Characterization of oskar translational activation and the oskar RNA function




Kanke, Matthew Robert

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Oskar (Osk) protein is required for posterior body patterning and establishment of the germline in Drosophila. Coordination of osk mRNA localization and translational regulation ensures Osk protein expression is confined to the oocyte posterior. Proper expression requires repression of osk RNA during transport and activation upon localization. Once activated, osk mRNA is translated into two protein isoforms, Long and Short Osk. Here I describe an element in the 5’ end of osk mRNA that is highly conserved across multiple Drosophila species and required for osk translational activation. This 5’ element is located in a region that is also protein coding for the longer Osk isoform and assays were designed to disentangle the effects that mutations had on protein and RNA function. The 5’ element is needed for efficient Osk translation, but only in the absence of Long Osk translation from the same transcript, suggesting a redundant role. Although the 5’ element was previously implicated in a posterior-specific relief of repression, here I provide evidence that the 5’ element acts as a general enhancer of translation, independent of localization and repression. In addition to its protein coding role, osk mRNA has a non-coding function. Egg chambers lacking osk mRNA fail to form a karyosome and arrest mid-oogenesis. RNA function depends on the presence of the osk 3’ UTR in the oocyte. Here I demonstrate that osk mRNA influences distribution of regulators. In the absence of osk mRNA these regulators dissociate from ribonucleoproteins in the germ cells and accumulate in the follicle cells. I find that the osk 3’ UTR performs multiple roles contributing to RNA function. Multiple binding sites act to sequester the translational repressor Bruno in one role. Another involves sequences not bound by Bruno near the 3’ end of osk. In contrary to disruption of Bruno sequestration, which requires mutation of multiple binding sites, mutation of a single site was sufficient to disrupt RNA function. However, disruption of either role recapitulates the failure of karyosome formation and the accumulation of regulators in the follicle cells.


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