Slx9p (Ygr081cp) is a nonessential yeast protein previously linked genetically with the DNA helicase Sgs1p. Here we report that Slx9p is involved in ribosome biogenesis in the yeast Saccharomyces cerevisiae. Deletion of SLX9 results in a mild growth defect and a reduction in the level of 18S rRNA. Co-immunoprecipitation experiments showed that Slx9p is associated with 35S, 23S, and 20S pre-rRNA, as well as U3 snoRNA and, thus, is a bona fide component of pre-ribosomes. The most striking effects on prerRNA processing resulting from deletion of SLX9 is the accumulation of the mutually exclusive 21S and 27SA2 pre-rRNA. Furthermore, deletion of SLX9 is synthetically lethal with mutations in Rrp5p that block cleavage at either site A2 or A3. We conclude that Slx9p has a unique role in the processing events responsible for separating the 66S and 43S pre-ribosomal particles. Interestingly, homologs of Slx9p were found only in other yeast species, indicating that the protein has been considerably less well conserved during evolution than the majority of trans-acting processing factors.

Sof1p is a trans-acting protein that is essential for biogenesis of the 40S ribosomal subunits in the yeast Saccharomyces cerevisiae. Because of its involvement in the early cleavage steps of precursor rRNA, its interaction with Nop1p and its ability to coprecipitate U3 snoRNA, Sof1p has so far been regarded as a protein that is specific to the U3 snoRNP. To determine whether a site exists within U3 snoRNA with which Sof1p directly or indirectly associates, we studied the ability of ProtA-tagged Sof1p to coimmunoprecipitate mutant versions of U3 snoRNA. None of the tested mutations had a significant effect on the recovery of mutant U3 from cell extracts. Further coimmunoprecipitation experiments, using cells that could be genetically depleted for either Sof1p or U3 snoRNA demonstrated that the two factors associate independently of each other with the 35S precursor RNA. Indeed, association between Sof1p and U3 snoRNA was abolished in cells in which 35S pre-rRNA transcription was blocked. Finally, we found that an overall reduction in the levels of box C/D snoRNPs by genetic depletion of the common Nop58p protein did not affect coprecipitation of 35S pre-rRNA by Sof1p. From these data, we conclude that Sof1p does not assemble into the 90S preribosome as part of the U3, or any other box C/D, snoRNP. The early and independently assembling trans-acting factor Rrp5p also proved to be dispensable for assembly of Sof1p.The production of eukaryotic ribosomes is a highly dynamic process that starts in the nucleolus, a subregion of the nucleus. Here, the 18S, 5.8S, and 25-28S rRNA genes are transcribed into a single large precursor rRNA by RNA polymerase I, while transcription of the 5S rRNA genes by RNA polymerase III gives rise to a separate pre-5S rRNA transcript. Most of the processing of these precursors into the mature rRNA species and their assembly with ribosomal proteins also occurs in the nucleolus.The RNA polymerase I transcript (35S pre-rRNA in yeast) contains, in addition to the three mature rRNAs, two external (5Ј and 3Ј external transcribed spacer [ETS]) and two internal (internal transcribed spacer 1 [ITS1] and ITS2) transcribed spacers (Fig. 1). Removal of the spacer elements involves an ordered series of endo-and exonucleolytic cleavages (for reviews see references 19, 24, and 31). Large complexes containing different sets of trans-acting proteins, ribosomal proteins, and small nucleolar RNAs (snoRNAs) are formed on the various precursor rRNA species to guide this maturation, which has been studied in most detail in the yeast Saccharomyces cerevisiae. The earliest complex, formed on the yeast 35S precursor, is the 80S/90S preribosome. Within this particle, first the 5Ј ETS is removed by cleavages at sites A0 and A1. Cleavage at site A2, or alternatively, at site A3, within ITS1 then separates the 90S preribosome into the 43S and 66S preribosomal subunits that contain the 20S and the 27SA2, or 27SA3, precursor rRNAs, respectively. The former particle is exported to the cytoplasm where the 20S pre-rRNA is ...