A study of a bacteria commonly used in dairy fermentation revealed how organized RNA helixes, called riboswitches, can combine different signals to keep gene expression in check. Researchers from the University of Michigan analyzed the process by which the binding of manganese (Mn2+) stabilizes a riboswitch from Lactococcus lactis (L. lactis), preventing the formation of a hairpin structure that stops gene transcription. The potential of utilizing central, flexible RNA riboswitches as a molecular fulcrum in a first-class lever system could be harnessed for the development of innovative antibacterial medications.

The research article, “A nascent riboswitch helix orchestrates robust transcriptional regulation through signal integration,” was published in Nature Communications.

Rapid mRNA regulation with riboswitches

Due to competition between different species, bacteria must quickly adapt to temporary nutritional resources and external threats, such as antibiotics and toxins, to survive. Bacterial riboswitches possess the ability to accurately determine the result of gene expression, either during transcription elongation or translation initiation.

A riboswitch typically consists of two intertwined domains: (1) a conserved aptamer region that can identify a specific ligand, causing structural changes, and (2) a less conserved expression platform that regulates the expression of downstream gene(s) after receiving signals from the aptamer. Riboswitches, similar to other structured RNAs, are thought to undergo folding during transcription, influenced by both ligand binding and transcription events. However, the mechanisms by which these processes are coordinated for effective regulation are poorly comprehended.

Riboswitches can control transcription through signal integration

An RNA motif, commonly found in bacterial riboswitches and in various human and plant pathogens, known as yybP-ykoY, plays a significant role in regulating genes related to Mn2+ homeostasis through the control of transcription or translation. The mechanism involves the specific detection of Mn2+ at concentrations below one millimolar, distinguishing it from the more abundant millimolar magnesium ion (Mg2+) and other divalent ions in the cell.

In this study, Adrien Chauvier, PhD, and colleagues from the laboratory of Nils G. Walter, PhD, utilized the extensively studied Mn2+-sensing riboswitch from L. lactis as a model system to examine the influence of the transcription machinery on a complex nascent RNA structure. The process of transcription can take one of two pathways, depending on whether there is a sufficient amount of Mn2+ in the cell’s surroundings.

Researchers at the University of Michigan employed a combination of single-molecule and biochemical methods to ascertain the mechanism by which the P1.1 “switch” helix, a segment of the riboswitch, orchestrates the interactions between a single Mn2+ ion and the transcribing RNA polymerase (RNAP). Without Mn2+, the 3′ segment of P1.1 tends to separate and form a hairpin structure at the terminator, which prevents transcription. The pre-organized P1.1 helix remains stable during riboswitch transcription, enabling it to quickly detect Mn2+ and effectively control gene expression in response to environmental changes. The binding of Mn2+ stabilizes the P1.1 helix, which prevents the formation of an intrinsic terminator hairpin and consequently halts the transcription of the downstream gene.

These findings provide an in-depth explanation of how various intracellular signals influence riboswitch folding, enabling bacterial gene expression to consistently adjust to an ever-changing environment. The mechanistic single-molecule techniques established here will aid in the dissection of many more complex dynamic interactions between transcription and RNA folding, which could exploited in the development of new antibacterial medications.

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