The functions of the RNA helicase Prp43 and interacting G-patch proteins in ribosome biogenesis

RNA helicases play a crucial role in ribosome biogenesis, as they are believed to drive structural rearrangements by modulating RNA-RNA or RNA-protein interactions. However, their exact functions and substrates are largely unknown. Prp43 is an RNA helicase of the yeast Saccharomyces cerevisiae that is involved in mRNA splicing, but also in ribosome biogenesis. Three physical interaction partners of Prp43 contain a G-patch, which is a sequence motif with up to now unknown function. One of these G-patch proteins, Ntr1, has been shown to act together with Prp43 in splicing and to stimulate the helicase activity of Prp43. The other two G-patch proteins, Pfa1 and Pxr1, seem to have functions in ribosome biogenesis. The role of Pfa1 and Prp43 in 40S subunit biogenesis was already characterized by me in the course of a different project. However, most likely Prp43 also functions in other steps of ribosome maturation. According to my model, Prp43 acts together with Pxr1 in early steps of ribosome biogenesis, and with Pfa1 not only in 40S, but also in 60S biogenesis. The aim of the proposed project is to analyze the functions of Prp43 and the interacting G-patch proteins in ribosome biogenesis. In the first part of the project, the interaction of Prp43 with G-patch proteins will be characterized. Further, I will try to identify the common molecular function of G-patch proteins, which could be the targeting of Prp43 to specific substrates, or the induction of its helicase activity. The focus of the second part of the project is the functional characterization of Pxr1, which likely acts together with Prp43 in early steps of ribosome biogenesis. In the third part of the project I will analyze the function of Pfa1 and Prp43 in 60S biogenesis. For the fourth part of the project, all the knowledge from the other three parts will be combined to predict the approximate sites of action of Prp43 and the respective G-patch proteins. These sites will be assayed for structural changes that are induced by Prp43. The analysis of Prp43 as described in this proposal is of special interest because it will improve our understanding of the structural changes occurring in maturating pre-ribosomal particles. In addition, the investigation of the G- patch proteins will give new insights into how specific steps in ribosome biogenesis are regulated. Consequently, the proposed project will be an important contribution to a more mechanistic understanding of ribosome biogenesis.