Microbiome characterization by high-throughput transfer RNA sequencing and modification analysis.

Area of Science:

• Microbiology
• Genomics
• Molecular Biology

Background:

• High-throughput sequencing advances microbial insights but struggles with protein translation dynamics.
• Transfer RNAs (tRNAs) are crucial for protein synthesis and offer a tractable target for physiological studies.

Purpose of the Study:

• To develop a direct sequencing method (tRNA-seq) and software (tRNA-seq-tools) for analyzing microbial tRNA sequences, abundance, and modifications.
• To gain physiological insights into microbial communities, particularly in the context of diet.

Main Methods:

• Developed tRNA-seq, a direct sequencing approach for microbial tRNA transcripts.
• Created tRNA-seq-tools for recovering tRNA sequences, abundance profiles, and post-transcriptional modifications.
• Applied tRNA-seq to analyze cecal samples from mice on high-fat and low-fat diets.

Main Results:

• tRNA-seq effectively distinguished microbial communities based on diet, comparable to 16S ribosomal RNA gene amplicon sequencing.
• Identified taxon- and diet-dependent variations in tRNA post-transcriptional modifications.
• Provided taxon-specific, in situ insights into tRNA gene expression dynamics within complex microbiomes.

Conclusions:

• tRNA-seq is a powerful tool for studying microbial physiology and protein translation dynamics.
• Post-transcriptional modifications of tRNAs vary significantly with microbial taxa and host diet.
• This approach enhances our understanding of microbial community function in environmental and host-associated settings.