A mechanism to enhance mRNA splicing fidelity: the RNA-dependent ATPase Prp16 governs usage of a discard pathway for aberrant lariat intermediates.
PRP16 encodes an RNA-dependent ATPase required for the second step of mRNA splicing in S. cerevisiae. We have isolated seven alleles of PRP16 that, like the original allele prp16-1, allow splicing of introns with a mutant branch site (UACUAAC to UACUACC), by forming lariat intermediates at the mutant C nucleotide. Every suppressor mutation maps to the region of PRP16 common to RNA-dependent ATPases. We purified three of the mutant proteins and found that all exhibit reduced ATPase activity, as does Prp16-1. An in vivo analysis of the steady-state levels of the splicing intermediates and products provides evidence for a pathway, under the genetic control of PRP16, to discard incorrectly branched substrates. We propose that decreasing the rate of ATP hydrolysis by Prp16 allows aberrantly formed lariat intermediates more time to proceed through the productive rather than the discard branch of this pathway.
Burgess
S M
SM
Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Guthrie
C
C
D000251:Adenosine Triphosphatases / Q000235:genetics*; D000483:Alleles; D000595:Amino Acid Sequence; D004252:DNA Mutational Analysis; D005656:Fungal Proteins / Q000235:genetics*; D005800:Genes, Fungal; D016153:Genes, Suppressor; D008969:Molecular Sequence Data; D020365:RNA Helicases; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012333:RNA, Messenger / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins
D005656:Fungal Proteins; D000072260:RNA Splicing Factors; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D000251:Adenosine Triphosphatases; C075669:PRP16 protein, S cerevisiae; C027900:RNA-dependent ATPase; D020365:RNA Helicases