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Multiple metal-binding cores are required for metalloregulation by M-box riboswitch RNAs.

ABSTRACT: Riboswitches are regulatory RNAs that control downstream gene expression in response to direct association with intracellular metabolites or metals. Typically, riboswitch aptamer domains bind to a single small-molecule metabolite. In contrast, an X-ray crystallographic structural model for the M-box riboswitch aptamer revealed the absence of an organic metabolite ligand but the presence of at least six tightly associated magnesiums. This observation agrees well with the proposed role of the M-box riboswitch in functioning as a sensor of intracellular magnesium, although additional nonspecific metal interactions are also undoubtedly required for these purposes. To gain greater functional insight into the metalloregulatory capabilities of M-box RNAs, we sought to determine whether all or a subset of the RNA-chelated magnesium ions were required for riboswitch function. To accomplish this task, each magnesium-binding site was simultaneously yet individually perturbed through random incorporation of phosphorothioate nucleotide analogues, and RNA molecules were investigated for their ability to fold in varying levels of magnesium. These data revealed that all of the magnesium ions observed in the structural model are important for magnesium-dependent tertiary structure formation. Additionally, these functional data revealed a new core of potential metal-binding sites that are likely to assist formation of key tertiary interactions and were previously unobserved in the structural model. It is clear from these data that M-box RNAs require specific binding of a network of metal ions for partial fulfillment of their metalloregulatory functions.

SUBMITTER: Wakeman CA

PROVIDER: S-EPMC2755058 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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Multiple metal-binding cores are required for metalloregulation by M-box riboswitch RNAs.

Wakeman Catherine A CA Ramesh Arati A Winkler Wade C WC

Journal of molecular biology 20090717 3

Riboswitches are regulatory RNAs that control downstream gene expression in response to direct association with intracellular metabolites or metals. Typically, riboswitch aptamer domains bind to a single small-molecule metabolite. In contrast, an X-ray crystallographic structural model for the M-box riboswitch aptamer revealed the absence of an organic metabolite ligand but the presence of at least six tightly associated magnesiums. This observation agrees well with the proposed role of the M-bo ...[more]

PMID: 19619558

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Project description:T-box riboswitches are a widespread class of structured noncoding RNAs in Gram-positive bacteria that regulate the expression of amino acid-related genes. They form negative feedback loops to maintain steady supplies of aminoacyl-transfer RNAs (tRNAs) to the translating ribosomes. T-box riboswitches are located in the 5' leader regions of mRNAs that they regulate and directly bind to their cognate tRNA ligands. T-boxes further sense the aminoacylation state of the bound tRNAs and, based on this readout, regulate gene expression at the level of transcription or translation. T-box riboswitches consist of two conserved domains-a 5' Stem I domain that is involved in specific tRNA recognition and a 3' antiterminator/antisequestrator (or discriminator) domain that senses the amino acid on the 3' end of the bound tRNA. Interaction of the 3' end of an uncharged but not charged tRNA with a thermodynamically weak discriminator domain stabilizes it to promote transcription readthrough or translation initiation. Recent biochemical, biophysical, and structural studies have provided high-resolution insights into the mechanism of tRNA recognition by Stem I, several structural models of full-length T-box-tRNA complexes, mechanism of amino acid sensing by the antiterminator domain, as well as kinetic details of tRNA binding to the T-box riboswitches. In addition, translation-regulating T-box riboswitches have been recently characterized, which presented key differences from the canonical transcriptional T-boxes. Here, we review the recent developments in understanding the T-box riboswitch mechanism that have employed various complementary approaches. Further, the regulation of multiple essential genes by T-boxes makes them very attractive drug targets to combat drug resistance. The recent progress in understanding the biochemical, structural, and dynamic aspects of the T-box riboswitch mechanism will enable more precise and effective targeting with small molecules. © 2019 IUBMB Life, 2019 © 2019 IUBMB Life, 71(8):1167-1180, 2019.

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Multiple metal-binding cores are required for metalloregulation by M-box riboswitch RNAs.

Publications

Multiple metal-binding cores are required for metalloregulation by M-box riboswitch RNAs.

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