Prof. Dr. Wahle
PROJECT INFORMATION :
Our project deals with the post-transcriptional regulation of the Drosophila nanos mRNA. Localized translation of this RNA at the posterior pole is essential for the formation of the anterior-posterior axis of the embryo. However, only a small fraction of the nanos mRNA is localized at the posterior pole; most of it is distributed throughout the embryo. This non-localized RNA is translationally repressed and also deadenylated and degraded during the first ~ three hours of development.
We have previously developed a cell-free system derived from early Drosophila embryos in which appropriate RNA constructs are both deadenylated and translationally repressed in a specific manner depending on motifs called SREs (Smaug Recognition Elements), which serve as binding sites for the repressor protein Smaug. The protein complex forming on the SREs and mediating repression and deadenylation has been isolated and analyzed by mass spectrometry. We are planning to reconstitute this complex from its seven identified components with the aim of elucidating the mechanisms of translational repression and deadenylation. We are also trying to get a biochemical access to the mechanism of piRNA-dependent regulation of nanos that has been discovered by our cooperation partner Martine Simonelig in Montpellier.
KEY TECHNOLOGIES :
- Protein expression
- Protein purification
- In vitro reconstitution)
Temme, C., Zhang, L., Kremmer, E., Ihling, C., Chartier, A., Sinz, A., Simonelig, M., Wahle, E. (2010) Subunits of the Drosophila CCR4-NOT complex and their roles in mRNA deadenylation. RNA 16, 1356-1370
Jeske, M., Moritz, B., Anders, A., Wahle, E. (2011) Smaug assembles an ATP-dependent stable complex repressing nanos mRNA translation at multiple levels. EMBO J. 30, 90-103
Berndt, H., Harnisch, C., Rammelt, C., Stöhr, N., Zirkel, A., Dohm, J. C., Himmelbauer, H., Tavanez, J.-P., Hüttelmaier, S., Wahle, E. (2012) Maturation of mammalian H/ACA box snoRNAs: PAPD5-dependent adenylation and PARN-dependent trimming. RNA 18, 958-972
Schönemann, L., Kühn, U., Martin, G., Schäfer, P., Gruber, A. R., Keller, W., Zavolan, M., Wahle, E. (2014) Reconstitution of CPSF active in polyadenylation: Recognition of the polyadenylation signal by WDR33. Genes Dev. 28, 2381-2393.
Harnisch, C., Cuzic-Feltens, S., Dohm, J. C., Götze, M., Himmelbauer, H., Wahle, E. (2016) Oligoadenylation of 3' decay intermediates promotes cytoplasmic mRNA degradation in Drosophila cells. RNA 22, 428-442.