Outer Membrane Regulation in Acinetobacter: Controlling the Shield against Antibiotics.

Acinetobacter baumannii is a bacterium the CDC categorizes as an urgent threat due to its distinctive level of antibiotic resistance. In A. baumannii, BfmRS is a global regulatory system that controls much of this resistance through regulating the cell envelope of the bacterium. However, the mechanisms with which BfmRS controls envelope synthesis and antibiotic resistance are unclear. Whether BfmRS regulates certain outer membrane proteins to control antibiotic resistance was studied. To test this hypothesis, reporters of gene expression of two outer membrane proteins, Omp25 and CarO, were constructed and then using the fluorescent protein GFP, the effect of BfmRS activating or inactivating mutations on the strength of gene expression was measured. The two genes were also deleted from the bacterial chromosome, and the resulting change in resistance to colistin, an important "last line of defense" drug against A. baumannii, were determined. From these experiments, the outer membrane proteins were indeed found to be regulated by BfmRS and to contribute to intrinsic colistin resistance. These findings suggest that Omp25 and CarO are important targets of regulation by BfmRS related to antibiotic resistance. Potential binding motifs in their promoter regions that may determine this regulation were also identified. For next steps, we will examine the role of these motifs on the strength of BfmRS-dependent gene expression by testing truncated promoters with fewer predicted BfmRS target sites. The binding motifs will also be modified through substitution mutations and then effects on gene expression levels will be measured. The effect of Omp25 and CarO on permeability to additional antibiotics and molecules will be tested. Altogether, this work offers insight into the mechanisms of BfmRS control of the cell envelope and drug resistance, which can help us use the system as a potential target for new treatments.