Global fitness landscapes of the Shine-Dalgarno sequence.

Shine-Dalgarno sequences (SD) in prokaryotic mRNA facilitate protein translation by pairing with rRNA in ribosomes. Although conventionally defined as AG-rich motifs, recent genomic surveys reveal great sequence diversity, questioning how SD functions. Here, we determined the molecular fitness (i.e., translation efficiency) of 49 synthetic 9-nt SD genotypes in three distinct mRNA contexts in Escherichia coli We uncovered generic principles governing the SD fitness landscapes: (1) Guanine contents, rather than canonical SD motifs, best predict the fitness of both synthetic and endogenous SD; (2) the genotype-fitness correlation of SD promotes its evolvability by steadily supplying beneficial mutations across fitness landscapes; and (3) the frequency and magnitude of deleterious mutations increase with background fitness, and adjacent nucleotides in SD show stronger epistasis. Epistasis results from disruption of the continuous base pairing between SD and rRNA. This "chain-breaking" epistasis creates sinkholes in SD fitness landscapes and may profoundly impact the evolution and function of prokaryotic translation initiation and other RNA-mediated processes. Collectively, our work yields functional insights into the SD sequence variation in prokaryotic genomes, identifies a simple design principle to guide bioengineering and bioinformatic analysis of SD, and illuminates the fundamentals of fitness landscapes and molecular evolution.

The diversity of Shine-Dalgarno sequences sheds light on the evolution of translation initiation.

Shine-Dalgarno (SD) sequences, the core element of prokaryotic ribosome-binding sites, facilitate mRNA translation by base-pair interaction with the anti-SD (aSD) sequence of 16S rRNA. In contrast to this paradigm, an inspection of thousands of prokaryotic species unravels tremendous SD sequence diversity both within and between genomes, whereas aSD sequences remain largely static. The pattern has led many to suggest unidentified mechanisms for translation initiation. Here we review known translation-initiation pathways in prokaryotes. Moreover, we seek to understand the cause and consequence of SD diversity through surveying recent advances in biochemistry, genomics, and high-throughput genetics. These findings collectively show: (1) SD:aSD base pairing is beneficial but nonessential to translation initiation. (2) The 5' untranslated region of mRNA evolves dynamically and correlates with organismal phylogeny and ecological niches. (3) Ribosomes have evolved distinct usage of translation-initiation pathways in different species. We propose a model portraying the SD diversity shaped by optimization of gene expression, adaptation to environments and growth demands, and the species-specific prerequisite of ribosomes to initiate translation. The model highlights the coevolution of ribosomes and mRNA features, leading to functional customization of the translation apparatus in each organism.