The riboswitch-mediated control of sulfur metabolism in bacteria

Abstract

Many operons in Gram-positive bacteria that are involved in methionine (Met) and cysteine (Cys) biosynthesis possess an evolutionarily conserved regulatory leader sequence (S-box) that positively controls these genes in response to methionine starvation. Here, we demonstrate that a feed-back regulation mechanism utilizes S-adenosyl-methionine as an effector. S-adenosyl-methionine directly and specifically binds to the nascent S-box RNA, causing an intrinsic terminator to form and interrupt transcription prematurely. The S-box leader RNA thus expands the family of newly discovered riboswitches, i.e., natural regulatory RNA aptamers that seem to sense small molecules ranging from amino acid derivatives to vitamins.M odulation of the structures of leader transcripts is a common mode of regulation of synthetic and metabolic operons in prokaryotes. Switching between two alternative RNA structures, with one causing transcription termination, is made in response to some external event, e.g., ribosome pausing (in Gram-negative bacteria), binding of a regulatory protein, or tRNA (in Gram-positive bacteria) (reviewed in ref. 1). Recently, three regulons responsible for vitamin biosynthesis have been shown to use flavinmononucleotide, thiamin pyrophosphate (TPP), and adenosyl-cobalamin to directly bind their cognate leader RNAs, changing their structures and functions (2-5). Such leader RNA aptamers were called riboswitches to reflect their unique ability to switch between two conformations in response to binding of a small molecule without the help of proteins (3, 5, 6). The evolutionarily conserved sequences encoding vitamin-sensing riboswitches have been found in the genomes of many Gram-positive and Gram-negative bacteria (7-10), and also in some archaea (10), fungi, and plant species (A.S.M. and E.N., unpublished observations). In this work, we tested the hypothesis that riboswitches are not just a peculiar attribute of vitamin biosynthetic operons but are more common in nature and are also involved in control of amino acid biosynthesis.Grundy and Henkin (11) found that at least 11 transcription units in Bacillus subtilis that are mostly involved in Cys and Met synthesis possess a leader element that includes an intrinsic transcription terminator, competing antiterminator, and a conserved element (S-box) that functions as an anti-antiterminator. Based on their genetic and physiological studies, the authors postulated that the S-box serves as a target for repression during growth in the presence of Met (11,12). Northern blot experiments substantiated a model of S-box gene regulation by transcription antitermination in response to Met (13). It has also been shown that overexpression of S-adenosyl-methionine (SAM) synthase leads to Met auxotrophy in B. subtilis, suggesting that SAM is an effector of Met biosynthesis in this bacteria (14). This notion was corroborated recently. The transcription profile of S-box genes in several Met auxotroph strains of B. subtilis was analyzed by using DNA arrays. The results argued that...

Confirmation of a second natural preQ1 aptamer class in Streptococcaceae bacteria

“…Four different aptamer classes recognizing Sadenosylmethionine (AdoMet or SAM) have been identified to date. SAM-I or S-box (Epshtein et al 2003;McDaniel et al 2003;Winkler et al 2003), SAM-II (Corbino et al 2005), SAM-III or SMKbox (Fuchs et al 2006), and SAM-IV ) often regulate the same genes in different organisms (e.g., SAM synthase, homoserine O-succinyltransferase). The discovery of a second natural preQ 1 aptamer class shows that SAM is not a special case and that additional metabolites may be bound by multiple distinct RNA architectures to control similar sets of genes in different organisms.…”

Time-resolved NMR methods resolving ligand-induced RNA folding at atomic resolution

“…In general, the energetic difference between alternate RNA conformations is very small, and the equilibrium distribution is strongly affected through the binding of proteins (6), ions (7), or small metabolites (8)(9)(10), or by structural modifications (11). Alternate RNA structures are stabilized by different Watson-Crick base-pairing interactions (12).…”

S -adenosylmethionine directly inhibits binding of 30S ribosomal subunits to the S MK box translational riboswitch RNA

“…Included in this regulon is the metK gene, which encodes S-adenosylmethionine (SAM) synthetase, the enzyme responsible for the synthesis of SAM from methionine and ATP. The S box system uses SAM as the effector molecule both in vivo (15,16) and in vitro (15,17,18), and regulation occurs primarily at the level of premature termination of transcription (14,15). Translational regulation has also been predicted in certain S box genes, predominantly those found in Gram-negative organisms (refs.…”