Cajal body-specific small nuclear RNAs: a novel class of 2'-O-methylation and pseudouridylation guide RNAs

Abstract

Cajal (coiled) bodies are conserved subnuclear organelles that are present in the nucleoplasm of both animal and plant cells. Although Cajal bodies were ®rst described nearly 100 years ago, their function has remained largely speculative. Here, we describe a novel class of human small nuclear RNAs that localize speci®cally to Cajal bodies. The small Cajal bodyspeci®c RNAs (scaRNAs) are predicted or have already been demonstrated to function as guide RNAs in site-speci®c synthesis of 2¢-O-ribose-methylated nucleotides and pseudouridines in the RNA polymerase II-transcribed U1, U2, U4 and U5 spliceosomal small nuclear RNAs (snRNAs). Our results provide strong support for the idea that the Cajal body, this mysterious nuclear organelle, provides the cellular locale for post-transcriptional modi®cation of spliceosomal snRNAs.

Small Cajal Body–specific RNAs ofDrosophilaFunction in the Absence of Cajal Bodies

“…Specific evidence for involvement of the CB in snRNA modification came from the discovery of U85 scaRNA (Jády and Kiss, 2001) and the demonstration that it and other scaRNAs are localized in the CB of HeLa cells (Darzacq et al, 2002;Jády et al, 2003;Richard et al, 2003). Localization in the CB is mediated by an evolutionarily conserved sequence (UGAG), referred to as the CB-specific localization sequence or CAB box.…”

“…Modification takes place in the nucleus after import of the U5 snRNA from the cytoplasm . CB localization of U85 scaRNA was originally demonstrated by fluorescent in situ hybridization (FISH) in mammalian and Drosophila cultured cells (Darzacq et al, 2002;Richard et al, 2003), and subsequently in various tissues of the fly (Liu et al, 2006b(Liu et al, , 2009.…”

“…However, the guide RNAs that mediate modification of the snRNAs are preferentially found in another nuclear organelle, the Cajal body (CB). These guide RNAs are called small CB-specific RNAs (scaRNAs).The first scaRNA to be studied in detail was U85 scaRNA (Jády and Kiss, 2001;Darzacq et al, 2002). U85 is a remarkable double guide RNA that contains both box C/D and box H/ACA motifs.…”

“…Modification takes place in the nucleus after import of the U5 snRNA from the cytoplasm . CB localization of U85 scaRNA was originally demonstrated by fluorescent in situ hybridization (FISH) in mammalian and Drosophila cultured cells (Darzacq et al, 2002;Richard et al, 2003), and subsequently in various tissues of the fly (Liu et al, 2006b(Liu et al, , 2009.CBs are small nuclear bodies found in a wide variety of eukaryotic organisms, including mammals, amphibians, insects, and plants. They are most often recognized by immunostaining for the marker protein coilin Raska et al, 1991) and by their unusually high concentration of splicing snRNPs relative to the rest of the nucleus (Carmo-Fonseca et al, 1991;Matera and Ward, 1993;Spector, 1993).…”

The C-terminal domain of coilin interacts with Sm proteins and U snRNPs

“…These 2′-O-methyl sugar-and pseudouridine-base modifications of the U snRNPs (Darzacq et al 2002;Jady et al 2003;Verheggen et al 2002) are guided by small CB-specific RNAs (scaRNAs) and are crucial for proper pre-mRNA splicing in vivo (Yu et al 1998). …”