Genome-Wide Analysis of Translation in the Early Drosophila Embryo

Regulation of translation plays an essential role in Drosophila embryogenesis, as loss or misexpression of regulatory proteins can cause lethal development defects. As a result of its importance in these early developmental events, there has been considerable study of translation in the early embryo. Here, I complement those studies with an investigation into translation on a genome-wide scale. In chapter 2, I describe my development and optimization of polysome-gradient protocols to study translation in the mature oocyte and early embryo on a genome-wide scale. In chapter 3, I combine the protocols developed in chapter 2 with microarray analysis to study the role of Smaug, a sequence specific RNA-binding protein, in regulating translation on a genome-wide scale. Comparisons of the list of mRNAs that co-immunoprecipitate with Smaug with those that are translationally repressed by Smaug and those that are degraded by Smaug, suggests that Smaug directly regulates the translation and stability of a large number of mRNAs in the early embryo. I also developed a new algorithm to search for Smaug binding sites, which are called Smaug Recognition Elements (SREs), in mRNAs. The presence of SREs was shown to predict Smaug binding and translational regulation. Taken together these data indicate that Smaug plays a direct and global role in regulating the translation and stability of a large fraction of the mRNAs in the early Drosophila embryo.Ph