Posttranscription Initiation Control of Tryptophan Metabolism in Bacillus subtilis by the trp RNA-Binding Attenuation Protein (TRAP), anti-TRAP, and RNA Structure

Abstract

2Organisms utilize a wide range of regulatory mechanisms to control gene expression. While regulation of transcription initiation is a common regulatory strategy, it is now apparent that this is only the starting point. Bacteria have developed several sophisticated regulatory mechanisms that allow the organism to modulate gene expression after transcription has initiated. In addition, several subtle mechanisms allow organisms to finetune the final level of any particular gene product. Several of these mechanisms, which operate after transcription initiation, are crucial for regulating tryptophan metabolism in Bacillus subtilis.The B. subtilis trpEDCFBA operon contains six of the seven genes that are required for the biosynthesis of tryptophan from chorismic acid, the common aromatic amino acid precursor (Fig. 1). The trp operon is present within a histidine and aromatic amino acid supraoperon. In addition to the trp operon promoter driving expression of this operon, the promoter from the upstream aroFBH operon contributes to trp operon expression. Since the first terminator for the aro operon is the terminator in the trp leader, transcriptional readthrough results in transcription of the trp operon structural genes (23). trpG, the remaining tryptophan biosynthetic gene, is present in an operon primarily concerned with folic acid biosynthesis ( Fig. 1) (38). Since there is no evidence for regulation of initiation from the trpEDCFBA promoter, it appears that the greater than 1,000-fold regulation observed for TrpE synthesis occurs after transcription has initiated. The trp RNA-binding attenuation protein (TRAP) plays a central role in controlling tryptophan metabolism by sensing the concentration of tryptophan in the cell (for previous reviews, see references 3, 20, 23). Another recently identified protein called anti-TRAP (AT) antagonizes TRAP activity (41). Since expression of the gene encoding AT responds to the accumulation of uncharged tRNA Trp , it is now apparent that B. subtilis regulates tryptophan biosynthesis by sensing the levels of both tryptophan and uncharged tRNA Trp in the cell (35, 41). TRAP regulates tryptophan biosynthesis by participating in transcription attenuation and translational control mechanisms. In the transcription attenuation mechanism, TRAP is responsible for the decision to terminate transcription in the trp operon leader region or to allow transcription to proceed into the trp structural genes by sensing the level of tryptophan in the cell (e.g., 4, 9, 27, 33). TRAP is also responsible for regulating translation of trpE and trpG. In the case of trpE, TRAP binding promotes formation of an RNA structure that sequesters the trpE Shine-Dalgarno (SD) sequence (14, 27, 31). Interestingly, the trpE SD sequence is more than 100 nucleotides downstream from the TRAP binding site. In contrast, TRAP regulates TrpG synthesis by binding to a segment of the trpG message that contains the SD sequence (7,16,38,44). In addition to the tryptophan biosynthetic genes, TRAP regulates expression of a p...

Crystal structure of Bacillus subtilis anti-TRAP protein, an antagonist of TRAP/RNA interaction

“…Six of these genes are arranged in the trpECDFBA suboperon of the aro supraoperon, and the seventh, trpG, is located in the folate operon (1). When cellular levels of tryptophan are elevated TRAP becomes activated, the apparent dissociation constant of tryptophan binding to TRAP being 5-7.5 M (5, 6).…”

“…I n Bacillus subtilis, both the level of intracellular free tryptophan and the level of uncharged tRNA Trp are used as regulatory signals for modulation of trp operon expression (1)(2)(3)(4). The first regulatory mechanism involves the trp RNA-binding attenuation protein (TRAP), an 11-subunit oligomer that is activated by free tryptophan to bind to a specific segment of trp mRNA.…”

“…This binding prevents formation of an antiterminator hairpin, thus permitting formation of a transcription terminator hairpin, which halts transcription of the operon (1,(8)(9)(10)(11). TRAP also regulates translation of trpE (10,11), trpG (pabA) (12,13), trpP (yhaG) (14,15), and ycbK (16) by binding to specific triplet repeat segments of these mRNAs and inhibiting translation initiation (1,2).…”

A Renaissance of Metabolite Sensing and Signaling: From Modular Domains to Riboswitches

“…An excellent example of this is the Trp RNA binding attenuation protein (TRAP) in Bacillus subtilis, which is responsible for both transcriptional attenuation and translational control of Trp synthetic pathway genes (Babitzke and Gollnick, 2001). The enzymes necessary for the synthesis of Trp from chorismate in B. subtilis are coded by the trpEDCFBA operon.…”

Recycling of a regulatory protein by degradation of the RNA to which it binds

“…By contrast, in Bacillus subtilis the trp leader antiterminator structure is the default. Formation of the terminator structure depends on binding of 1 TRAP, a regulatory protein that can bind trp leader RNA only when it is activated by tryptophan (3)(4)(5). In conditions of low tryptophan, TRAP does not bind trp leader RNA, allowing formation of the antiterminator structure and transcription of the trp operon structural genes.…”