title: Protein localisation by electron microscopy reveals the architecture of the yeast spliceosomal B complex.
abstract: The spliceosome assembles on a pre-mRNA intron by binding of five snRNPs and numerous proteins, leading to the formation of the pre-catalytic B complex. While the general morphology of the B complex is known, the spatial arrangement of proteins and snRNP subunits within it remain to be elucidated. To shed light on the architecture of the yeast B complex, we immuno-labelled selected proteins and located them by negative-stain electron microscopy. The B complex exhibited a triangular shape with main body, head and neck domains. We located the U5 snRNP components Brr2 at the top and Prp8 and Snu114 in the centre of the main body. We found several U2 SF3a (Prp9 and Prp11) and SF3b (Hsh155 and Cus1) proteins in the head domain and two U4/U6 snRNP proteins (Prp3 and Lsm4) in the neck domain that connects the main body with the head. Thus, we could assign distinct domains of the B complex to the respective snRNPs and provide the first detailed picture of the subunit architecture and protein arrangements of the B complex.
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Rigo
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors.forename: Chengfu
authors.initials: C
authors.lastname: Sun
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Fabrizio
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany bkastne@mpibpc.mpg.de reinhard.luehrmann@mpibpc.mpg.de.
authors.forename: Berthold
authors.initials: B
authors.lastname: Kastner
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany bkastne@mpibpc.mpg.de reinhard.luehrmann@mpibpc.mpg.de.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Lührmann
mesh_terms: D011485:Protein Binding; D021381:Protein Transport; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D017381:Spliceosomes / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D017411:Ribonucleoproteins, Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C104158:BRR2 protein, S cerevisiae; D020365:RNA Helicases
keywords: B complex; electron microscopy; immunolabelling; protein localisation; spliceosome