EslB Is Required for Cell Wall Biosynthesis and Modification in Listeria monocytogenes

Abstract

Lysozyme is an important component of the innate immune system. It functions by hydrolysing the peptidoglycan (PG) layer of bacteria. The human pathogen Listeria monocytogenes is intrinsically lysozyme resistant. The peptidoglycan N-deacetylase PgdA and O-acetyltransferase OatA are two known factors contributing to its lysozyme resistance. Furthermore, it was shown that the absence of components of an ABC transporter, here referred to as EslABC, leads to reduced lysozyme resistance. How its activity is linked to lysozyme resistance is still unknown. To investigate this further, a strain with a deletion in eslB, coding for a membrane component of the ABC transporter, was constructed in L. monocytogenes strain 10403S. The eslB mutant showed a 40-fold reduction in the minimal inhibitory concentration to lysozyme. Analysis of the PG structure revealed that the eslB mutant produced PG with reduced levels of O-acetylation. Using growth and autolysis assays, we show that the absence of EslB manifests in a growth defect in media containing high concentrations of sugars and increased endogenous cell lysis. A thinner PG layer produced by the eslB mutant under these growth conditions might explain these phenotypes. Furthermore, the eslB mutant had a noticeable cell division defect and formed elongated cells. Microscopy analysis revealed that an early cell division protein still localized in the eslB mutant indicating that a downstream process is perturbed. Based on our results, we hypothesize that EslB affects the biosynthesis and modification of the cell wall in L. monocytogenes and is thus important for the maintenance of cell wall integrity. IMPORTANCE The ABC transporter EslABC is associated with the intrinsic lysozyme resistance of Listeria monocytogenes. However, the exact role of the transporter in this process and in the physiology of L. monocytogenes is unknown. Using different assays to characterize an eslB deletion strain, we found that the absence of EslB not only affects lysozyme resistance, but also endogenous cell lysis, cell wall biosynthesis, cell division and the ability of the bacterium to grow in media containing high concentrations of sugars. Our results indicate that EslB is by a yet unknown mechanism an important determinant for cell wall integrity in L. monocytogenes.

Not Just Transporters: Alternative Functions of ABC Transporters in Bacillus subtilis and Listeria monocytogenes

“…EslA and EslB are predicted to form an ABC transporter; however, it is still unknown whether the transmembrane protein EslC is also part of the ABC transporter or whether it has an independent role of EslAB. Bacterial two-hybrid experiments indicated that EslB and EslC interact with each other [ 80 ], and we will thus refer to the transporter as the EslABC transporter. Phenotypic analysis of strains lacking either EslA or EslB revealed a strong increase in lysozyme sensitivity, and deletion of eslC does not affect lysozyme resistance of L. monocytogenes [ 80 , 81 , 82 ].…”

“…Bacterial two-hybrid experiments indicated that EslB and EslC interact with each other [ 80 ], and we will thus refer to the transporter as the EslABC transporter. Phenotypic analysis of strains lacking either EslA or EslB revealed a strong increase in lysozyme sensitivity, and deletion of eslC does not affect lysozyme resistance of L. monocytogenes [ 80 , 81 , 82 ]. This observation does not rule out that EslC might be part of the ABC transporter; however, it suggests that EslA and EslB have an independent role of EslC.…”

“…The PG produced by an eslB mutant is more deacetylated and less O -acetylated as compared to the PG of the L. monocytogenes wildtype strain. Furthermore, deletion of eslB results in the production of a thinner PG layer, and thus, the lysozyme sensitivity of the eslB mutant is likely caused by both, the decrease in O -acetylation and PG layer thickness [ 80 ]. The reduction in cell wall thickness in the eslB mutant also leads to an increased cell lysis and EslB is thus required for the maintenance of cell wall integrity in L. monocytogenes [ 80 ].…”

“…Furthermore, deletion of eslB results in the production of a thinner PG layer, and thus, the lysozyme sensitivity of the eslB mutant is likely caused by both, the decrease in O -acetylation and PG layer thickness [ 80 ]. The reduction in cell wall thickness in the eslB mutant also leads to an increased cell lysis and EslB is thus required for the maintenance of cell wall integrity in L. monocytogenes [ 80 ]. In addition to its importance for PG biosynthesis, EslB is required for proper cell division in L. monocytogenes as cells lacking EslB form elongated cells.…”

“…In addition to its importance for PG biosynthesis, EslB is required for proper cell division in L. monocytogenes as cells lacking EslB form elongated cells. Interestingly, localization studies with the early cell division protein ZapA, which is required for FtsZ filament stabilization [ 85 ], suggest that these elongated cells form multiple Z-rings [ 80 ]. Thus, a process downstream of the recruitment of early cell division proteins, but prior to septum formation seems to be disturbed in absence of EslB.…”