Sno Storm in the Nucleolus: New Roles for Myriad Small RNPs

Abstract

trimethyl guanosine (TMG) cap structure. Phylogenetic comparison of U3 snoRNAs from various species re-Department of Molecular Biophysics and vealed conserved sequence elements called boxes C Biochemistry (UGAUGA/U) and D (CUGA), which are also present in Howard Hughes Medical Institute the next vertebrate snoRNAs (U8 and U13) to be identi-Yale University fied. Boxes C and D direct binding of the abundant New Haven, Connecticut 06536nucleolar protein fibrillarin, a target of autoantibodies, and are necessary for the accumulation and stability of these snoRNAs. Yet, as more snoRNAs were discov-Biologists have known for decades that the nucleolus ered, not all exhibited the hallmarks of U3, U8, and U13: is the compartment of the eukaryotic cell most densely some possessed an unmodified terminal 5Ј monophospacked with RNA. But few would have guessed that, in phate instead of the TMG cap and some lacked the addition to the ribosomal RNAs (rRNAs) at various characteristic box C and D sequences (Table 1). Progstages of maturation, the nucleolus contains a multitude ress in assigning functions was made by deletion/rescue of discrete small RNA molecules. These small nucleolar experiments in yeast and in Xenopus oocytes. Certain RNAs, or snoRNAs, are responsible not only for orches-snoRNAs were found to be essential for growth (U3, trating the cleavage events that cut the long pre-rRNA U14, snR10 [temperature sensitive], and snR30) in yeast into 18S, 5.8S, and 28S molecules, but also for adding and/or for specific cleavage steps in pre-rRNA profinishing touches to rRNAs as they assemble into the cessing (U3, U8, and U22) in Xenopus. Curiously, deleultimate products of the nucleolus, the ribosomal subtion of other yeast snoRNA genes had no detectable units. These finishing touches include remodeling of cereffect on growth rate or rRNA maturation (reviewed by tain rRNA uridines into pseudouridines and tagging of Maxwell and Fournier, 1995). numerous ribose moieties with methyl groups. The A completely unanticipated mode of biogenesis for amazing recent realization is that each of these modifithe particularly small (60-90 nucleotides) and less abuncations is directed by its own specific snoRNA that is dant ‫01ف(‬ 4 copies per cell) members of the vertebrate packaged (as are all small nuclear RNAs) into a ribobox C/D snoRNA family then emerged (reviewed by nucleoprotein (snoRNP) particle. Maxwell and Fournier, 1995). Rather than being tran-Suggesting some important function, methylated and scribed from their own genes, these snoRNAs are intronpseudouridylated residues are confined to the most encoded. Liberated by exonucleolytic processing of exhighly conserved portions of rRNA sequences and are cised introns, these stable intronic fragments possess absent from the discarded regions of pre-rRNA. Verte-5Ј monophosphates instead of TMG caps. SnoRNA host brate rRNAs contain approximately 100 methylated suggenes most commonly specify proteins involved in ars, 95 pseudourdines, and 10 methylated bases (Matranslation or ribosome bi...

Pseudouridines in spliceosomal snRNAs

“…In the 1990s, though, a major breakthrough occurred when a large number of box H/ACA guide RNAs were found to direct rRNA pseudouridylation (Balakin et al, 1996;Ganot et al, 1997;Ni et al, 1997;Smith and Steitz, 1997;Yu et al, 2005). …”

“…About 15 years ago, though, several laboratories began to turn their attention to the mechanisms and functions of these modifications in spliceosomal snRNAs and rRNAs (Bachellerie et al, 1995;Cavaillé et al, 1996;KissLászló et al, 1996;Tycowski et al, 1996;Ganot et al, 1997;Ni et al, 1997;Smith and Steitz, 1997;Tycowski et al, 1998;Yu et al, 1998;Lowe and Eddy, 1999). Multiple effective assays and systems have since been developed for RNA modification research [reviewed in (Wu et al, 2011b)].…”

Two mutant forms of the S1/TPR-containing protein Rrp5p affect the 18S rRNA synthesis in Saccharomyces cerevisiae

“…The nucleolus compartment of eukaryotic cells is devoted to the assembly of large and small ribosomal subunits (reviewed by Mélèse & Xue, 1995)+ The association of almost 80 ribosomal proteins into 60S and 40S subunits occurs concomitantly with the maturation of ribosomal RNA (rRNA) molecules+ The rRNAs have a mixed transcriptional origin+ The nucleolar RNA polymerase I transcribes a single, large rRNA precursor 35S which is processed into three mature rRNAs (18S, 5+8S and 25S)+ The fourth eukaryotic rRNA molecule (5S) is transcribed independently by RNA polymerase III+ The 5S, 5+8S and 25S rRNA species constitute the RNA molecules of mature 60S subunits whereas mature 40S subunits contain only 18S rRNA+ The maturation of the pre-rRNA 35S polymerase I transcript is largely studied in yeast and numerous small nucleolar RNAs (snoRNAs) and protein components involved in this process have been characterized (Eichler & Craig, 1994;Venema & Tollervey, 1995;Tollervey & Kiss, 1997)+ The 35S primary transcript is successively matured by both endonucleolytic and exonucleolytic cleavages that generate stable rRNA precursors whose identification by in vivo labeling or by Northern hybridization has led to the processing pattern drawn in Figure 1+ The nucleic cleavages contribute to a precise excision of both external (59 and 39 ETS) and internal transcribed spacers (ITS1 and ITS2) that interrupt the 35S rRNA sequence+ Aberrations in any of the cleavage steps result in a defective maturation that modifies the reference pattern shown in Figure 1+ In such cases, an increase or decrease in intermediate precursors is evident whereas some aberrant and sometimes deadend pre-rRNA species are detected+ This observation of altered profiles is classically used to determine at which step either a given mutation or the depletion of a cellular component will affect the rRNA maturation process+ These approaches have indeed revealed that a large number of trans-acting factors are required for the same processing step (Venema & Tollervey, 1995)+ For example, the snoRNAs U3, U14, snR30, and snR10 are all essential (or at least important in the case of snR10) for efficient cleavage at sites A 0 , A 1 , and A 2 (Tollervey, 1987;Li et al+, 1990;Hughes & Ares, 1991;Morrissey & Tollervey, 1993; see Fig+ 1B)+ The snoRNAs are believed to function as small nucleolar ribonucleoprotein (snoRNP) complexes rather than as free RNAs (Maxwell & Fournier, 1995) and some nucleolar proteins were shown to be physically associated with snoRNAs (Maxwell & Fournier, 1995;Smith & Steitz, 1997)+ Two RNP particles involved in the yeast rRNA processing are considered in this article: the abovementioned snoRNP complex (Venema & Tollervey, 1995) and the RNase MRP complex …”

Nucleolar localization elements in U8 snoRNA differ from sequences required for rRNA processing

“…The nucleolus is a prominent organelle in the nucleus of eukaryotic cells+ It is the site of ribosome biogenesis, which involves ribosomal RNA (rRNA) synthesis, modification and processing, and association with ribosomal proteins+ Several of these events are mediated by small nucleolar RNAs (snoRNAs)+ There are about 200 snoRNA species in the nucleolus, most of which are used to guide the modifications that occur on pre-rRNA: 29-O-ribose methylation by snoRNA members of the Box C/D family (Cavaillé et al+, 1996;Kiss-László et al+, 1996Maden, 1996;Nicoloso et al+, 1996;Tollervey, 1996;Tycowski et al+, 1996;Maden & Hughes, 1997;Smith & Steitz, 1997), and pseudouridine formation by snoRNA members of the Box H/ACA family (Ganot et al+, 1997;Ni et al+, 1997;Smith & Steitz, 1997)+ The function of these modifications in rRNA is not yet understood, and the snoRNAs utilized for rRNA modifications are dispensable+ In contrast, there are a handful of snoRNAs that are essential for viability of the cell, and are required for rRNA processing events that remove the external transcribed spacers and internal transcribed spacers from the rRNA precursor (summarized in Gerbi, 1995;Maxwell & Fournier, 1995;Sollner-Webb et al+, 1995;Venema & Tollervey, 1995)+ These include U3 (Kass et al+, 1990;Savino & Gerbi, 1990;Hughes & Ares, 1991;Hughes, 1996), U8 (Peculis & Steitz, 1993, U14 (Jarmolowski et al+, 1990;Li et al+, 1990;Li & Fournier, 1992;Liang & Fournier, 1995;Dunbar & Baserga, 1998;Lange et al+, 1998), U22 (Tycowski et al+, 1994), and E1 (ϭU17), E2, and E3 (Mishra & Elicieri, 1997)+ U3 and U14 are used for rRNA processing both in metazoa and in yeast, but the other snoRNAs listed above have only been identified in metazoa+ There are some additional snoRNAs that are essential for rRNA processing that have only been found in yeast+ Of all the snoRNAs that have been identified, U8...…”