Figure 1: Crystal structure of the 10-subunit yeast exosome bound to a T4-lysozyme fused Ski7 (turquoise) and RNA (black). Core subunits of the exosome are grey, cap subunits in shades of yellow and the nuclease Rrp44 is shown in pink.
The Kowalinski group investigates the architecture and mechanisms of macromolecular complexes involved in cellular RNA editing.
Previous and current research
RNA editing is a conserved post-transcriptional processing step that inserts, deletes or alters the nucleotide bases of an RNA transcript. This results in an RNA sequence that differs from the one encoded in the genome. RNA editing affects protein diversity through sequence alterations but also has effects on RNA structure, stability and localisation. We are investigating how exactly a specific RNA is selected for modification and how this process is regulated.
During my PhD thesis I investigated how the innate immune receptor RIG-I triggers an inflammatory response upon recognition of viral RNA. Solving several crystal structures of RIG-I with and without ligand RNA enabled us to draw a conclusive picture of the conformational changes that lead to activation of the receptor (Kowalinski et al., Cell, 2011). In my postdoctoral work, I studied how the helicase-containing Ski complex aids the degradation of cellular RNAs by the exosome complex. Based on the structure of the exosome bound to Ski7 we understood how compartment specificity of the exosomal co-factors is achieved and moreover could identify the human homologue on evidence of the structure (Kowalinski et al., MolCell, 2016). In collaborative efforts we found that the Ski-complex directly connects the translational machinery (the ribosome) with mRNA degradation (the exosome) (Schmidt, Kowalinski et al., Science 2016).
Future projects and goals
In the future, the Kowalinski group wants to gain insight into the mechanisms that specify certain RNA for modification and how the editing is controlled. To this end, we will not only reconstitute recombinant complexes in vitro but also purify native endogenous complexes for structural investigation. We will use X-ray crystallography, cryo-electron microscopy and scattering techniques like SAXS or SANS combined with biophysical methods and biochemical assays to assess the structure-function relationship within these complexes. We will study Trypanosoma brucei, the causative agent of the sleeping sickness, as a model system. Later on we would like to also study dynamic details with single molecule techniques as well as systemic effects via RNA sequencing techniques.