Post-transcriptional pseudouridylation in mRNA as well as in some major types of noncoding RNAs.

Pseudouridylation is a post-transcriptional isomerization reaction that converts a uridine to a pseudouridine (Ψ) within an RNA chain. Ψ has chemical properties that are distinct from that of uridine and any other known nucleotides. Experimental data accumulated thus far have indicated that Ψ is present in many different types of RNAs, including coding and noncoding RNAs. Ψ is particularly concentrated in rRNA and spliceosomal snRNAs, and plays an important role in protein translation and pre-mRNA splicing, respectively. Ψ has also been found in mRNA, but its function there remains essentially unknown. In this review, we discuss the mechanisms and functions of RNA pseudouridylation, focusing on rRNA, snRNA and mRNA. We also discuss the methods, which have been developed to detect Ψs in RNAs. This article is part of a Special Issue entitled: mRNA modifications in gene expression control edited by Dr. Soller Matthias and Dr. Fray Rupert.

Guide-substrate base-pairing requirement for box H/ACA RNA-guided RNA pseudouridylation.

Box H/ACA RNAs are a group of small RNAs found in abundance in eukaryotes (as well as in archaea). Although their sequences differ, eukaryotic box H/ACA RNAs all share the same unique hairpin-hinge-hairpin-tail structure. Almost all of them function as guides that primarily direct pseudouridylation of rRNAs and spliceosomal snRNAs at specific sites. Although box H/ACA RNA-guided pseudouridylation has been extensively studied, the detailed rules governing this reaction, especially those concerning the guide RNA-substrate RNA base-pairing interactions that determine the specificity and efficiency of pseudouridylation, are still not exactly clear. This is particularly relevant given that the lengths of the guide sequences involved in base-pairing vary from one box H/ACA RNA to another. Here, we carry out a detailed investigation into guide-substrate base-pairing interactions, and identify the minimum number of base pairs (8), required for RNA-guided pseudouridylation. In addition, we find that the pseudouridylation pocket, present in each hairpin of box H/ACA RNA, exhibits flexibility in fitting slightly different substrate sequences. Our results are consistent across three independent pseudouridylation pockets tested, suggesting that our findings are generally applicable to box H/ACA RNA-guided RNA pseudouridylation.

Posttranscriptional RNA Pseudouridylation.

Pseudouridine (Ψ) is the most abundant posttranscriptional modification in noncoding RNAs. Pseudouridines are often clustered in important regions of rRNAs (ribosomal RNAs), snRNAs (small nuclear RNAs), and tRNAs (transfer RNAs), contributing to RNA function. Pseudouridylation is governed by two independent mechanisms. The first involves single protein enzymes called pseudouridine synthases (PUSs) that alone recognize the substrate and catalyze the isomerization of uridine to pseudouridine (RNA-independent pseudouridylation). The second is an RNA-guided pseudouridylation by a family of box H/ACA RNPs (ribonucleoproteins), each of which consists of a unique RNA (box H/ACA RNA) and four common core proteins (Cbf5/NAP57/Dyskerin, Nhp2/L7Ae, Nop10, and Gar1). The RNA component serves as a guide that base pairs with the substrate RNA and directs the enzyme (Cbf5) to carry out the pseudouridylation reaction at a specific site. The crystal structures of many PUSs have been solved in numerous organisms including E. coli and human. Several partial and complete crystal structures of archaea and yeast box H/ACA RNPs are available, providing a rich source of information regarding the molecular interactions between protein components and box H/ACA RNA. Over the years, several experimental systems have been developed to study the mechanism and function of pseudouridylation. Apart from noncoding RNA pseudouridylation, recent experiments have provided evidence of mRNA pseudouridylation as well. Despite remarkable progress, there is a need to accelerate efforts in order to understand the detailed mechanisms and functions of RNA pseudouridylation.