The Role of snRNAs in Spliceosomal Catalysis - Progress in Molecular Biology and Translational Science - page 215

modulated by the spliceosomal protein Snu114 and the C-terminal domain of
Prp8, which also improves the binding of Brr2 to one of its targets, the U4/U6
base-paired complex. 113,114
As mentioned above, Brr2 is involved in two important remodeling steps
in the spliceosomal cycle, namely the unwinding of the base-pairing inter-
action of U6 and U4 snRNAs as the precatalytic B complex is transitioning
to the catalytically competent B * complex ( Fig. 6.7 ), thus allowing the for-
mation of U6/U2 complex; and the release of U6 from its interaction with
U2 after the second step of splicing. These two functions of Brr2 are in turn
regulated by Snu114, which is a U5 snRNP-associated GTPase that has sig-
nificant homology to the ribosomal translocase EF-G. 114 Through an ele-
gant series of experiments, Staley and colleagues 114 showed that Snu114
functions as a classic regulatory G protein and signal-dependent switch
for regulating the molecular remodeling function of Brr2. Binding of
GDP to Snu114 prevented Brr2 from performing its function in both
spliceosomal assembly and disassembly, whereas replacing GDP with
GTP or a nonhydrolyzable GTP analog removed this inhibition. These
findings are supported by previous observations indicating that Snu114
extensively interacts with U5-specific proteins Prp8, Prp28, and
Brr2. 86,115 Further, Brenner and colleagues 115 showed that truncation of
the C-terminal domain of Snu114 allows the initial stages of the
spliceosomal assembly to occur, but it blocks the release of U4 during
the conversion of B to B * complex ( Fig. 6.7 ). Finally, mutations in the
GTPase domain of Snu114 prevent its interaction with Prp8 and U5
snRNA and block the assembly of U5 snRNP particle. 115 It is plausible that
the interaction of Snu114 with Prp8 through its GTPase domain may be
one of the mechanisms by which Prp8 and other factors regulate the
spliceosomal cycle.
Another set of helicase proteins function to remodel the existing
spliceosomal conformation to the next stage in the spliceosomal cycle. Per-
haps the best-studied example of these is Prp16, which associates with the
spliceosomes before the first step of splicing and is thought to function by
remodeling the catalytic core to a conformation that is conducive to catalysis
of the second splicing step ( Fig. 6.7 ). 5,17,112 The direct molecular target of
Prp16 is not yet determined. However, it has been shown to directly interact
with the premessenger RNA in the 3 0 splice site region and also to have a
role in remodeling of one of the U6/U2 helices. 55,116 Yet another group of
proteins involved in remodeling the spliceosomal core include 7 proteins in
human and 8-11 proteins in yeast that associate with Prp19 through a