A possible tertiary rRNA interaction between expansion segments ES3 and ES6 in eukaryotic 40S ribosomal subunits

Abstract

Eukaryotic 16S-like ribosomal RNAs contain 12 so-called expansion segments, i.e., sequences not included in the RNA secondary structure core common to eubacteria, archaea, and eukarya. Two of these expansion segments, ES3 and ES6, are juxtaposed in the recent three-dimensional model of the eukaryotic 40S ribosomal subunit. We have analyzed ES3 and ES6 sequences from more than 2900 discrete eukaryotic species, for possible sequence complementarity between the two expansion segments. The data show that ES3 and ES6 could interact by forming a helix consisting of seven to nine contiguous base pairs in almost all analyzed species. We, therefore, suggest that ES3 and ES6 form a direct RNA-RNA contact in the ribosome.Eukaryotic 18S ribosomal RNA is considerably longer than its prokaryotic homolog 16S rRNA (Clark 1987). Despite differences in size and sequence, 18S rRNA and 16S rRNA share a common structural core (Gutell et al. 1985) in which the additional nucleotides found in 18S rRNA are inserted at specific positions. The inserted nucleotides form extra sequence elements called variable regions (Neefs and De Wachter 1990) or expansion segments (ES; Gerbi 1996). The length and sequence of these expansion segments varies considerably between organisms.18S rRNA contains 12 expansion segments (Gerbi 1996). One of these expansion segments, referred to as ES6, is located in the central domain of 18S rRNA (Fig. 1). With an average length of 250 nucleotides (Neefs and De Wachter 1990), this is the largest expansion segment found in 18S rRNA. ES6 can be divided in two halves based on sequence variability. The 5Ј half exhibits extensive variability, whereas the sequence of the 3Ј half is more conserved. Sequences corresponding to the latter half are absent in eubacterial and archaeal 16S rRNA (Cannone et al. 2002). Several attempts to construct a secondary structure model for the 3Ј half have been made (Nickrent and Sargent 1991;Hancock and Vogler 1998;Wuyts et al. 2000). However, because of the sequence conservation, structure prediction is difficult, and this part of ES6 is therefore left unstructured in the phylogenetic models of 18S rRNA secondary structure ( Fig. 1; Cannone et al. 2002).The location of ES6 within the three-dimensional structure of the yeast 40S subunit has recently been determined using cryo-electron microscopy (Spahn et al. 2001). The expansion segment is located on the back of the so-called body of the 40S subunit (Fig. 1, inset). The segment is seen as two separate densities, one positioned across the back of the 40S body, whereas the second density is located at the side of the lower part of the body with its lower end close to the so-called left foot (Spahn et al. 2001).The left foot contains another expansion segment, called ES3 (Spahn et al. 2001), only found in eukaryotes (Cannone et al. 2002). ES3 is very variable in size and sequence (Gutell et al. 1985;Wuyts et al. 2002), but the phylogenetic data indicate a similar basic structure for ES3 in different organisms ( Fig. 1; Cannone et al...

Secondary structure of two regions in expansion segments ES3 and ES6 with the potential of forming a tertiary interaction in eukaryotic 40S ribosomal subunits

“…Figure 3B summarizes the experimental results obtained in isolated intact wheat ribosomes. As seen in the figure, the two sequences in ES3 and ES6 indicated to be in direct contact (Alkemar and Nygard 2003) were only accessible for double-strand-specific modification.…”

“…The two expansion segments contain a complementary sequence motif that allows formation of a 7-9-bp helix in most eukaryotes (Alkemar and Nygard 2003). Here we have analyzed the secondary structure of the two regions in ES3 and ES6 to determine the possibility of a tertiary interaction between these two eukaryote-specific sequences in intact ribosomes.…”

“…The nucleotides are numbered in accordance with Figure 2. The proposed tertiary interaction between sequences in yeast ES3 and ES6 is based on the consensus sequences (Alkemar and Nygard 2003).…”

“…We have recently suggested that the two juxtaposed expansion segments ES3 and ES6 may be in direct contact in the 40S subunit (Alkemar and Nygard 2003). This suggestion was based on a comparative sequence analysis of ES3 and ES6 sequences from six eukaryotic taxa, representing >2900 discrete species.…”

“…This suggestion was based on a comparative sequence analysis of ES3 and ES6 sequences from six eukaryotic taxa, representing >2900 discrete species. The sequence analysis showed that ES3 and ES6 contained two complementary sequence motifs with the consensus sequences AAAACCAAU and GUUG-GUUUU, respectively, that allowed formation of a helix containing seven to nine contiguous base pairs in ∼95% of the species analyzed (Alkemar and Nygard 2003).…”