Equally parsimonious pathways through an RNA sequence space are not equally likely

Area of Science:

• Molecular Biology
• Evolutionary Biology
• Bioinformatics

Background:

• 5S ribosomal RNA (rRNA) plays a crucial role in protein synthesis within Escherichia coli.
• Understanding the evolutionary pathways of functional RNA sequences is key to deciphering molecular evolution.
• Previous work established an experimental system to test 5S rRNA functionality in vivo.

Purpose of the Study:

• To investigate the functional validity of intermediate sequences along evolutionary trajectories of 5S rRNA.
• To determine if evolutionary paths through RNA sequence space are constrained by functional requirements.
• To assess the complexity of interdependencies within RNA sequence space.

Main Methods:

• Utilized a previously developed experimental system to test 5S rRNA functionality in Escherichia coli.
• Analyzed alternative, equally parsimonious evolutionary trajectories connecting known functional 5S rRNA sequences.
• Examined the in vivo validity of all sequence intermediates along these trajectories.

Main Results:

• Out of 20 intermediate sequences examined across two trajectories, only eight were found to be clearly invalid.
• Seven of 30 possible evolutionary paths consisted exclusively of functionally valid sequences.
• The validity of some intermediate sequences was unpredictable based on current structural knowledge, indicating complex sequence-structure-function relationships.

Conclusions:

• A significant proportion of intermediate sequences along plausible evolutionary paths are functionally valid, supporting the idea that RNA evolution is constrained by function.
• The complexity of RNA sequence space and its interdependencies may be underestimated.
• Ancestral RNA sequences, when inferred using parsimony, should also meet functional validity criteria, which can be experimentally tested.