A Mira-GFP screen : looking for missing genes in bicoid mRNA localisation

• Main Author: Chang, C-W.
• Published: University of Cambridge 2007
• Subjects: 591.35
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597452

During <i>Drosophila </i>oogenesis, <i>bicoid</i> mRNA is localised to the anterior of the oocyte, where it is translated after egg deposition to pattern the head. However, only a few genes involved in the <i>bcd</i> mRNA localisation have been identified in classic genetic screens. In this thesis, I describe a genetic screen for suppressors of Miranda-GFP, which was designed to identify missing genes required for the <i>bcd</i> mRNA localisation. Mira is not endogenously expressed during oogenesis. However, when Mira-GFP is expressed in the female germ line, the resulting embryos show a maternal-effect lethal bicaudal phenotype. <i>oskar </i>mRNA is localised to both anterior and posterior ends of these embryos and co-localises with Staufen to both ends of the oocyte. Stau is double-strand RNA binding protein and is required for <i>osk</i> and <i>bcd</i> mRNA localisation during oogenesis and asymmetrical <i>prospero</i> mRNA localisation in neuroblasts. Mira functions as an adaptor to couple Stau/<i>pros</i> mRNA to the basal side of neuroblasts. Mutants in genes required for <i>bcd</i> mRNA localisation, such as <i>exuperantia</i> and <i>swallow,</i> suppress the Mira-GFP bicaudal phenotype. <i>Osk </i>mRNA diffuses from the anterior of these oocytes in a similar way to <i>bcd</i> mRNA, indicating that Mira-GFP couples Stau/<i>osk</i> mRNA to the anterior localisation pathway. I further confirmed this by showing that the anterior localisation of Mira-GFP is microtubule-dependent, like <i>bcd</i> mRNA. In order to find genes required for <i>bcd</i> mRNA localisation, I performed a screen for dominant suppressors of the Mira-GFP bicaudal phenotype. I have identified and characterised a number of interesting mutants from this screen, and have shown that several of them cause defects in <i>bcd</i> mRNA localisation.