The MarR Family Regulator BmrR Is Involved in Bile Tolerance of Bifidobacterium longum BBMN68 via Controlling the Expression of an ABC Transporter

Abstract

In order to colonize the human gastrointestinal tract and exert their beneficial effects, bifidobacteria must effectively cope with toxic bile salts in the intestine; however, the molecular mechanism underlying bile tolerance is poorly understood. In this study, heterologous expression of a MarR family transcriptional regulator, BmrR, significantly reduced the ox bile resistance ofLactococcus lactisNZ9000, suggesting that BmrR might play a role in the bile stress response.In silicoanalysis combined with reverse transcription-PCR assays demonstrated thatbmrRwas cotranscribed withbmrAandbmrB, which encoded multidrug resistance (MDR) ABC transporters. Promoter prediction and electrophoretic mobility shift assays revealed that BmrR could autoregulate thebmrRABoperon by binding to thebmrbox (ATTGTTG-6nt-CAACAAT) in the promoter region. Moreover, heterologous expression ofbmrAandbmrBinL. lactisyielded 20.77-fold higher tolerance to 0.10% ox bile, compared to the wild-type strain. In addition, ox bile could disrupt the DNA binding activity of BmrR as a ligand. Taken together, our findings indicate that thebmrRABoperon is autoregulated by the transcriptional regulator BmrR and ox bile serves as an inducer to activate the bile efflux transporter BmrAB in response to bile stress inBifidobacterium longumBBMN68.IMPORTANCEBifidobacteria are natural inhabitants of the human intestinal tract. Some bifidobacterial strains are used as probiotics in fermented dairy production because of their health-promoting effects. Following consumption, bifidobacteria colonize the lower intestinal tract, where the concentrations of bile salts remain nearly 0.05% to 2.0%. Bile salts, as detergent-like antimicrobial compounds, can cause cellular membrane disruption, protein misfolding, and DNA damage. Therefore, tolerance to physiological bile stress is indeed essential for bifidobacteria to survive and to exert probiotic effects in the gastrointestinal tract. InB. longumBBMN68, the MarR-type regulator BmrR was involved in the bile stress response by autoregulating thebmrRABoperon, and ox bile as an inducer could increase the expression of the BmrAB transporter to enhance the bile tolerance of BBMN68. Our study represents a functional analysis of thebmrRABoperon in the bile stress response, which will provide new insights into bile tolerance mechanisms inBifidobacteriumand other bacteria.

The genus Bifidobacterium: from genomics to functionality of an important component of the mammalian gut microbiota

“…At the same time, they may activate membrane-associated efflux pumps able to extrude bile salts and bile acids accumulated in the cytoplasm. Specifically, several multidrug transporters belonging to the ATP-binding cassette or the major facilitator superfamily have been identified as mediators of bile tolerance in B. breve and B. longum strains [70] , [92] , [93] . However, other defence mechanisms may be exploited by bifidobacterial taxa to limit the harmful effects of bile, i.e.…”

The effect of bile acids on the growth and global gene expression profiles in Akkermansia muciniphila

“…ABC transporters are divided into several groups with different characteristics associated with the uptake of nutrients and export of drugs (Locher 2016). ABC transporter BmrAB, comprising of 652-and 671-amino acid proteins, is required for bile acid tolerance in Bifidobacterium longum BBMN68 (Xu et al 2019). The amino acid sequence of BmrA (652 aa, BBMN68_1797) in B. longum is similar to the ABC transporter ATP-binding protein AMUC_RS00905 (34% identity; 54% similarity, 594 aa).…”

Stress Response in Bifidobacteria

“…In B. longum BBMN68, another bile-induced MDR transporter system was described recently (231). The transport system was found to be encoded within a polycistronic operon that consists of three genes, encoding the heterodimeric ABC transporter BmrAB and the MarR-family transcriptional regulator BmrR (231). In silico structural analysis and DNA-binding analysis of BmrR indicated that the expression of the bmrRAB operon is autoregulated by BmrR, which functions as a repressor.…”

“…In silico structural analysis and DNA-binding analysis of BmrR indicated that the expression of the bmrRAB operon is autoregulated by BmrR, which functions as a repressor. Under nonstress conditions, BmrR is assumed to bind to an inverted repeat sequence (ATTGTTG-N 6 -CAACAAT) in the promoter region of the operon, while being released from the promoter when some component of the bile interacts with BmrR and causes conformational changes in the DNA binding site (231). Heterologous overexpression of the bmrA gene and bmrB gene of B. longum BBMN68 in L. lactis significantly increased the host's resistance to ox bile (231).…”

“…Under nonstress conditions, BmrR is assumed to bind to an inverted repeat sequence (ATTGTTG-N 6 -CAACAAT) in the promoter region of the operon, while being released from the promoter when some component of the bile interacts with BmrR and causes conformational changes in the DNA binding site (231). Heterologous overexpression of the bmrA gene and bmrB gene of B. longum BBMN68 in L. lactis significantly increased the host's resistance to ox bile (231). Similarly, in B. breve UCC2003, the expression of three consecutive genes encoding a MarR regulator and the heterodimeric ABC transporter were also induced upon bile stress, mainly in the form of oxgall (228).…”