Accurately controlled termination of transcription is critical for gene expression and adaptation to changing environments. The aim of this study was to define transcription termination sites in Mycobacterium tuberculosis at nucleotide resolution. To this end, we applied Term-seq to cultures of M. tuberculosis and established that by far the most abundant mechanism of transcription termination is Rho-dependent. Moreover, we find that conditional or premature termination of transcription regulates a large proportion of genes and that this is often associated with translated upstream open reading frames (uORFs). Finally, our results show that overlapping ORFs are abundant in many cases in the absence of a recognisable Shine-Dalgarno sequence, indicating tightly coordinated/coupled translation. Our results provide a detailed image and comprehensive catalogue of novel M. tuberculosis cis-regulatory elements, where Rho-dependent conditional termination of transcription and translational coupling together play major roles in gene expression control.
Vitamin B12 (B12), an essential cofactor in all domains of life, is produced de novo by only a small subset of prokaryotes, but B12-sensing riboswitches are some of the most widely distributed riboswitches in bacteria.Mycobacterium tuberculosis, the causative agent of the ongoing tuberculosis pandemic, encodes two distinct vitamin B12 riboswitches. One controls the expression ofmetE, encoding a B12-independent methionine synthase, while the other is located upstream ofppe2, a PE/PPE family gene whose function is still unresolved. Here, we analyse ligand sensing, secondary structure architecture, and gene expression control mechanisms of these two riboswitches. Our results provide the first evidence of direct ligand binding bymetEandppe2riboswitches and show that the two switches exhibit different preferences for natural isoforms of B12, use distinct regulatory and structural elements, and act as translational OFF switches. Based on our results, we propose that theppe2switch represents a new Class IIc of B12-sensing riboswitches. Moreover, we have identified small translated open reading frames (uORFs) upstream of bothmetEandppe2, which modulate the expression of the respective downstream genes in opposite directions. Translation of themetEriboswitch uORF suppresses MetE expression, while translation of the uORF in theppe2switch is essential for PPE2 expression via the synthesis of a uORF-PPE2 fusion protein. In summary, our findings reveal an unexpected diversity and complexity of B12-dependent cis-regulation inM. tuberculosis, with potential implications for host-pathogen interactions.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.