Integrated Proteomic and Transcriptomic Analyses Reveal the Roles of Brucella Homolog of BAX Inhibitor 1 in Cell Division and Membrane Homeostasis of Brucella suis S2

Abstract

BAX inhibitor 1 (BI-1) is an evolutionarily conserved transmembrane protein first identified in a screening process for human proteins that suppress BAX-induced apoptosis in yeast cells. Eukaryotic BI-1 is a cytoprotective protein that suppresses cell death induced by multiple stimuli in eukaryotes. Brucella, the causative agent of brucellosis that threatens public health and animal husbandry, contains a conserved gene that encodes BI-1-like protein. To explore the role of the Brucella homolog of BI-1, BrBI, in Brucella suis S2, we constructed the brbI deletion mutant strain and its complemented strain. brbI deletion altered the membrane properties of Brucella suis S2 and decreased its resistance to acidic pH, H2O2, polymyxin B, and lincomycin. Additionally, deleting brbI led to defective growth, cell division, and viability in Brucella suis S2. We then revealed the effect of brbI deletion on the physiological characteristics of Brucella suis S2 via integrated transcriptomic and proteomic analyses. The integrated analysis showed that brbI deletion significantly affected the expression of multiple genes at the mRNA and/or protein levels. Specifically, the affected divisome proteins, FtsB, FtsI, FtsL, and FtsQ, may be the molecular basis of the impaired cell division of the brbI mutant strain, and the extensively affected membrane proteins and transporter-associated proteins were consistent with the phenotype of the membrane properties’ alterations of the brbI mutant strain. In conclusion, our results revealed that BrBI is a bacterial cytoprotective protein involved in membrane homeostasis, cell division, and stress resistance in Brucella suis S2.

Alr Gene in Brucella suis S2: Its Role in Lipopolysaccharide Biosynthesis and Bacterial Virulence in RAW264.7

“…Mutant strains were confirmed by PCR using alr -qF and alr -qR primers. To obtain the complemented strain, we constructed an expression plasmid (pBBARpc) containing a Flag tag [ 3 ]. The full-length alr open reading frame was amplified using alr -F and alr -R primers, and the PCR product was cloned into the laboratory’s pre-existing pBBARpc vector to generate the recombinant pBBARpc- alr plasmid.…”

“…Brucella is a Gram-negative bacterium that grows extracellularly and which replicates within both phagocytic and non-phagocytic host cells [ 3 ]. Despite lacking many of the classical virulence factors that other pathogenic bacteria possess, including invasive proteases, exotoxins, capsules, and fimbriae, Brucella has evolved a remarkable ability to evade host immunity and cause chronic infection.…”

Regulation of the Gene for Alanine Racemase Modulates Amino Acid Metabolism with Consequent Alterations in Cell Wall Properties and Adhesive Capability in Brucella spp.

“…PI freely penetrates damaged cell membranes and binds to intracellular DNA, which leads to the emission of red fluorescence. Therefore, the fluorescence microscopy detection of PI staining within cells indirectly reflects changes in the Brucella cell membrane [14]. No red fluorescence was observed in wild-type, ∆alr, and C∆alr strains without PI treatment.…”

“…For antibiotic resistance testing, cultures were placed in a 96-well enzyme-linked immunosorbent assay (ELISA) plate (at 2 × 10 4 CFUs/well) and were exposed to polymyxin B at concentrations of 1200, 600, and 300 µg/mL or lincomycin at concentrations of 500, 375, and 250 µg/mL at 37 • C for 1 h. The cultures were tenfold diluted and plated onto TSA agar, followed by incubation at 37 • C for 3 days. The CFUs after antibiotic treatment were counted and compared to those of untreated cultures to calculate the survival rate for each sample [14].…”

Brucella Manipulates Host Cell Ferroptosis to Facilitate Its Intracellular Replication and Egress in RAW264.7 Macrophages

“…Evidence supports that the virulence of Brucella is completely dependent on its ability to survive and replicate in host cells [9][10][11]. As a stealthy pathogen, Brucella has evolved numerous strategies to defend host bactericidal responses and then establish intracellular multiplication niches [12,13]. It has been reported that Brucella infection induces the premature death of neutrophils to promote their phagocytosis by macrophages [14,15].…”