smith (Creator)

Institution

East Carolina University (ECU )

Web Site: http://www.ecu.edu/lib/

Advisor

Xiaoping Pan

Abstract: Although much is known about the addictive effects of nicotine the molecular mechanisms of nicotine-induced effects remain largely unclear. Specifically little is known about the effects of nicotine on gene expression including gene expression controlled by miRNAs. miRNAs may play a key role in regulating gene expression in response to nicotine exposure due to the fact that expression profiles of the miRNAs will be altered. These effects on gene expression may be associated with long-term use and the "addictive" behavior that is often exhibited with use of nicotine. Our goals are to explore the toxicity of nicotine on Caenorhabditis elegans (C. elegans) including survival reproduction and gene expression and to develop C. elegans as a model organism to assess the toxicity of various xenobiotics. We hypothesize that 1) Nicotine affects survival and reproduction of C. elegans; 2) The expression of several important genes including the genes coding for the nicotinic acetylcholine receptor and for oxidative stress response will be affected by nicotine exposure; and 3) The expression of miRNA genes will be changed and related to the selected protein coding gene expression. In survival trials we tested a range of doses to obtain a 24- hour dose-response data. The 24-hour lethal dose-20 (LD20) in C. elegans corresponds to dose of ~3.16 ppm (19.5 uM) of nicotine. A reproduction study revealed that even at low nicotine exposure levels egg-laying is affected. Using qRT-PCR we found that the expression of several egg-laying and oxidative stress related genes were altered by nicotine which may be regulated by miRNAs. We were able to analytically determine that the expression patterns of the selected protein coding genes were dose-related. In the miRNA assay we analyzed the expression of four miRNAs (Cel-mir-70 Cel-mir-58 Cel-mir-790 Cel-mir-253.) which were selected by in-silico prediction of miRNAs that potentially target our protein-coding genes of interest. We found that individual miRNA expression profiles varied among the different concentrations indicating that the nicotine concentration induces a differential miRNA expression. At 3.16 ppm where the protein-coding genes are the most active miRNAs are also up-regulated indicating there is a complex system of regulation based on more than one miRNA many miRNAs may target the same gene. Therefore we believe that miRNAs may play a key role in controlling protein-coding gene expression. The understanding of this relationship between the toxicant (nicotine) and its effects on miRNAs and their targeted genes will lead to a greater understanding of mechanisms of nicotine-related addiction. 

Additional Information

Publication

Thesis

Date: 2012

Keywords

Biology, Biochemistry, nicotine

Subjects

Caenorhabditis elegans

Nicotine addiction

Gene expression

RNA

Animal models in research

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