Abstract
Neonatal meningitis, caused by Escherichia coli K1, is a serious central nervous system disease. We have established that macrophages serve as permissive niches for E. coli K1 to multiply in the host and for attaining a threshold level of bacterial load, which is a prerequisite for the onset of the disease. Here, we demonstrate experimentally that three N-glycans in FcγRIa interact with OmpA of E. coli K1 for binding to and entering the macrophages. Adoptive transfer of FcγRIa(-/-) bone marrow-derived macrophages transfected with FcγRIa into FcγRIa(-/-) newborn mice renders them susceptible to E. coli K1-induced meningitis. In contrast, mice that received bone marrow-derived macrophages transfected with FcγRIa in which N-glycosylation sites 1, 4, and 5 are mutated to alanines exhibit resistance to E. coli K1 infection. Our molecular dynamics and simulation studies predict that N-glycan 5 exhibits strong binding at the barrel site of OmpA formed by loops 3 and 4, whereas N-glycans 1 and 4 interact with loops 1, 3, and 4 of OmpA at tip regions. Molecular modeling data also suggest no role for the IgG binding site in the invasion process. In agreement, experimental mutations in IgG binding site had no effect on the E. coli K1 entry into macrophages in vitro or on the onset of meningitis in newborn mice. Together, this integration of experimental and computational studies reveals how the N-glycans in FcγRIa interact with the OmpA of E. coli K1 for inducing the disease pathogenesis.
Highlights
Escherichia coli OmpA interacts with Fc␥RI ␣-chain (Fc␥RIa) to invade macrophages
N-Glycosylation Sites 1, 4, and 5 in the Extracellular Domain of Fc␥RIa Are Important for E. coli K1 Binding to and Invasion of RAW 264.7 Macrophages—We previously demonstrated experimentally that OmpA of E. coli K1 interacts with GlcNAc1– 4GlcNAc epitopes of glycoproteins and that Fc␥RIa expression is critical for the entry into macrophages [6, 9, 10]
This study demonstrates three important pieces of evidence to confirm that E. coli K1 OmpA interaction with Fc␥RIa in macrophages is critical for the onset of meningitis in newborn mice: First, we showed clearly that three N-glycans (NG1, NG4, and NG5) present in the extracellular domains of Fc␥RIa interact with OmpA for binding to and invasion of macrophages and that there is no role for IgG binding region in this interaction
Summary
Escherichia coli OmpA interacts with Fc␥RI ␣-chain (Fc␥RIa) to invade macrophages. Results: Lack of three N-glycans in Fc␥RIa prevents E. coli invasion of macrophages and the onset of meningitis. Experimental mutations in IgG binding site had no effect on the E. coli K1 entry into macrophages in vitro or on the onset of meningitis in newborn mice. Together, this integration of experimental and computational studies reveals how the N-glycans in Fc␥RIa interact with the OmpA of E. coli K1 for inducing the disease pathogenesis. This integration of experimental and computational studies reveals how the N-glycans in Fc␥RIa interact with the OmpA of E. coli K1 for inducing the disease pathogenesis Fc␥R comprises a multigene family divided into three classes (Fc␥RI, II, and III), which are defined
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