Anaplasma phagocytophilum Outer Membrane Protein A Interacts with Sialylated Glycoproteins To Promote Infection of Mammalian Host Cells

ABSTRACTAnaplasma phagocytophilumis the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis (HGA).A. phagocytophilumbinding to sialyl Lewis x (sLex) and other sialylated glycans that decorate P selectin glycoprotein 1 (PSGL-1) and other glycoproteins is critical for infection of mammalian host cells. Here, we demonstrate the importance ofA. phagocytophilumouter membrane protein A (OmpA) APH_0338 in infection of mammalian host cells. OmpA is transcriptionally induced during transmission feeding ofA. phagocytophilum-infected ticks on mice and is upregulated during invasion of HL-60 cells. OmpA is presented on the pathogen's surface. Sera from HGA patients and experimentally infected mice recognize recombinant OmpA. Pretreatment ofA. phagocytophilumorganisms with OmpA antiserum reduces their abilities to infect HL-60 cells. The OmpA N-terminal region is predicted to contain the protein's extracellular domain. GlutathioneS-transferase (GST)-tagged versions of OmpA and OmpA amino acids 19 to 74 (OmpA19-74) but not OmpA75-205bind to, and competitively inhibitA. phagocytophiluminfection of, host cells. Pretreatment of host cells with sialidase or trypsin reduces or nearly eliminates, respectively, GST-OmpA adhesion. Therefore, OmpA interacts with sialylated glycoproteins. This study identifies the firstA. phagocytophilumadhesin-receptor pair and delineates the region of OmpA that is critical for infection.

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Anaplasma marginale Outer Membrane Protein A Is an Adhesin That Recognizes Sialylated and Fucosylated Glycans and Functionally Depends on an Essential Binding Domain

Infection and Immunity ◽

10.1128/iai.00968-16 ◽

2016 ◽

Vol 85 (3) ◽

Cited By ~ 9

Author(s):

Kathryn S. Hebert ◽

David Seidman ◽

Aminat T. Oki ◽

Jerilyn Izac ◽

Sarvani Emani ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Protein A ◽

Anaplasma Marginale ◽

Binding Domain ◽

Host Cells ◽

Content Type ◽

Outer Membrane Protein A ◽

Glycan Receptors

ABSTRACT Anaplasma marginale causes bovine anaplasmosis, a debilitating and potentially fatal tick-borne infection of cattle. Because A. marginale is an obligate intracellular organism, its adhesins that mediate entry into host cells are essential for survival. Here, we demonstrate that A. marginale outer membrane protein A (AmOmpA; AM854) contributes to the invasion of mammalian and tick host cells. AmOmpA exhibits predicted structural homology to OmpA of A. phagocytophilum (ApOmpA), an adhesin that uses key lysine and glycine residues to interact with α2,3-sialylated and α1,3-fucosylated glycan receptors, including 6-sulfo-sialyl Lewis x (6-sulfo-sLex). Antisera against AmOmpA or its predicted binding domain inhibits A. marginale infection of host cells. Residues G55 and K58 are contributory, and K59 is essential for recombinant AmOmpA to bind to host cells. Enzymatic removal of α2,3-sialic acid and α1,3-fucose residues from host cell surfaces makes them less supportive of AmOmpA binding. AmOmpA is both an adhesin and an invasin, as coating inert beads with it confers adhesiveness and invasiveness. Recombinant forms of AmOmpA and ApOmpA competitively antagonize A. marginale infection of host cells, but a monoclonal antibody against 6-sulfo-sLex fails to inhibit AmOmpA adhesion and A. marginale infection. Thus, the two OmpA proteins bind related but structurally distinct receptors. This study provides a detailed understanding of AmOmpA function, identifies its essential residues that can be targeted by blocking antibody to reduce infection, and determines that it binds to one or more α2,3-sialylated and α1,3-fucosylated glycan receptors that are unique from those targeted by ApOmpA.

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Anaplasma phagocytophilum Asp14 Is an Invasin That Interacts with Mammalian Host Cells via Its C Terminus To Facilitate Infection

Infection and Immunity ◽

10.1128/iai.00932-12 ◽

2012 ◽

Vol 81 (1) ◽

pp. 65-79 ◽

Cited By ~ 31

Author(s):

Amandeep Kahlon ◽

Nore Ojogun ◽

Stephanie A. Ragland ◽

David Seidman ◽

Matthew J. Troese ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Anaplasma Phagocytophilum ◽

Affinity Purification ◽

Surface Protein ◽

Surface Proteins ◽

Host Cells ◽

Mammalian Host ◽

Content Type

Anaplasma phagocytophilum, a member of the familyAnaplasmataceae, is the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis. The life cycle ofA. phagocytophilumis biphasic, transitioning between the noninfectious reticulate cell (RC) and infectious dense-cored (DC) forms. We analyzed the bacterium's DC surface proteome by selective biotinylation of surface proteins, NeutrAvidin affinity purification, and mass spectrometry. Transcriptional profiling of selected outer membrane protein candidates over the course of infection revealed thataph_0248(designatedasp14[14-kDaA. phagocytophilumsurface protein]) expression was upregulated the most duringA. phagocytophilumcellular invasion.asp14transcription was induced during transmission feeding ofA. phagocytophilum-infected ticks on mice and was upregulated when the bacterium engaged its receptor, P-selectin glycoprotein ligand 1. Asp14 localized to theA. phagocytophilumsurface and was expressed duringin vivoinfection. Treating DC organisms with Asp14 antiserum or preincubating mammalian host cells with glutathioneS-transferase (GST)–Asp14 significantly inhibited infection of host cells. Moreover, preincubating host cells with GST-tagged forms of both Asp14 and outer membrane protein A, anotherA. phagocytophiluminvasin, pronouncedly reduced infection relative to treatment with either protein alone. The Asp14 domain that is sufficient for cellular adherence and invasion lies within the C-terminal 12 to 24 amino acids and is conserved among otherAnaplasmaandEhrlichiaspecies. These results identify Asp14 as anA. phagocytophilumsurface protein that is critical for infection, delineate its invasion domain, and demonstrate the potential of targeting Asp14 in concert with OmpA for protecting against infection byA. phagocytophilumand otherAnaplasmataceaepathogens.

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Lack of Outer Membrane Protein A Enhances the Release of Outer Membrane Vesicles and Survival ofVibrio choleraeand Suppresses Viability ofAcanthamoeba castellanii

International Journal of Microbiology ◽

10.1155/2014/610190 ◽

2014 ◽

Vol 2014 ◽

pp. 1-6 ◽

Cited By ~ 9

Author(s):

Soni Priya Valeru ◽

Salah Shanan ◽

Haifa Alossimi ◽

Amir Saeed ◽

Gunnar Sandström ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Outer Membrane Proteins ◽

Protein A ◽

Membrane Vesicles ◽

Outer Membrane Vesicles ◽

Host Cells ◽

Wild Type ◽

Outer Membrane Protein A

Vibrio cholerae, the causative agent of the diarrhoeal disease cholera, survives in aquatic environments. The bacterium has developed a survival strategy to grow and survive insideAcanthamoeba castellanii. It has been shown thatV. choleraeexpresses outer membrane proteins as virulence factors playing a role in the adherence to interacted host cells. This study examined the role of outer membrane protein A (OmpA) and outer membrane vesicles (OMVs) in survival ofV. choleraealone and during its interaction withA. castellanii. The results showed that anOmpAmutant ofV. choleraesurvived longer than wild-typeV. choleraewhen cultivated alone. Cocultivation withA. castellaniienhanced the survival of both bacterial strains andOmpAprotein exhibited no effect on attachment, engulfment, and survival inside the amoebae. However, cocultivation of theOmpAmutant ofV. choleraedecreased the viability ofA. castellaniiand this bacterial strain released more OMVs than wild-typeV. cholerae. Surprisingly, treatment of amoeba cells with OMVs isolated from theOmpAmutant significantly decreased viable counts of the amoeba cells. In conclusion, the results might highlight a regulating rule forOmpAin survival ofV. choleraeand OMVs as a potent virulence factor for this bacterium towards eukaryotes in the environment.

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