Do DEAD-box proteins promote group II intron splicing without unwinding RNA?
The DEAD-box protein Mss116p promotes group II intron splicing in vivo and in vitro. Here we explore two hypotheses for how Mss116p promotes group II intron splicing: by using its RNA unwinding activity to act as an RNA chaperone or by stabilizing RNA folding intermediates. We show that an Mss116p mutant in helicase motif III (SAT/AAA), which was reported to stimulate splicing without unwinding RNA, retains ATP-dependent unwinding activity and promotes unfolding of a structured RNA. Its unwinding activity increases sharply with decreasing duplex length and correlates with group II intron splicing activity in quantitative assays. Additionally, we show that Mss116p can promote ATP-independent RNA unwinding, presumably via single-strand capture, also potentially contributing to DEAD-box protein RNA chaperone activity. Our findings favor the hypothesis that DEAD-box proteins function in group II intron splicing as in other processes by using their unwinding activity to act as RNA chaperones.
Del Campo
Mark
M
Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Tijerina
Pilar
P
Bhaskaran
Hari
H
Mohr
Sabine
S
Yang
Quansheng
Q
Jankowsky
Eckhard
E
Russell
Rick
R
Lambowitz
Alan M
AM
D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D007438:Introns; D009154:Mutation; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D012313:RNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D012313:RNA; C058150:MSS116 protein, S cerevisiae; D053487:DEAD-box RNA Helicases