Structural Dynamics of the Spliceosome during Assembly and Catalysis with Single Molecule Resolution

Splicing is an essential step in the maturation of all eukaryotic precursor messenger RNA (pre-mRNA) and required for proper translation of nearly all proteins, from yeast to humans. Anomalous splicing can be lethal for the cell and has been linked to numerous human diseases, such as cancer and neurodegenerative disorders. In spite of its importance in human health, the molecular mechanism of splicing is still not well understood. The challenge is that the spliceosome is a multi-megaDalton dynamic assembly of 5 small nuclear RNAs (snRNAs U1, U2, U4, U5 and U6) and >100 proteins that undergoes several highly conserved structural rearrangements.

Among these RNAs, the U2/U6 di-snRNA complex is of particular interest because it comprises the catalytic core of the spliceosome. Our laboratory first characterized the structural dynamics of this complex. Using single molecule fluorescence techniques, we have shown that, in the absence of proteins, both the yeast and human U2/U6 complexes adopt multiple conformations in dynamic equilibrium that correlate well with splicing activation steps in vivo.

The current project consists of elucidating the structure and dynamics of the U2/U6 complex in the context of actively assembling spliceosomal complexes purified from yeast extracts. This highly multi-disciplinary project will touch on multiple fields. The student will apply concepts in Molecular Biology, Genetics, Biochemistry, Biophysics and Mathematical modelling.