mRNA Degradation Rates Are Coupled to Metabolic Status in Mycobacteria

Abstract

The success of Mycobacterium tuberculosis (Mtb) as a human pathogen is due in part to its ability to survive stress conditions, such as hypoxia or nutrient deprivation, by entering non-growing states. In these low-metabolic states, Mtb can tolerate antibiotics and develop genetically encoded antibiotic resistance, making its metabolic adaptation to stress crucial for survival. Numerous bacteria, including Mtb, have been shown to reduce their rates of mRNA degradation under growth limitation and stress. While the existence of this response appears to be conserved across species, the underlying bacterial mRNA stabilization mechanisms remains unknown. To better understand the biology of non-growing mycobacteria, we sought to identify the mechanisms by which mRNA stabilization occurs using the non-pathogenic model Mycobacterium smegmatis. We found that mRNA half-life was responsive to energy stress, with carbon starvation and hypoxia causing global mRNA stabilization. This global mRNA stabilization was rapidly reversed when hypoxia-adapted cultures were re-exposed to oxygen, even in the absence of new transcription. The stringent response and RNase protein levels did not explain mRNA stabilization, nor did transcript abundance. This led us to hypothesize that metabolic changes during growth cessation impact the activity of degradation proteins, increasing mRNA stability. Indeed, bedaquiline and isoniazid, two drugs with opposing effects on cellular energy status, had opposite effects on mRNA half-lives in growth-arrested cells. Taken together, our results indicate that mRNA stability in mycobacteria is not directly regulated by growth status, but rather seems to be dependent on the status of energy metabolism.\n\nIMPORTANCEThe logistics of treating tuberculosis are difficult, requiring multiple drugs for at least six months. Mtb is able to survive within the human host in part by entering non-growing states in which it is metabolically less active, thus rendering it less susceptible to antibiotics. Basic knowledge on how Mtb survives during these low-metabolic states is incomplete, and we postulate that optimized energy resource management -such as transcriptome stabilization--is important for survival. Here we report that mRNA stabilization (increased mRNA half-lives) is a common feature of mycobacteria under stress (e.g. hypoxia and nutrient deprivation) but is not dependent on the mechanisms that have been most often postulated in the literature. Finally, we found that mRNA stability and growth status can be decoupled by a drug that causes growth arrest but increases metabolic activity, indicating that mRNA stability responds to metabolic status rather than to growth rate changes per se. Our findings suggest a need to re-orient the study of global mRNA stabilization to identify novel mechanisms that are presumably responsible.