Gene regulatory activity of newly discovered small RNA species in Drosophila melanogaster

• Main Author: Leong, Onn Kit
• Other Authors: School of Biological Sciences
• Format: Final Year Project
• Language: English
• Published: 2014
• Subjects: DRNTU::Science::Biological sciences::Genetics
• Online Access: http://hdl.handle.net/10356/60771
• Institution: Nanyang Technological University

siRNAs and miRNAs are well-characterized small regulatory RNAs in eukaryotic organisms. In Drosophila melanogaster, the miRNA and RNAi pathways are distinct from one another, despite being principally similar in their involvement of Dicer and Argonaute proteins for mounting repression. Interestingly, there exist many small RNAs that seem to impinge upon components of both pathways for biogenesis, as identified by small RNA libraries recently constructed from deep sequencing of Argonaute-bound RNAs from wild type and mutant flies. The present project identifies which well-established siRNA/miRNA processing factors are required for biogenesis of two selected DCR2-dependent, AGO1-loaded RNA candidate miRNA-like hairpins that are transcribed from the 3’UTR of known mRNA genes. Through knockdown, Luciferase assay and Northern blot analysis, we found out that processing of one of these novel hairpins (Glut4EF) requires a mix of factors from both miRNA/RNAi pathway, including Drosha, Pasha, DCR1, DCR2, Loqs and AGO1. Unexpectedly, we found that the Glut4EF hairpin can directly regulate host gene expression via a Drosha-dependent, RISC-independent manner. Results of this study fundamentally raises more questions than answers, but provides evidence for a novel small RNA processing pathway for these small RNA species and sets the momentum for future studies on such hairpins.