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

• Published in: PloS one Vol. 4; no. 9; p. e7202
• Main Authors: Hernández, Helena, Makarova, Olga V, Makarov, Evgeny M, Morgner, Nina, Muto, Yutaka, Krummel, Daniel Pomeranz, Robinson, Carol V
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 28.09.2009; Public Library of Science (PLoS)
• Subjects: Biochemistry; Biochemistry/Experimental Biophysical Methods; Biochemistry/Macromolecular Assemblies and Machines; Biochemistry/RNA Structure; Catalysis; Dynamic stability; Dynamics; Exons; HeLa Cells; Humans; Isoforms; Laboratories; Mass spectrometry; Mass Spectrometry - methods; Mass spectroscopy; Molecular biology; Molecular Conformation; Phosphorylation; Post-translation; Post-translational modifications; Protein Isoforms; Protein Processing, Post-Translational; Protein structure; Protein Structure, Quaternary; Proteins; Proteomics; Proteomics - methods; Quaternary; Quaternary structure; Recombinant Proteins - chemistry; Ribonucleic acid; Ribonucleoprotein, U1 Small Nuclear - chemistry; Ribonucleoprotein, U1 Small Nuclear - metabolism; Ribonucleoproteins (small nuclear); RNA; RNA - metabolism; RNA, Small Nuclear - metabolism; Scientific imaging; Spectrometry, Mass, Electrospray Ionization - methods; Spliceosomes - metabolism; Trends
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0007202

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-translationally 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.