title: A protein map of the yeast activated spliceosome as obtained by electron microscopy.
abstract: We have elucidated the spatial arrangement of proteins and snRNP subunits within the purified spliceosomal B(act) complex from Saccharomyces cerevisiae, using negative-stain immunoelectron microscopy. The B(act) spliceosome exhibits a mushroom-like shape with a main body connected to a foot and a steep and a shallow slope. The U5 core components, including proteins Snu114 and Prp8, are located in the main body and foot, while Brr2 is on the shallow slope. U2 snRNP components and the RNA helicase Prp2 were predominantly located in the upper regions of both slopes. While several proteins of the "nineteen complex" are located on the steep slope, Prp19, Cef1, and the U6 snRNA-binding protein Cwc2 are on the main body. Our results also indicate that the catalytic core RNP of the spliceosome resides in its main body. We thus assign distinct domains of the B(act) complex to its snRNP and protein components, and we provide first structural insights into the remodeling events at the spliceosome during its transformation from the B to the B(act) complex.
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Chengfu
authors.initials: C
authors.lastname: Sun
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Rigo
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Fabrizio
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Berthold
authors.initials: B
authors.lastname: Kastner
authors.affiliation: Department of Cellular Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Lührmann
mesh_terms: D053487:DEAD-box RNA Helicases / Q000378:metabolism; D011485:Protein Binding; D060066:Protein Interaction Maps; D020365:RNA Helicases / Q000378:metabolism; D000072260:RNA Splicing Factors / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000378:metabolism; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D017381:Spliceosomes / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: C513239:CEF1 protein, S cerevisiae; C543031:Cwc2 protein, S cerevisiae; C079877:PRP19 protein, S cerevisiae; C077018:PRP8 protein, S cerevisiae; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; C472240:SNU114 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C104158:BRR2 protein, S cerevisiae; C066490:PRP2 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases
keywords: Bact complex; electron microscopy; immunolabeling; protein localization; spliceosome