Inability of Prevotella bryantii to Form a Functional Shine-Dalgarno Interaction Reflects Unique Evolution of Ribosome Binding Sites in Bacteroidetes

Abstract

The Shine-Dalgarno (SD) sequence is a key element directing the translation to initiate at the authentic start codons and also enabling translation initiation to proceed in 5′ untranslated mRNA regions (5′-UTRs) containing moderately strong secondary structures. Bioinformatic analysis of almost forty genomes from the major bacterial phylum Bacteroidetes revealed, however, a general absence of SD sequence, drop in GC content and consequently reduced tendency to form secondary structures in 5′-UTRs. The experiments using the Prevotella bryantii TC1-1 expression system were in agreement with these findings: neither addition nor omission of SD sequence in the unstructured 5′-UTR affected the level of the reporter protein, non-specific nuclease NucB. Further, NucB level in P. bryantii TC1-1, contrary to hMGFP level in Escherichia coli, was five times lower when SD sequence formed part of the secondary structure with a folding energy -5,2 kcal/mol. Also, the extended SD sequences did not affect protein levels as in E. coli. It seems therefore that a functional SD interaction does not take place during the translation initiation in P. bryanttii TC1-1 and possibly other members of phylum Bacteroidetes although the anti SD sequence is present in 16S rRNA genes of their genomes. We thus propose that in the absence of the SD sequence interaction, the selection of genuine start codons in Bacteroidetes is accomplished by binding of ribosomal protein S1 to unstructured 5′-UTR as opposed to coding region which is inaccessible due to mRNA secondary structure. Additionally, we found that sequence logos of region preceding the start codons may be used as taxonomical markers. Depending on whether complete sequence logo or only part of it, such as information content and base proportion at specific positions, is used, bacterial genera or families and in some cases even bacterial phyla can be distinguished.

Defining the Bacteroides Ribosomal Binding Site

“…Our results, though more detailed, are in keeping with the findings by Accetto and Avgustin (20), who used the total gene contents of the organisms for their logos, containing alien as well as highly and moderately expressed genes. This explains why our logos are more extensive and detailed, as ribosomal genes are predicted to be among the most highly expressed genes and show high levels of occurrence of strong SD sequences (40), therefore providing a better starting point for identifying strong ribosomal binding sites through a sequence logo approach.…”

“…Some authors have suggested that translation initiation in the Bacteroidetes phylum is facilitated solely through ribosomal protein S1 (RPS1) (20,45). It has been demonstrated that translational enhancers derived from the tobacco mosaic virus, whose sequence fits the binding criteria for RPS1 (no guanosine) (46), can replace the SD sequence in a variety of Gram-negative bacteria (47).…”

“…This is seemingly akin to Gram-positive bacteria such as Bacillus subtilis, which have more-stringent criteria for their translation initiation signals and require a higher degree of complementarity to the 3= end of the 16S rRNA (19). Recently, Accetto and Avgustin (20) noted from the genome sequences of several representatives of the phylum Bacteroidetes that there is enrichment in adenine and thymine in the 5= untranslated region (UTR) of genes. Furthermore, they reported an inability of Prevotella bryantii (a member of the order Bacteroidales) to form a functional Shine-Dalgarno interaction, although it was somewhat surprising that the best-performing 5= UTR contained a perfect core SD sequence (5=-GGAGG-3=).…”

Polysaccharide utilization locus and CAZYme genome repertoires reveal diverse ecological adaptation of Prevotella species

“…The Bacteroidetes are special in several ways: the base composition and spacing of their promoters are distinct [6], the Shine-Dalgarno sequence is not used in translation initiation [1], and their capability to degrade a wide array of carbohydrates depends on the starch utilization system-like (Sus-like) multiprotein complexes that bind and partially degrade substrates prior to their transport into the periplasm where final breakdown occurs [37]. The central elements of the Sus-like systems are encoded by the paralogues of Bacteroides thetaiotaomicron VPI-5482 susC and susD whose products were shown to account for >60% of starch binding, while SusC also transfers oligosaccharides across the outer membrane [37].…”

Noncanonical Translation Initiation Comes of Age

“…Some leadered mRNAs lack recognizable SD-like sequences in their untranslated regions (UTRs). Moreover, analyses of sequenced genomes indicate that the SD-aSD interaction is completely absent in some archaea and bacteria (10,11). Among the alternative initiation mechanisms, proposals involving mRNA-rRNA complementarity outside the aSD region of 16S rRNA have, in general, not withstood scrutiny (12,13).…”