Two different ribosomal RNA species were probed with 1M7
This series contains 4 samples re-analyzed from GSE83821. The raw data associated with these samples in this series (GSM2856218-GSM286221) include the barcodes not present in the raw data from the original samples from GSE83821 (GSM2219115-GSM2219118).
In eukaryotes, biogenesis of ribosomes requires folding and assembly of the precursor rRNA (pre-rRNA) with a large number of proteins and snoRNPs into huge RNA-protein complexes. In spite of intense genetic, biochemical and high resolution cryo-EM studies in Saccharomyces cerevisiae, information about the conformation of the earliest 35S pre-rRNA is limited. To overcome this, we performed high-throughput SHAPE chemical probing on the 35S pre-rRNA associated with 90S pre-ribosomes. We focused our analyses on external (5´ETS) and internal (ITS1) transcribed spacers as well as the 18S region. We show that in the 35S pre-rRNA, the central region of the 18S is in a more open configuration compared to 20S pre-rRNA and that the central pseudoknot is not formed. The essential ribosome biogenesis protein Mrd1 influences the structure of the 18S part locally and is involved in organizing the central pseudoknot and surrounding structures. Our results demonstrate that the U3 snoRNA dynamically interacts with the 35S pre-rRNA and that Mrd1 is required for disrupting U3 snoRNA base-pairing interactions in the 5'ETS. We propose that the dynamic U3 snoRNA interactions and Mrd1 are essential for establishing the structure of the central region of 18S that is required for processing and 40S subunit function.
The ChemModSeq next generation sequencing data and processed data (counts for RT stops and read coverage over 35S) is available from the NCBI Gene Expression Omnibus (GEO) under accession number GSE106868. https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE106868