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Mg2+-Dependent Translational Speed Bump Acts to Regulate COMMENTARY + Mg2 -dependent translational speed bump acts to regulate gene transcription COMMENTARY Kelly T. Hughesa,1 DNA Transcription in Bacteria (7). Normally, an mRNA in bacteria is described as The central dogma of molecular biology states that having a 5′UTR between the transcriptional start-site DNA is transcribed (by RNA polymerase) to messenger and the translation initiation codon. The 5′UTR is often RNA that is then translated (by ribosomes) into protein used in translational control by controlling the mRNA (1). The regulation of gene expression can occur at any secondary structure to either expose or occlude the ri- level: transcription, mRNA stability, translation that in- bosome binding site upstream of the translation initia- cludes mRNA folding to reveal or occlude translation tion codon. Terminator sequences upstream of the initiation sites, and protein modification and stability. In structural genes of many bacterial operon systems are eukaryotes, the processes of transcription and trans- used to effect transcriptional control, such as in the lation are carried out in the cell nucleus and cytoplasm, case of riboswitches. Riboswitches allow metabolites to respectively. In bacteria, which lack a nucleus, the pro- control alternative mRNA secondary structures in the cesses of transcription and translation are coupled. This 5′UTR region to facilitate either the formation or de- provides an evolutionary target that is unique to bacteria stabilization of a transcriptional terminator (8, 9). for controlling gene expression. Many gene regulatory mechanisms in bacteria use transcription–translation Control of DNA Transcription by Charged tRNA coupling to affect whether transcription elongation The discovery of a role for charged tRNA species in through specific single or multigene operons continues transcriptional control of the histidine biosynthetic or terminates in response to changing concentrations of operons led to the discovery that translation played the metabolic product associated with a given operon. a critical role in the control of transcription in many In bacteria, termination of transcription occurs when amino acid biosynthetic operons (10). DNA sequencing RNA polymerase is preceded by a stretch of RNA that is of amino acid biosynthetic operons revealed the pres- not being translated, which occurs in the 3′-untranslated ence of 5′-translated, short, leader peptide regions RNA following the final translation termination codon of preceding the structural genes of these operons. These a given operon. Transcription termination in bacteria translated leader peptide regions were an integral occurs by one of two mechanisms that is either de- component of mechanisms that allowed for regulation pendent on the Rho transcriptional terminal factor or by of transcription of amino acid biosynthetic operons in the Rho-independent mechanism that involves the for- response to end product amino acid levels (7). The mation of a folded stem-loop structure in the RNA “true” end product sensed in the regulation of these followed by a poly-U sequence (2–4). Rho protein rec- amino acid biosynthetic operons was not a given amino ognizes untranslated RNA that is rich in cytosine base acid itself. Rather, the regulatory mechanisms sensed residues (5). levels of a specific amino acid in its charged form as an Suppression of transcription termination in gene aminoacyl tRNA. The leader peptide includes codons regulation was first characterized as a mechanism to for the specific amino acid end-product of that amino control the timing of gene expression during the infection acid biosynthetic operon. Between the leader peptide growth cycle of bacteriophage λ in Escherichia coli (6). coding sequence and the initiation codon of the first Here, antiterminator proteins, such as the λ N protein, gene in the biosynthetic operon resides a transcrip- must be transcribed, translated, and reach sufficient lev- tional attenuator sequence. Whether transcription els to bind termination sequences to prevent termination continues into the biosynthetic genes or attenuates at and allow transcription to continue into downstream the attenuator sequence depends on how the RNA genes that are needed later in the infection cycle. sequence that includes the attenuator folds. Alternative Suppression of transcription termination during RNA folding, that either promotes formation or de- transcription–translation coupling was later found to stabilization of the attenuator depends on the rate of regulate expression of amino acid biosynthetic operons translation by the ribosome through the leader peptide. aDepartment of Biology, University of Utah, Salt Lake City, UT 84112 Author contributions: K.T.H. wrote the paper. The author declares no conflict of interest. See companion article on page 15096. 1Email: [email protected]. www.pnas.org/cgi/doi/10.1073/pnas.1618222114 PNAS | December 27, 2016 | vol. 113 | no. 52 | 14881–14883 Downloaded by guest on September 28, 2021 + Because transcription and translation are coupled in bacteria, translation speedometer to sense Mg2 concentrations by Salmonella the initial ribosome that translates the leader peptide follows di- cells (17). The unique chemical interactions between magnesium + rectly behind the transcribing RNA polymerase complex (11). and phosphate make Mg2 essential in many cellular processes + When the ribosome reaches the leader peptide termination codon, involving nucleic acids. Mg2 is required for hundreds of enzymatic the untranslated sequence between the leader peptide stop codon reactions. Salmonella possesses three uptake systems for magne- + and the initiation codon of the first biosynthetic gene folds in a way sium ion. As an enteric pathogen, Salmonella uses low Mg2 con- that forms a transcriptional attenuator. When the levels of the amino centrations as one of multiple sensors in determining that it is inside acid end product of a specific amino acid biosynthetic pathway a host cell to control expression of genes required for virulence (18). drop, such that the codons in the leader peptide that code for that Magnesium and phosphate are also connected in controlling ex- amino acid are translated at a reduced rate, the unbound RNA se- pression of Salmonella virulence genes. A two-component regulatory quence between first translating ribosome and the transcribing RNA system, PhoQ-PhoP, originally described as a regulator of the phoN complex is extended to include bases in the 3′-end of the leader phosphatase gene, acts as an environmental sensor that effects peptide. Exposed 3′ leader peptide-coding bases are free to in- expression of 5% of the Salmonella genome, and is a primary teract with downstream RNA attenuator sequences, destabilize the regulator of genes required for Salmonella pathogenesis includ- + + attenuator form, and RNA polymerase continues to transcribe into ing Mg2 uptake systems (19). One Mg2 uptake system, mgtA,is the structural genes of the biosynthetic operon for that amino acid. This mechanism allows for the transcription of a given amino acid biosynthetic operon to depend on the levels of the specific amino In PNAS, Gall et al. demonstrate a novel, leader acyl tRNA for that operon and respond accordingly. peptide-dependent attenuation mechanism + One of the best-characterized attenuation control mechanisms that acts as a Mg2 -dependent translation + in bacteria for amino acid biosynthesis is that of the histidine speedometer to sense Mg2 concentrations by biosynthetic operon (his)ofSalmonella (12). The Salmonella his Salmonella operon includes nine structural genes. The first structural gene in cells. the his operon is preceded by a classic Rho-independent termi- nator, which is preceded by the his leader peptide sequence. The induced more than 100-fold in response to low magnesium in a his leader peptide codes for 16 amino acids, including 7 con- PhoQP-dependent manner (20, 21). secutive histidine codons (13). At high levels of charged histidyl aminoacyl tRNA, the first translating ribosome proceeds behind Slow Pro Translation as a Metabolite Sensor Mechanism RNA polymerase to the leader peptide termination codon, the In addition to PhoQP-dependent transcription of mgtA,Galletal. attenuator forms, and RNA polymerase comes off the DNA as now demonstrate a second control mechanism is involved that uses + transcription is terminated. At low histidyl-tRNA levels, the first aMg2 -dependent translational speedometer through a leader translating ribosome stalls in the stretch on consecutive histidine peptide sequence reminiscent of the attenuation control mecha- codons within the his leader peptide sequence and exposed RNA nisms in control of amino acid biosynthetic operon expression (17). + bases at the 3′-end of the his leader peptide interfere with the The Mg2 -dependent translation speedometer mechanism takes folding of the RNA into the attenuator form. advantage of the relatively slow translation of proline codons. The The properties of the his leader peptide in the his operon at- mgtA leader peptide contains a stretch of seven codons with four tenuation system were used to develop a translational speedometer proline codons spaced by intervening codons: Pro-Glu-Pro-Thr- that measures how fast ribosomes translate the codons in the his Pro-Leu-Pro. Anything that slows translation through the al- leader region that normally include the seven consecutive histidine ternating Pro codon region of the mgtA leader peptide results in codons (14). Substitution at position His5
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