Investigating the Roles of ADARs and A-to-I RNA Editing in Germline RNA Regulation

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PROJECT SUMMARY The complex events of gametogenesis and early embryonic development require tight regulation to ensure proper reproduction and healthy offspring. To achieve this regulation while maintaining the plasticity necessary for embryogenesis, germ cells rely heavily on RNA-level regulation of gene expression. RNA regulation plays a number of important roles in germ cell development and maintenance, including maintenance of germ cell- specific gene programs, protection of the germline genome from foreign or damaging sequences, and translational repression and storage of maternal transcripts necessary for early embryonic development prior to the onset of zygotic transcription. A diverse network of RNA binding proteins and small RNA pathways govern this crucial RNA regulation, many of which have overlapping or redundant roles to ensure proper regulatory fine-tuning. Adenosine DeAminases that act on RNA (ADARs), are RNA binding proteins that can influence the cellular fate of transcripts either by binding to RNA or by catalyzing the deamination of Adenosine to Inosine, known as A-to-I RNA editing. ADARs are present in all animals and have been shown to play important roles in development, innate immunity, and oncogenesis. While the roles of ADARs in germ cells has not been explored, my own preliminary data suggests ADARs are highly expressed in the germline and can influence hundreds of germline transcripts. Additionally, previous work from other labs suggests that ADARs act along with small RNA pathways to perform functions necessary for proper reproduction. As such, I hypothesize that ADARs play a role in the extensive RNA regulation that takes place in the germline. My preliminary experiments investigating the effects of ADARs on germline transcripts suggest that ADARs play a role on regulating ribosome biogenesis. Additionally, I have found that many germline-edited transcripts are known maternal transcripts, which are translationally repressed throughout the germline and stored for loading into the embryo. For these reasons, I hypothesize that ADARs play a role in translational repression that occurs throughout the germline during gametogenesis. To address this hypothesis, I will expand upon my investigation of the effects of ADARs on germline transcripts as well as interactions with other germline RNA regulators (Aim 1). I will assess the impact of ADARs on germline ribosome biogenesis and translational activity (Aim 2). Finally, I will investigate the impact of ADAR editing on maternally loaded transcripts and early embryonic RNA populations (Aim 3). Through these experiments, I will gain an understanding of the effects of ADARs on germline transcripts, and a bigger-picture understanding of their role in germ cell development and maintenance, early embryonic development, and reproduction in general. This work will contribute to our understanding of the complex processes that facilitate successful and healthy reproduction.