? The spliceosome is a large ribonucleoprotein machine comprised of 5 small nuclear RNAs (snRNAs) and ~80 proteins. Assembly of the spliceosome requires a complex series of rearrangements fueled by members of the DEAD-box family of RNA-dependent ATPases. The spliceosome also contains a single essential GTPase, Snu114, whose function is poorly understood. Activation of the fully assembled spliceosome for catalysis requires the displacement of U1 and U4 snRNPs by the U5 snRNP-associated ATPases Prp28 and Brr2. An important unanswered question is how the activity of these ATPases is regulated. Based on recent genetic analyses, the Guthrie lab has proposed that Snu114 controls this regulation and, moreover, that the activity of Snu114 is itself regulated by ubiquitination. Specifically, their data suggest that GTP hydrolysis by Snu114 produces a large conformational rearrangement of the spliceosome via changed interactions between the C-terminal domain of Snu114 and the large U5 snRNP protein Prp8. Since Prp8 has previously been proposed to negatively regulate the activity of Prp28 and Brr2, this GTP-dependent rearrangement could likely allow activation of these ATPases. This proposal will test the hypotheses that Snu114-dependent GTP hydrolysis activates the spliceosome for catalysis and that ubiquitination is important for the function of Snu114. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32GM077844-01
Application #
7112726
Study Section
Special Emphasis Panel (ZRG1-F08-G (20))
Program Officer
Tompkins, Laurie
Project Start
2006-06-01
Project End
2009-05-31
Budget Start
2006-06-01
Budget End
2007-05-31
Support Year
1
Fiscal Year
2006
Total Cost
$45,976
Indirect Cost
Name
University of California San Francisco
Department
Biochemistry
Type
Schools of Medicine
DUNS #
094878337
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Abelson, John; Blanco, Mario; Ditzler, Mark A et al. (2010) Conformational dynamics of single pre-mRNA molecules during in vitro splicing. Nat Struct Mol Biol 17:504-12
Maeder, Corina; Kutach, Alan K; Guthrie, Christine (2009) ATP-dependent unwinding of U4/U6 snRNAs by the Brr2 helicase requires the C terminus of Prp8. Nat Struct Mol Biol 16:42-8
Zhang, Lingdi; Xu, Tao; Maeder, Corina et al. (2009) Structural evidence for consecutive Hel308-like modules in the spliceosomal ATPase Brr2. Nat Struct Mol Biol 16:731-9