MICRORNA CONTROLS OF NEVEROUS SYSTEM DEVELOPMENT AND FUNCTIONS IN DROSOPHILA MELANOGASTER

Ph.DDOCTOR OF PHILOSOPH

Control of Drosophila Type I and Type II central brain neuroblast proliferation by bantam microRNA

Post-transcriptional regulation of stem cell self-renewal by microRNAs is emerging as an important mechanism controlling tissue homeostasis. Here, we provide evidence that bantam microRNA controls neuroblast number and proliferation in the Drosophila central brain. Bantam also supports proliferation of transit-amplifying intermediate neural progenitor cells in type II neuroblast lineages. The stem cell factors brat and prospero are identified as bantam targets acting on different aspects of these processes. Thus, bantam appears to act in multiple regulatory steps in the maintenance and proliferation of neuroblasts and their progeny to regulate growth of the central brain

<i>miR-989</i> Is Required for Border Cell Migration in the <i>Drosophila</i> Ovary

<div><p>microRNAs (miRNAs) are small non-coding RNAs that regulate gene expression by destabilizing target transcripts and/or inhibiting their translation. miRNAs are thought to have roles in buffering gene expression to confer robustness. miRNAs have been shown to play important roles during tissue development to control cell proliferation, differentiation and morphogenesis. Many miRNAs are expressed in the germ line of <i>Drosophila</i>, and functions have been reported for a few miRNAs in maintenance of stem cell proliferation during oogenesis. Here, we analyse the function of <i>Drosophila miR-989</i> in oogenesis. <i>miR-989</i> is abundant in ovaries. Mutants lacking <i>miR-989</i> did not display gross abnormalities affecting egg chamber formation or maturation. However, the migration of the border cell cluster was severely delayed in <i>miR-989</i> mutant egg chambers. We demonstrate that <i>miR-989</i> function is required in the somatic cells in the egg chamber, not in germ line cells for border cell migration. Loss of <i>miR-989</i> from a fraction of the border cell cluster was sufficient to impair cluster migration as a whole, suggesting a role in border cells. Gene ontology analysis reveals that many predicted <i>miR-989</i> target mRNAs are implicated in regulating cell migration, cell projection morphogenesis, cell adhesion as well as receptor tyrosine kinase and ecdysone signalling, consistent with an important regulatory role for <i>miR-989</i> in border cell migration.</p></div

Recombinase-Mediated Cassette Exchange Provides a Versatile Platform for Gene Targeting: Knockout of miR-31b

A series of vectors has been designed to enhance the versatility of targeted homologous recombination. Recombinase-mediated cassette exchange permits sequential targeting at any locus and improves flexibility in making user-defined mutations. Application of RMCE to delete an intronic microRNA gene is described

A miRNA sensor suggest somatic miR-<i>989</i> activity.

<p>Stage 8 (A, B) and Stage 10A (C, D) eggchambers expressing a <i>miR-989</i> GFP sensor in a wild type (A, C) or in a <i>miR-989</i> mutant background (B, D). While the GFP sensor was expressed at similar levels in the germ line cells in both genetic backgrounds, it was not detectable in the somatic follicle cells in the wild-type egg chambers. However, follicle cells were positively labelled by the sensor in the absence of <i>miR-989</i>. Note that the sensor also labels the border cells in (D) (arrow).</p

Morphology of mid-oogenesis egg chambers and border cell migration.

<p>Mid-oogenesis egg chambers labelled with Phalloidin (green) and border cell marker α-Slbo (white). The germ line derived nurse cell (NC) cluster and oocyte (O) as well as the somatic follicular epithelium (FE), which encapsulates the germ line cells, are identified. A Stage S8 egg chamber. Slbo-positive border cells form in the FE anterior to the NC cluster (arrow). B Stage S9 egg chamber. The FE migrates towards the oocyte where it forms a columnar epithelium. Follicle cells stretch over the NC cluster to form a flat epithelium. The border cells (arrow) migrate through the NC cluster, roughly in parallel to the leading edge of the migrating external follicle cell sheet (arrowheads). C Stage S10A egg chamber. Migration of the border cell cluster and the migrating FE have essentially completed. D Stage S10B egg chamber. The centripetal follicle cells migrate over the anterior face of the oocyte (arrowheads).</p