Identification of the Neuroinvasive Pathogen Host Target, LamR, as an Endothelial Receptor for the Treponema pallidum Adhesin Tp0751

Abstract

Treponema pallidum subsp. pallidum is the causative agent of syphilis, a human-specific sexually transmitted infection that causes a multistage disease with diverse clinical manifestations. Treponema pallidum undergoes rapid vascular dissemination to penetrate tissue, placental, and blood-brain barriers and gain access to distant tissue sites. The rapidity and extent of T. pallidum dissemination are well documented, but the molecular mechanisms have yet to be fully elucidated. One protein that has been shown to play a role in treponemal dissemination is Tp0751, a T. pallidum adhesin that interacts with host components found within the vasculature and mediates bacterial adherence to endothelial cells under shear flow conditions. In this study, we further explore the molecular interactions of Tp0751-mediated adhesion to the vascular endothelium. We demonstrate that recombinant Tp0751 adheres to human endothelial cells of macrovascular and microvascular origin, including a cerebral brain microvascular endothelial cell line. Adhesion assays using recombinant Tp0751 N-terminal truncations reveal that endothelial binding is localized to the lipocalin fold-containing domain of the protein. We also confirm this interaction using live T. pallidum and show that spirochete attachment to endothelial monolayers is disrupted by Tp0751-specific antiserum. Further, we identify the 67-kDa laminin receptor (LamR) as an endothelial receptor for Tp0751 using affinity chromatography, coimmunoprecipitation, and plate-based binding methodologies. Notably, LamR has been identified as a receptor for adhesion of other neurotropic invasive bacterial pathogens to brain endothelial cells, including Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae, suggesting the existence of a common mechanism for extravasation of invasive extracellular bacterial pathogens. IMPORTANCE Syphilis is a sexually transmitted infection caused by the spirochete bacterium Treponema pallidum subsp. pallidum. The continued incidence of syphilis demonstrates that screening and treatment strategies are not sufficient to curb this infectious disease, and there is currently no vaccine available. Herein we demonstrate that the T. pallidum adhesin Tp0751 interacts with endothelial cells that line the lumen of human blood vessels through the 67-kDa laminin receptor (LamR). Importantly, LamR is also a receptor for meningitis-causing neuroinvasive bacterial pathogens such as Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae. Our findings enhance understanding of the Tp0751 adhesin and present the intriguing possibility that the molecular events of Tp0751-mediated treponemal dissemination may mimic the endothelial interaction strategies of other invasive pathogens.

Evidence that immunization with TP0751, a bipartite Treponema pallidum lipoprotein with an intrinsically disordered region and lipocalin fold, fails to protect in the rabbit model of experimental syphilis

“…[ 45 ] identified four regions, distributed around the lipocalin barrel, that mediate TP0751-laminin interactions. Of these, 'peptide 10' (p10) recently also was shown to interact with the laminin receptor (LamR) [ 74 ]. Mapping of p10 onto a 3D model for TP0751 97-228 reveals that it corresponds to two positively charged β strands (strands 5 and 6) with adjoining loop ( S3 Fig ).…”

“…In the absence of TonBdependent uptake pathways, the mechanism(s) for scavenging of heme by T. pallidum and transport across the OM are unclear; this enigma is part of the broader conundrum of transition metal acquisition by the syphilis spirochete [4]. In any event, while the small molecule binding properties of TP751 require more investigation, the results presented herein provide proof of principle for a biologically plausible alternative to the atypical binding sites on the external façade of the lipocalin barrel proposed from peptide mapping [45,74]. The structural similarities between the TP0751 lipocalin domain and Tpp17 are intriguing given the marked difference in their levels of expression.…”

“…Previously reported laminin-binding peptides [ 45 ] p4 (residues 97–111, cyan), p6 (residues 116–138, orange), p10 (residues 172–195, magenta) and p11 (residues 196–209, yellow) mapped onto the structure of TP0751 97-228 . p10 also has also been reported to interact with LamR [ 74 ]. TP0751 97-228 is shown in a different orientation compared to other figures to facilitate presentation of the grid box and the locations of laminin-binding peptides.…”

Characterization of Treponema pallidum Dissemination in C57BL/6 Mice

“…Studies have shown that T. pallidum can break through the blood-brain barrier at the beginning of infection and colonize the brain tissue in both humans and experimental rabbits. Tp92 and Tp0751 may be involved in this process by mediating the adhesion of T. pallidum to host cells, although its exact mechanism needs to be further elucidated ( 31 , 32 ). Our results suggested that the brain may be an important site for spirochetal invasion, demonstrating consistent PCR positivity.…”

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“…Studies have shown that T. pallidum can break through the blood-brain barrier at the beginning of infection and colonize the brain tissue in both humans and experimental rabbits. Tp92 and Tp0751 may be involved in this process by mediating the adhesion of T. pallidum to host cells, although its exact mechanism needs to be further elucidated (31,32). Our results suggested that the brain may be an important site for spirochetal invasion, demonstrating consistent PCR positivity.…”