title: Imp3 unfolds stem structures in pre-rRNA and U3 snoRNA to form a duplex essential for small subunit processing.
abstract: Eukaryotic ribosome biogenesis requires rapid hybridization between the U3 snoRNA and the pre-rRNA to direct cleavages at the A0, A1, and A2 sites in pre-rRNA that liberate the small subunit precursor. The bases involved in hybridization of one of the three duplexes that U3 makes with pre-rRNA, designated the U3-18S duplex, are buried in conserved structures: box A/A' stem-loop in U3 snoRNA and helix 1 (H1) in the 18S region of the pre-rRNA. These conserved structures must be unfolded to permit the necessary hybridization. Previously, we reported that Imp3 and Imp4 promote U3-18S hybridization in vitro, but the mechanism by which these proteins facilitate U3-18S duplex formation remained unclear. Here, we directly addressed this question by probing base accessibility with chemical modification and backbone accessibility with ribonuclease activity of U3 and pre-rRNA fragments that mimic the secondary structure observed in vivo. Our results demonstrate that U3-18S hybridization requires only Imp3. Binding to each RNA by Imp3 provides sufficient energy to unfold both the 18S H1 and the U3 box A/A' stem structures. The Imp3 unfolding activity also increases accessibility at the U3-dependent A0 and A1 sites, perhaps signaling cleavage at these sites to generate the 5' mature end of 18S. Imp4 destabilizes the U3-18S duplex to aid U3 release, thus differentiating the roles of these proteins. Protein-dependent unfolding of these structures may serve as a switch to block U3-pre-rRNA interactions until recruitment of Imp3, thereby preventing premature and inaccurate U3-dependent pre-rRNA cleavage and folding events in eukaryotic ribosome biogenesis.
authors: Binal N
authors: BN
authors: Shah
authors: Xin
authors: X
authors: Liu
authors: Carl C
authors: CC
authors: Correll
mesh_terms: D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012337:RNA, Ribosomal, 18S / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020537:RNA, Small Nucleolar / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C120533:IMP3 protein, S cerevisiae; C120534:IMP4 protein, S cerevisiae; D012322:RNA Precursors; D012331:RNA, Fungal; D012337:RNA, Ribosomal, 18S; D020537:RNA, Small Nucleolar; C117378:RNA, U3 small nucleolar; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins
keywords: RNA conformational switch; ribonucleoprotein; ribosome synthesis; yeast
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