title: Sad1 counteracts Brr2-mediated dissociation of U4/U6.U5 in tri-snRNP homeostasis.
abstract: The yeast Sad1 protein was previously identified in a screen for factors involved in the assembly of the U4/U6 di-snRNP particle. Sad1 is required for pre-mRNA splicing both in vivo and in vitro, and its human orthologue has been shown to associate with U4/U6.U5 tri-snRNP. We show here that Sad1 plays a role in maintaining a functional form of the tri-snRNP by promoting the association of U5 snRNP with U4/U6 di-snRNP. In the absence of Sad1, the U4/U6.U5 tri-snRNP dissociates into U5 and U4/U6 upon ATP hydrolysis and cannot bind to the spliceosome. The separated U4/U6 and U5 can reassociate upon incubation more favorably in the absence of ATP and in the presence of Sad1. Brr2 is responsible for mediating ATP-dependent dissociation of the tri-snRNP. Our results demonstrate a role of Sad1 in maintaining the integrity of the tri-snRNP by counteracting Brr2-mediated dissociation of tri-snRNP and provide insights into homeostasis of the tri-snRNP.
authors.affiliation: Molecular Cell Biology, Taiwan International Graduate Program, Institute of Molecular Biology, Academia Sinica, and Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Yu-Hsin
authors.initials: YH
authors.lastname: Huang
authors.forename: Che-Sheng
authors.initials: CS
authors.lastname: Chung
authors.forename: Der-I
authors.initials: DI
authors.lastname: Kao
authors.forename: Tzu-Chung
authors.initials: TC
authors.lastname: Kao
authors.forename: Soo-Chen
authors.initials: SC
authors.lastname: Cheng
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D064447:Checkpoint Kinase 2 / Q000737:chemistry / Q000378:metabolism*; D006706:Homeostasis; D054730:Protein Interaction Domains and Motifs; D020365:RNA Helicases / Q000378:metabolism*; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000378:metabolism*; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C472240:SNU114 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C440571:Snu66 protein, S cerevisiae; D000255:Adenosine Triphosphate; D064447:Checkpoint Kinase 2; C067022:RAD53 protein, S cerevisiae; C104158:BRR2 protein, S cerevisiae; D020365:RNA Helicases