Pervasive, non-functional translation in mycobacterium tuberculosis

Open reading frame (ORF) boundaries in bacterial genomes have largely been drawn by gene prediction algorithms. However, these algorithms often fail to predict ORFs with non-canonical features, including those that are short, overlapping, or lack 5' UTRs. Recent developments in genome-scale mapping of translation have facilitated the empirical identification of open reading frames (ORFs). Here, we use ribosome profiling approaches to map initiating and elongating ribosomes in Mycobacterium tuberculosis. Thus, we identify over 1,000 novel ORFs, revealing that much of the M. tuberculosis genome encodes proteins in overlapping reading frames, and/or on both strands. Most of the novel ORFs are short (sORFs), impeding their identification by traditional methods. The strong codon bias that characterizes annotated mycobacterial ORFs is not evident in the aggregate novel sORFs, and hence most are unlikely to encode functional proteins. Thus, our data suggest that bacterial transcriptomes are subject to pervasive translation that occurs as a result of the relatively low specificity requirements of initiating ribosomes. We speculate that the inefficiency of expressing spurious sORFs may be offset by positive contributions to M. tuberculosis biology through cis and trans regulatory activities of a small subset.