Pseudouridine modification in Caenorhabditis elegans spliceosomal snRNAs: unique modifications are found in regions involved in snRNA-snRNA interactions

As a prelude to determining the function of or requirement for this modification in snRNAs, we have mapped the positions of Ψ in U1, U2, U4, U5, and U6 snRNAs from worms using a specific primer extension method. As with other species, C. elegans U2 snRNA has the greatest number of Ψ residues, with nine, located in the 5' half of the U2 snRNA. U5 snRNA has three Ψs, in or near the loop of the large stem-loop that dominates the structure of this RNA. U6 and U1 snRNAs each have one Ψ, and two Ψ residues were found in U4 snRNA.The total number of Ψs found in the snRNAs of C. elegans is significantly higher than the minimal amount found in yeasts but it is lower than that seen in sequenced vertebrate snRNAs. When the actual sites of modification on C. elegans snRNAs are compared with other sequenced snRNAs most of the positions correspond to modifications found in other species. However, two of the positions modified on C. elegans snRNAs are unique, one at position 28 on U2 snRNA and one at position 62 on U4 snRNA. Both of these modifications are in regions of these snRNAs that interact with U6 snRNA either in the spliceosome or in the U4/U6 small nuclear ribonucleoprotein particle (snRNP) and the presence of Ψ may be involved in strengthening the intermolecular association of the snRNAs.Pseudouridine (Ψ) is an abundant modified nucleoside found in RNA, at one time considered the fifth nucleoside of RNA [1]. This modification has been found in many types of RNA and is particularly abundant in small stable RNAs such as transfer RNA (tRNA) and small nuclear RNA (snRNA) [2,3]. Because of its structure, with the carbon at the 5 position of the uracil ring attached to the sugar rather than the nitrogen at the 1 position, Ψ is potentially more versatile in its hydrogen bonding interactions [4]. The presence of Ψ appears to strengthen stems in RNA secondary structures and to stabilize base stacking in loops [5-7]. The splicing of pre-messenger RNAs (pre-mRNAs) in Xenopus oocyte