Biogenesis & Action of small RNAs
Our group is interested in unraveling the molecular determinants that allow the establishment of a potent RNA interference (RNAi) response in which small RNAs (sRNAs) are used to silence the expression of genes. RNAi pathways that target RNAs from repetitive sequences serve to maintain genome integrity, whereas pathways that target mRNAs allow the regulation of gene expression during development or in adaptation to the environment. We are particularly interested in understanding how sRNAs are produced from larger precursor molecules and how sRNAs induce heterochromatin formation in the nuclear RNAi pathway.
We address these questions by combining various structural biology techniques with quantitative biochemistry. Through our studies, we will thus gain insights into the catalytic activities and regulation and assign the functions to the individual subunits of the large macromolecular multi-protein complexes involved in these pathways. Our research will provide fundamental knowledge of the mechanism of gene silencing and in general on the regulation of gene expression.
Projects
sRNA Action
After biogenesis, mature sRNAs associate with Argonaute proteins to form RNA-induced silencing complexes (RISC), which then act as the actual effector regulating gene expression. RISC can regulate gene expression by either transcriptional gene silencing in the nucleus or post-transcriptional gene silencing in the cytosol. We are interested to understand the nuclear RNAi pathway, in which a RISC induces histone modification and consequently, gene silencing.
piRNA Biogenesis
piRNAs are a distinct class of small RNAs that associate with the PIWI protein and repress transposons in the germline of animals, thereby contributing to animal fertility maintenance. Although several key players of piRNA biogenesis have been identified genetically, the mechanism of piRNA is poorly understood. We are focusing on piRNA biogenesis in C. elegans and want to know how mature piRNAs are generated from longer precursors.
Approach
Our group uses an integrated structural biology approach, with structural biological techniques forming the core (X-ray crystallography & single-particle electron microscopy) and complemented by biochemical and biophysical approaches. Our research focuses on the mechanistic characterization of protein complexes, which we obtain by two complementary methods. In the bottom-up approach, we produce and purify individual components or protein subcomplexes, which then can be used for the gradual build-up of increasingly larger assemblies. In the top-down approach, a single subunit of a protein complex carries an affinity tag, which allows purifying stable complexes from endogenous sources that can be utilized for structural studies but also for the identification of new interaction partners by mass spectrometry.