The “Fifth” RNA Nucleotide: A Role for Ribosomal RNA Pseudouridylation in Control of Gene Expression at the Translational Level - Biophysical Approaches to Translational Control of Gene Expression - page 251

snoRNP components and their function, please refer to the following reviews
(Filipowicz and Pogacic 2002 ; Ishitani et al. 2008 ; Reichow et al. 2007 ) . It is clear
that significant advances have been made in uncovering the components implicated
in methylation of rRNA. However, how distinct methyl modifications affect ribo-
some function in eukaryotic cells, and more importantly, how this process is regu-
lated remains poorly understood. For example, from a mechanistic viewpoint, it
remains to be addressed whether methylation imposes conformational changes in
rRNA, influences ribosome composition, and/or affects RNA-RNA or RNA-protein
binding. Complementary to studies investigating rRNA methylation, there has been
a wealth of research focused on how Y, one of the most abundant base modifications
among diverse RNA species including transfer RNA (tRNA), rRNA, and small
nuclear RNA (snRNA), modulates RNA function (Ofengand 2002 ) . In the remain-
der of this chapter, we will focus on the biophysical approaches employed to iden-
tify the chemical properties and functions of Y residues in modulating gene
expression at the translation level.

13.2.2 Pseudouridylation of rRNA

13.2.2.1 Structure and Properties of Pseudouridine

Often referred to as the “fifth RNA nucleotide,” Y was fi rst identi fi ed by chromato-
graphic analysis of tRNA from baking yeast by the Allen laboratory in 1957 (Davis
and Allen 1957 ). The chemical structure of Y, a 5-ribosyl isomer of uridine, was
subsequently solved and characterized by the Allen and Cohn laboratories (Cohn
1959 ; Yu and Allen 1959 ). Distinct from the four canonical RNA nucleotides, Y is
the only nucleotide with a C-C rather than an N-C glycosyl bond (Fig. 13.1 ). It has
been proposed that the presence of a free N1-H provides an additional hydrogen
bond donor site (d), which may be exploited for novel RNA-RNA and RNA-protein
interactions. NMR studies indicate that Y is predominantly found in the anti
configuration in RNA, providing the ideal spatial arrangement for the formation of
water bridges that connect the N1-H group of the Y to the backbone phosphate of
the preceding residue (Davis et al. 1998 ; Yarian et al. 1999 ) . The water bridge con-
fers 5¢ rigidity, which restricts the mobility of the RNA backbone at the site of
pseudouridylation, thus stabilizing the RNA and, in particular, its folding domains
(Arnez and Steitz 1994 ; Davis 1995 ). Pseudouridylation of RNA also appears to be
critical for stabilizing RNA-RNA binding by influencing the rigidity of the sugar-
phosphate backbone and enhancing base stacking (reviewed in Charette and Gray
2000 ). In this respect, it is not surprising that Y has been referred to as the “molecu-
lar glue” that reinforces necessary RNA conformation (Ofengand 2002 ) . Overall, it
seems likely that specific conformational changes in the tertiary structure of rRNA,
imposed by Y modifications, may stabilize rRNA, affect its binding to RNAs and/
or proteins, and therefore impact the overall structure of the ribosome.