MicroRNAs with 3' Modification Regulate Specific Target Genes

MicroRNAs are small RNA molecules known to regulate pathways in all mammalian cells. In brain cancer cells microRNA-21 and its targets show significant correlation in expression and in the future may be utilized as biomarkers of prognosis. Our main focus is to determine the role of a novel version of microRNA-21, with 3’ end modification. We hypothesize that the modified version of microRNA-21 regulates a specific subset of messenger RNAs. In earlier studies, our laboratory had cloned microRNA-21 to determine if it has a 3’ end modification. As a result, 2 different types of microRNA were obtained: a mature microRNA and a second microRNA that had an unusual 3’ modification consisting of a high number of G residues. We used the sequence of the modified microRNA to search for a gene that contains the same sequence and found PHF5A gene. We hypothesized that modified microRNA-21 regulates the expression of PHF5A. In order to confirm the regulation, we first identified the cell line DAOY that does express both modified microRNA-21 and PHF5A. We examined GEO Profiles data and determined that PHF5A is differentially expressed in neural stem cells. We also showed using a microRNA target prediction program, RNA-22, that PHF5A was very likely a target for microRNA-21. In order to prove that PHF5A is regulated by microRNA-21, we constructed a luciferase reporter vector, containing the novel binding site for microRNA-21 in PHF5A. The goal of our project is to confirm that modified microRNA-21 specifically regulates a gene in brain cancer cells.