Chapter 15 : Regulatory Aspects of rRNA Modification and Pre-rRNA Processing

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The major types of posttranscriptional modification are the isomerization of uracil to pseudouridine, 2′-O-methylation of the ribose moieties and base methylation. In addition to their roles in pre-rRNA modification, both Dim1p and Cbf5p are required for pre-rRNA processing, and Pet56p is required for synthesis of mitochondrial large ribosomal subunits. This chapter discusses data on Dim1p and Cbf5p that provide insights on the type of systems that may act to coordinate the many steps in ribosome synthesis and the origins of the snoRNA-directed rRNA modification systems present in eukaryotic cells. Uncoupling of the dimethylation and prerRNA processing defects showed that Dimlp rather than the dimethylation activity is required for prerRNA processing. This observation did not, however, determine whether Dim1p is itself directly required for pre-rRNA processing. The binding of Dim1p to the pre-rRNA may therefore be monitored to ensure that it fulfills its functions in both modification and assembly. Ribosome synthesis is a highly dynamic process during which a vast number of processing, modification, and assembly reactions occur simultaneously. The requirement for Cbf5p in pre-rRNA processing appears to be quite different from the requirement for Dim1p. Eukaryotic ribosome synthesis involves very complex pre-rRNA processing and assembly pathways. These include numerous steps occurring in different cellular compartments and requiring a plethora of RNA and proteins, many of them in the form of snoRNPs that only transiently associate with the preribosomal particles. The archaea may hold the final clues to understanding the origins of the small nucleolar ribonucleoproteins (snoRNPs)-directed systems of rRNA modification.

Citation: Lafontaine D, Tollervey D. 1998. Regulatory Aspects of rRNA Modification and Pre-rRNA Processing, p 281-288. In Grosjean H, Benne R (ed), Modification and Editing of RNA. ASM Press, Washington, DC. doi: 10.1128/9781555818296.ch15
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Image of Figure 1
Figure 1

Structure of the yeast pre-rRNA and its processing pathway. (A) The 35S pre-rRNA. The sequences encoding the mature 18S, 5.8S and 25S rRNAs (thick lines) are flanked by the 5′ and 3′ external transcribed spacers (5′ ETS and 3′ ETS) and separated by internal transcribed spacers 1 and 2 (ITS1 and ITS2). Sites of pre-rRNA processing are indicated with uppercase letters (A to E). The site of dimethylation is represented by m A. (B) The pre-rRNA processing pathway. Pseudouridine formation occurs shortly after or during transcription. Pseudouridine synthesis is targeted by the Η + ACA snoRNAs and requires Cbf5p assisted by Gar1p. Processing of the primary 35S precursor starts at site A, yielding the 33S pre-rRNA. This molecule is subsequently processed at sites A and A, giving rise successively to the 32S pre-rRNA and to the 20S and 27SsA precursors. Cleavage at A separates the pre-rRNAs destined for the small and large ribosomal subunit. The 20S precursor is dimethylated by Dim1p and then cleaved endonucleolytically at site D to yield the mature 18S rRNA. The 27SA precursor is processed by two alternative pathways to form the mature 5.8S and 25S rRNAs. The major pathway involves cleavage at a second site in ITS1, A, rapidly followed by exonucleolytic digestion to site B1, generating the 27SB precursor. Approximately 15% of the 27SA molecules are processed by an alternative pathway at site B1, producing the 27SB pre-rRNA. At the same time as processing at B1 is completed, the 3′-end of mature 25S rRNA is generated by processing at site B2. The subsequent processing of both 27SB species appears to follow a similar pathway. Cleavage at sites C and C releases the mature 25S rRNA and the 7S pre-rRNAs, which undergo rapid 3′→5′ exonuclease digestion to site E, generating the mature 3′-end of 5.8S rRNA. Cbf5p and Dim1p are required for the early cleavages at sites A and A; loss of these cleavages inhibits formation of the 20S and 27SA pre-rRNAs, preventing synthesis of 18S rRNA. In addition, Cbf5p is required for efficient processing at site A and efficient processing of the 27SB and 7S pre-rRNAs in ITS2.

Citation: Lafontaine D, Tollervey D. 1998. Regulatory Aspects of rRNA Modification and Pre-rRNA Processing, p 281-288. In Grosjean H, Benne R (ed), Modification and Editing of RNA. ASM Press, Washington, DC. doi: 10.1128/9781555818296.ch15
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Image of Figure 2
Figure 2

3′-end of the SSU-rRNA in and yeast cytoplasmic ribosomes. Divergent nucleotides are boxed. A precise deletion of the anti-Shine-Dalgarno box (CCUCC) has occurred in the eukaryotic SSU-rRNA. The twin adenosine substrates (1518–1519 in and 1779–1780 in yeast) of Dim1p are universally conserved.

Citation: Lafontaine D, Tollervey D. 1998. Regulatory Aspects of rRNA Modification and Pre-rRNA Processing, p 281-288. In Grosjean H, Benne R (ed), Modification and Editing of RNA. ASM Press, Washington, DC. doi: 10.1128/9781555818296.ch15
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Figure 3

Pre-rRNA processing in the wild type (A) and (B) and (C) mutants. (A) Simplified version of the wild-type pre-rRNA processing pathway, as described in Fig. 1 . (B) strains and strains are inhibited in cleavage at sites A and A. Consequently, the 22S pre-rRNA accumulates and no 18S rRNA is made. The 27SA pre-rRNA is normally processed to 25S and 5.8S rRNAs. (C) strains are inhibited in cleavage at sites A, A and A. In this case the 23S pre-rRNA is accumulated and no 18S rRNA is made. The 27SA pre-rRNA can be processed to 25S and 5.8S rRNAs, although processing of the 27SB and 7S pre-rRNAs is delayed.

Citation: Lafontaine D, Tollervey D. 1998. Regulatory Aspects of rRNA Modification and Pre-rRNA Processing, p 281-288. In Grosjean H, Benne R (ed), Modification and Editing of RNA. ASM Press, Washington, DC. doi: 10.1128/9781555818296.ch15
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