Riboswitches for control of gene expression

Protein repressors and corepressors are not the only way in which bacteria control gene transcription. In many eukaryotes), it turns out that the regulation of the level of certain metabolites can also be controlled by newly discovered riboswitches. A riboswitch often a part of a messenger RNA (mRNA), and has a specific binding site for the metabolite (or a close relative), but only some (not all) mRNA molecules carry such a riboswitch. Numerous fundamental metabolic pathways in bacteria are regulated by this genetic control of a riboswitch. Elements called ‘riboswitches’ were discovered towards the end of the last century and in the early years of the present century (1999-2002). A riboswitch resides in the non-coding region (5′-UTR; UTR region) of the mRNA encoding a protein involved in the same metabolic pathway, which the riboswitch harbored by the mRNA regulates. These riboswitches selectively bind metabolites and modulate gene expression in response to changing ligand concentrations in the medium (Fig. 1).

A riboswitch (as a part of mRNA) is composed of two functional and sometimes distinct structural domains: (i) one binding domain is a natural ‘aptamer’* that binds to the metabolite, and (ii) the other domain is an ‘expression platform’ that harnesses allosteric changes in RNA structure, brought about by aptamer ligand complex formation, to control the expression of the adjacent gene expression platforms have been identified that control transcription termination or translation. In early 2005, at least seven fundamental metabolites were already known which could bind with riboswitches, contributing to the regulation of at least 68 genes in Bacillus subtilis. These targets included the following: (i) the nucleobases, guanine and adenine, (ii) the amino acid lysine, and (iii) coenzymes thiamine pyrophosphate (TPP), FMN, SAM, and B12 (adenosylcobalamin or AdoCbl).

Several classes of riboswitches have also been detected through database searches. These riboswitches were found to be distributed widely amongst prokaryotic organisms, and examples have been identified in archaeans also. Sequence variants of the natural aptamer for thiamine pyrophosphate (TPP) also have been identified in certain fungal and plant mRNAs, where they bind the coenzyme with affinities that match those of their prokaryotic counterparts. These findings indicate that riboswitches are a fundamental and widespread form of genetic

Riboswitches for Control of Transcription

Some of the metabolites that bind to riboswitches and bring about control at the transcription level include the following: (i) the purines, adenine, and guanine; (ii) the amino acids, glycine, and lysine (iii) flavin mononucleotide (the prosthetic group of NADH dehydrogenase), (iv) S-adenosyl methionine (that donates methyl groups to many molecules, including DNA, (v) the cap at the 5 ends of messenger RNA,

In each of the above cases, the riboswitch regulates transcription of genes involved in the metabolism of that molecule, The metabolite binds to the growing mRNA and induces an allosteric change that (i) for some genes causes further synthesis of the mRNA to terminate before forming a functional product, and (ii) for other enhances completion of synthesis of the genes, mRNA. In both cases, the riboswitch controls the level of metabolite. It is a kind of feedback inhibition, where the end product of a biosynthetic pathway binds not the enzyme of the pathway to bring about allosteric modification in the enzyme, but instead binds mRNA for such an enzyme, so that the cell does not have to waste its resources in synthesizing the enzyme.

Riboswitches for control of gene expression
Diverse strategies for gene regulation with the help of riboswitches, which have been linked to both transcriptional attenuation (a) and transcriptional inhibition (b) mechanisms; characterized thermosensors also use a translational inhibition strategy(c)

Riboswitches for Control of Translation

Several examples are available, where a riboswitch is involved in the regulation of gene expression binds to a part of the riboswitch, the latter undergoes allosteric modifications, thus rendering the translation level, so that when a metabolite binds to a part of the riboswitch, the later undergoes allosteric modification, thus rendering the mRNA component of riboswitch incapable of translation.

Control of thiamine synthesis.

If thiamine pyrophosphate (TPP), the active form of thiamine (vitamin B1) is available in the culture medium of E. coli, it binds to a messenger RNA, whose protein product is an enzyme needed to synthesize thiamine from the ingredients in a minimal medium. Binding induces an allosteric shift in the structure of the mRNA so that it can no longer bind to a ribosome and thus cannot be translated into the enzyme. Thus E. coli no longer waste resources on synthesizing a preformed.

Control of vitamin B12 transporter synthesis.

If sufficient vitamin B12 is present in the cell, it binds to the mRNA which encodes a vitamin that is an available protein (cobalamine-transport protein) needed to import the vitamin B12 from the culture medium. This, too, induces an allosteric shift in the mRNA prevents it from binding a ribosome. Thus E.coli no longer wastes resources on a transporter for a vitamin, enough of which is synthesizing an already present within the cell.

Some gram-positive Control of synthesis of an enzyme of sugar biosynthetic pathway.

Some gram-positive bacteria (E. coli is gram-negative) use a riboswitch to control the level of sugar needed to synthesize their cell wall. In this case, as the concentration of the sugar builds up, it binds to the messenger RNA (mRNA) encoding an enzyme involved in sugar biosynthesis. Such a binding renders the mRNA incapable of synthesizing the protein.

Mechanism of Riboswitch Regulation

Three different mechanisms for the regulation of gene expression through riboswitches (RNA sensors) are depicted in Figure 1.

Transcriptional termination mechanism.

As shown in Figure 1(a), in the absence of the relevant metabolite, part of the terminator is bound by an anti-terminator rendering it non-functional. Transcription proceeds past the polyuridine stretch following the terminator sequence extends into the coding sequence. In the presence of the relevant metabolite, on the other hand, ligand-bound RNA adopts an alternative conformation in which the anti-terminator is bound by an anti-anti-terminator. This allows the formation of the structurally distinctive terminator hairpin, which induces the release of RNA polymerase following the poly-uridine tract, thus permitting no transcription of the coding sequence that follows the terminator.

Translational inhibition mechanism.

In this mechanism, as shown in Figure 1(b), in the absence of cofactor-binding, the anti-Shine-Dalgarno (anti-SD) ribosome binding sequence is sequestered by an anti-anti-SD sequence; this conformation allows ribosome binding to the SD box and translation to occur. Cofactor binding restructures the RNA so that the anti-SD sequence is allowed to pair with the SD box, thus inhibiting translation.

A thermosensor from L. monocytogenes prfA.

In this mechanism, a stem structure adjacent to the SD box prevents translation at lower temperatures, but an increase in ambient temperature following host infection leads to melting of this structure, allowing the ribosome to access the SD box and translate prfA (Fig. 1(c)).