ATP can be dispensable for prespliceosome formation in yeast.
The first ATP-dependent step in pre-mRNA splicing involves the stable binding of U2 snRNP to form the prespliceosome. We show that a prespliceosome-like complex forms in the absence of ATP in yeast extracts lacking the U2 suppressor protein CUS2. These complexes display the same pre-mRNA and U snRNA requirements as authentic prespliceosomes and can be chased through the splicing pathway, indicating that they are a functional intermediate in the spliceosome assembly pathway. ATP-independent prespliceosome-like complexes are also observed in extracts containing a mutant U2 snRNA. Loss of CUS2 does not bypass the role of PRP5, an RNA helicase family member required for ATP-dependent prespliceosome formation. Genetic interactions between CUS2 and a heat-sensitive prp5 allele parallel those observed between CUS2 and U2, and suggest that CUS2 mediates functional interactions between U2 RNA and PRP5. We propose that CUS2 enforces ATP dependence during formation of the prespliceosome by brokering an interaction between PRP5 and the U2 snRNP that depends on correct U2 RNA structure.
Perriman
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R
Center for Molecular Biology of RNA, Sinsheimer Laboratories, University of California, Santa Cruz, Santa Cruz, California 95064, USA.
Ares
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D000255:Adenosine Triphosphate / Q000378:metabolism*; D001483:Base Sequence; D053487:DEAD-box RNA Helicases; D017931:DNA Primers; D005656:Fungal Proteins / Q000378:metabolism; D020365:RNA Helicases / Q000378:metabolism; D012322:RNA Precursors / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D012342:RNA, Small Nuclear / Q000235:genetics; D016601:RNA-Binding Proteins / Q000378:metabolism; D011994:Recombinant Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017381:Spliceosomes; D015534:Trans-Activators / Q000378:metabolism
C114067:CUS2 protein, S cerevisiae; D017931:DNA Primers; D005656:Fungal Proteins; D012322:RNA Precursors; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D000255:Adenosine Triphosphate; C064447:PRP5 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases