Transcriptional Termination Control of a Novel ABC Transporter Gene Involved in Antibiotic Resistance in Bacillus subtilis

Abstract

In members of one of the subfamilies of the bacterial ATP binding cassette (ABC) transporters, the two nucleotide binding domains are fused as a single peptide and the proteins have no membrane-spanning domain partners. Most of the ABC efflux transporters of this subfamily have been characterized in actinomycetes, producing macrolide, lincosamide, and streptogramin antibiotics. Among 40 ABC efflux transporters of Bacillus subtilis, five proteins belong to this subfamily. None of these proteins has been functionally characterized. We examined macrolide, lincosamide, and streptogramin antibiotic resistance in insertional disruptants of the genes that encode these proteins. It was found that only a disruptant of vmlR (formerly named expZ) showed hypersensitivity to virginiamycin M and lincomycin. Expression of the vmlR gene was induced by the addition of these antibiotics in growth medium. Primer extension analysis revealed that transcription of the vmlR gene initiates at an adenosine residue located 225 bp upstream of the initiation codon. From the analysis of the vmlR and lacZ fusion genes, a 52-bp deletion from ؉159 to ؉211 resulted in constitutive expression of the vmlR gene. In this region, a typical -independent transcriptional terminator was found. It was suggested that the majority of transcription ends at this termination signal in the absence of antibiotics, whereas under induced conditions, RNA polymerase reads through the terminator, and transcription continues to the downstream vmlR coding region, resulting in an increase in vmlR expression. No stabilization of vmlR mRNA occurred under the induced conditions.

Characterization of sal (A), a Novel Gene Responsible for Lincosamide and Streptogramin A Resistance in Staphylococcus sciuri

“…It might be speculated, in light of recently reported findings, that a ubiquitous microorganism like S. sciuri, which has been isolated in soil, sand, and marsh grass (31), has maintained an indigenous Sal(A) determinant to survive within lincosamide-containing environments. This is probably what also happens with many other species of the Bacilli class, as recently exemplified by molecular characterization of VmlR in B. subtilis (32) and Lsa(A) in E. faecalis (8), which confer the same intrinsic resistance properties, the LS A resistance phenotype, upon the bacteria in which they reside.…”

Regulatory RNAs in Bacillus subtilis: a Gram-Positive Perspective on Bacterial RNA-Mediated Regulation of Gene Expression

“…The next top-10 segment in the all-independent category is S198 (48/62 genomes, MFE Z-score Ϫ5.1), which forms the leader region of the vlmR-encoded ABC transporter that was briefly addressed above. Transcription termination-mediated control of read-through transcription into vlmR has been reported and seems to be mediated by S198 (58). However, this function does not necessarily exclude a second regulatory mechanism for S198 as an sRNA, especially since such a dual-function RNA has been reported in Listeria monocytogenes (116).…”

“…However, when translation of BmrB is slowed down by the addition of ribosome-targeted antibiotics, an antiterminator structure forms that allows transcription to continue into the bmrCD operon (50). Interestingly, another TA system was reported in the 5=UTR of vmlR, which is also responsible for the efflux of ribosome-targeting antibiotics in B. subtilis (58). The 5=UTR of vmlR S198 also contains multiple putative ORFs, but it remains to be investigated whether these ORFs are involved in TA and to what extent the respective TA regulation mechanism is similar to that of bmrCD (50).…”

A New Evolutionary Variant of the Streptogramin A Resistance Protein, Vga(A) LC , from Staphylococcus haemolyticus with Shifted Substrate Specificity towards Lincosamides

“…Another vga gene, vga(B), has 59% nucleotide identity with vga(A) and confers only lowlevel resistance to streptogramin A but substantially increases the level of resistance to pristinamycin, a mixture of streptogramin A and streptogramin B (4,13). A new determinant, vmlR, conferring inducible virginamycin M (streptogramin A) and lincomycin resistance has been found in Bacillus subtilis (24). The predicted amino acid sequence of VmlR has 32%, 33%, and 29% amino acid identity to Vga(B), Vga(A), and Lmr(C), respectively.…”