Role of MicroRNAs and RNA-Binding Proteins in Addiction-Related Gene Expression

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DESCRIPTION (provided by applicant): Although post-transcriptional mechanisms play a vital role in the control of gene expression, their role in the establishment of addictive behaviors has received very little attention. Amongst these, mRNA stability is estimated to control about 10% of all human genes. One of the most studied cis-acting elements is the AU-rich element (ARE) present in the 3'untranslated region (3'UTR) of several unstable mRNAs. These sequences are targets of RNA-binding proteins such as the neuronal-specific and plasticity-associated RNA-binding protein HuD. We have recently identified three new binding motifs for HuD; two of which are U-rich and thus termed HuD-AREs. Using bioinformatics analyses, we found that there is an overrepresentation of HuD-ARE motifs in the 3' UTRs of mRNAs that are associated with mechanisms of addiction, including BDNF and GAP-43. ARE sequences are not only targets of RNA-binding proteins but also they were recently shown to be targets of specific microRNAs. In the case of BDNF and GAP-43, the same ARE motifs that are recognized by HuD also contain target sequences for miR-495. Given that the overlap between sequences recognized by RNA-binding proteins and microRNAs and the overrepresentation of these sites in the 3' UTRs of addiction-related genes (ARGs), we propose that RNA-binding proteins such as HuD could compete with microRNAs such as miR-495 for the post-transcriptional control of genes associated with substance abuse. To test this hypothesis we propose: 1) To use bioinformatics tools to mine data from public domain databases and from our own microarray studies to investigate the prevalence and co-localization of HuD-AREs and microRNA sites in the 3'UTR of genes associated with drug addiction and 2) To experimentally test the functional competition between HuD and miR-495 on the stability of the BDNF and GAP-43 mRNAs in vitro and to assess the significance of these interactions in vivo in an animal model of cocaine-self-administration. The experiments described in this R03 grant are the first steps to address the possible interactions of microRNAs and RNA-binding proteins in the control of addiction-related gene expression, a novel and important regulatory process. PUBLIC HEALTH RELEVANCE: Although post-transcriptional mechanisms play a vital role in the control of gene expression, their role in the establishment of addictive behaviors has received very little attention. Our preliminary results indicate that 1) genes associated with drug addiction have an unusual high number of post-transcriptional regulatory elements, 2) these elements are recognized by two key post-transcriptional regulators, microRNAs and RNA-binding proteins and 3) these molecules could compete to control gene expression. The experiments described in this R03 grant are the first steps to address the possible interactions of microRNAs and RNA-binding proteins in the control of addiction-related gene expression, a novel and important regulatory process.