A role for ubiquitin in the spliceosome assembly pathway.
The spliceosome uses numerous strategies to regulate its function in mRNA maturation. Ubiquitin regulates many cellular processes, but its potential roles during splicing are unknown. We have developed a new strategy that reveals a direct role for ubiquitin in the dynamics of splicing complexes. A ubiquitin mutant (I44A) that can enter the conjugation pathway but is compromised in downstream functions diminishes splicing activity by reducing the levels of the U4/U6-U5 small nuclear ribonucleoprotein (snRNP). Similarly, an inhibitor of ubiquitin's protein-protein interactions, ubistatin A, reduces U4/U6-U5 triple snRNP levels in vitro. When ubiquitin interactions are blocked, ATP-dependent disassembly of purified U4/U6-U5 particles is accelerated, indicating a direct role for ubiquitin in repressing U4/U6 unwinding. Finally, we show that the conserved splicing factor Prp8 is ubiquitinated within purified triple snRNPs. These results reveal a previously unknown ubiquitin-dependent mechanism for controlling the pre-mRNA splicing pathway.
Bellare
Priya
P
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, 2205 Tech Drive, Evanston, Illinois 60208, USA.
Small
Eliza C
EC
Huang
Xinhua
X
Wohlschlegel
James A
JA
Staley
Jonathan P
JP
Sontheimer
Erik J
EJ
D009154:Mutation; D054730:Protein Interaction Domains and Motifs; D011804:Quinolines / Q000494:pharmacology; D012326:RNA Splicing; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D017381:Spliceosomes / Q000378:metabolism*; D013425:Sulfanilic Acids / Q000494:pharmacology; D025801:Ubiquitin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
C077018:PRP8 protein, S cerevisiae; D011804:Quinolines; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D017415:Ribonucleoprotein, U5 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; D013425:Sulfanilic Acids; D025801:Ubiquitin; C492595:ubistatin A