Genome-wide analysis of polyadenylation and translation in Xenopus

Massachusetts General Hospital, Boston MA

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Abstract

DESCRIPTION (provided by applicant): Xenopus laevis and Xenopus tropicalis are major model organisms useful for the study of vertebrate development, germ cell biology, and various aspects of cell cycle control. Historically, X. laevis has been the preferred model frog due to ease of culture, large size, and availability of material for biochemical purifications. However, partly due to the allotetraploid nature of X. laevis it has lacked a sequenced and annotated genome until recently. The recent release of a draft of the Xenopus laevis genome sequence facilitates application of genome-wide approaches to study gene expression in this important model organism. Genome-wide approaches that assay gene expression by measuring the levels of transcripts at particular time points have been very successful at identification of nove factors important for many processes. However, knowledge of the steady state level of a particular transcript does not equate to precise knowledge of the timing of protein expression from that transcript. In organisms such as Xenopus where early development is programmed by translation of maternal mRNAs in the absence of transcription it is especially important to couple measurement of RNA abundance to measurement of translational output. In this proposal we describe the use of a novel mRNA-sequencing strategy that identifies mRNAs that show patterns of regulated expression during late oogenesis and early development when transcription is silent. In addition, we will use a recently developed method to measure RNA association with ribosomes to measure protein expression and provide a list of noncoding transcripts. Finally, we will use the information gained to identify novel proteins and RNAs that are important for the completion of meiosis. This proposal will use novel methods and the recent genome sequence of Xenopus laeivs to gain a deeper understanding of posttranscriptional regulation of gene expression and use this information to identify factors important for a key developmental time point. Finally, this proposal will generate many resources for the entire Xenopus community and ensure that Xenopus is a key model organism in the future.