Isoforms of U1-70k control subunit dynamics in the human spliceosomal U1 snRNP

Most human protein-encoding genes contain multiple exons that are spliced together, frequently in alternative arrangements, by the spliceosome. It is established that U1 snRNP is an essential component of the spliceosome, in human consisting of RNA and ten proteins, several of which are post- translationally modified and exist as multiple isoforms. Unresolved and challenging to investigate are the effects of these post translational modifications on the dynamics, interactions and stability of the particle. Using mass spectrometry we investigate the composition and dynamics of the native human U1 snRNP and compare native and recombinant complexes to isolate the effects of various subunits and isoforms on the overall stability. Our data reveal differential incorporation of four protein isoforms and dynamic interactions of subunits U1-A, U1-C and Sm-B/B’. Results also show that unstructured post- ranslationally modified C-terminal tails are responsible for the dynamics of Sm-B/B’ and U1-C and that their interactions with the Sm core are controlled by binding to different U1-70k isoforms and their phosphorylation status in vivo. These results therefore provide the important functional link between proteomics and structure as well as insight into the dynamic quaternary structure of the native U1 snRNP important for its function.This work was funded by: BBSRC (OVM), BBSRC and EPSRC (HH and NM), EU Prospects (HH), European Science Foundation (NM), the Royal Society (CVR), and fellowship from JSPS and HFSP (YM and DAPK respectively)

Imaging Mass Spectrometry: Hype or Hope?

Imaging mass spectrometry is currently receiving a significant amount of attention in the mass spectrometric community. It offers the potential of direct examination of biomolecular patterns from cells and tissue. This makes it a seemingly ideal tool for biomedical diagnostics and molecular histology. It is able to generate beautiful molecular images from a large variety of surfaces, ranging from cancer tissue sections to polished cross sections from old-master paintings. What are the parameters that define and control the implications, challenges, opportunities, and (im)possibilities associated with the application of imaging MS to biomedical tissue studies. Is this just another technological hype or does it really offer the hope to gain new insights in molecular processes in living tissue? In this critical insight this question is addressed through the discussion of a number of aspects of MS imaging technology and sample preparation that strongly determine the outcome of imaging MS experiments