L30 binds the nascent RPL30 transcript to repress U2 snRNP recruitment.
The mechanisms of pre-mRNA splicing regulation are poorly understood. Here we dissect how the Saccharomyces cerevisiae ribosomal L30 protein blocks splicing of its pre-mRNA upon binding a kink-turn structure including the 5' splice site. We show that L30 binds the nascent RPL30 transcript without preventing recognition of the 5' splice site by U1 snRNP but blocking U2 snRNP association with the branch site. Interaction of the factors BBP and Mud2 with the intron, relevant for U2 snRNP recruitment, is not affected by L30. Furthermore, the functions of neither the DEAD-box protein Sub2 in the incipient spliceosome nor the U2 snRNP factor Cus2 on branch site recognition are required for L30 inhibition. These findings contrast with the effects caused by binding a heterologous protein to the same region, completely blocking intron recognition. Collectively, our data suggest that L30 represses a spliceosomal rearrangement required for U2 snRNP association with the transcript.
Macías
Sara
S
Centre de Regulació Genòmica, Doctor Aiguader 88, 08003 Barcelona, Spain.
Bragulat
Mireia
M
Tardiff
Daniel F
DF
Vilardell
Josep
J
D001483:Base Sequence; D037502:Galectin 3 / Q000494:pharmacology*; D007438:Introns; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D012326:RNA Splicing; D012331:RNA, Fungal / Q000737:chemistry / Q000187:drug effects / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000737:chemistry* / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism; D014158:Transcription, Genetic
D037502:Galectin 3; D012331:RNA, Fungal; D011994:Recombinant Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins