title: Brr2 plays a role in spliceosomal activation in addition to U4/U6 unwinding.
abstract: Brr2 is a DExD/H-box RNA helicase that is responsible for U4/U6 unwinding, a critical step in spliceosomal activation. Brr2 is a large protein (∼250 kD) that consists of an N-terminal domain (∼500 residues) with unknown function and two Hel308-like modules that are responsible for RNA unwinding. Here we demonstrate that removal of the entire N-terminal domain is lethal to Saccharomyces cerevisiae and deletion of the N-terminal 120 residues leads to splicing defects and severely impaired growth. This N-terminal truncation does not significantly affect Brr2's helicase activity. Brr2-Δ120 can be successfully assembled into the tri-snRNP (albeit at a lower level than the WT Brr2) and the spliceosomal B complex. However, the truncation significantly impairs spliceosomal activation, leading to a dramatic reduction of U5, U6 snRNAs and accumulation of U1 snRNA in the B(act) complex. The N-terminal domain of Brr2 does not seem to be directly involved in regulating U1/5'ss unwinding. Instead, the N-terminal domain seems to be critical for retaining U5 and U6 snRNPs during/after spliceosomal activation through its interaction with snRNAs and possibly other spliceosomal proteins, revealing a new role of Brr2 in spliceosomal activation in addition to U4/U6 unwinding.
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
authors: Lingdi
authors: L
authors: Zhang
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
authors: Xueni
authors: X
authors: Li
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
authors: Ryan C
authors: RC
authors: Hill
authors: College of Bioscience and Bioengineering, Hebei University of Science and Technology, Shijiazhuang 050018, P. R. China.
authors: Yan
authors: Y
authors: Qiu
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
authors: Wenzheng
authors: W
authors: Zhang
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.
authors: Kirk C
authors: KC
authors: Hansen
authors: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA rui.zhao@ucdenver.edu.
authors: Rui
authors: R
authors: Zhao
mesh_terms: D005800:Genes, Fungal; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D020365:RNA Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics / Q000378:metabolism; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000737:chemistry / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D012322:RNA Precursors; D012331:RNA, Fungal; D012342:RNA, Small Nuclear; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C104158:BRR2 protein, S cerevisiae; D020365:RNA Helicases