title: Identification of heat shock protein 60 as the ligand on Histoplasma capsulatum that mediates binding to CD18 receptors on human macrophages.
abstract: Histoplasma capsulatum (Hc), is a facultative intracellular fungus that binds to CD11/CD18 receptors on macrophages (Mphi). To identify the ligand(s) on Hc yeasts that is recognized by Mphi, purified human complement receptor type 3 (CR3, CD11b/CD18) was used to probe a Far Western blot of a detergent extract of Hc cell wall and cell membrane. CR3 recognized a single 60-kDa protein, which was identified as heat shock protein 60 (hsp60). Biotinylation of viable yeasts, followed by precipitation with streptavidin-coated beads, and Western blotting with anti-hsp60 demonstrated that hsp60 was on the surface of Hc yeasts. Electron and confocal microscopy revealed that hsp60 resided on the yeast cell wall in discrete clusters. Recombinant hsp60 (rhsp60) inhibited attachment of Hc yeasts to Mphi. Recombinant hsp60 and Abs to CD11b and CD18 inhibited binding of yeasts to Chinese hamster ovary cells transfected with CR3 (CHO3). Polystyrene beads coated with rhsp60 bound to Mphi, and attachment was inhibited by Abs to CD11 and CD18. Freeze/thaw extract (F/TE), a preparation of Hc yeast surface proteins that contained hsp60, inhibited the attachment of Hc yeasts to Mphi. Depletion of hsp60 from F/TE removed the capacity of F/TE to block binding of Hc to Mphi. Interestingly, rhsp60 did not inhibit binding of Hc yeasts to dendritic cells (DC), which recognize Hc via very late Ag 5. Moreover, F/TE inhibited attachment of Hc to DC even when depleted of hsp60. Thus, Hc hsp60 appears to be a major ligand that mediates attachment of Hc to Mphi CD11/CD18, whereas DC recognize Hc via a different ligand(s).
authors.affiliation: Department of Cell Biology, Neurobiology, and Anatomy, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0560, USA.
authors.forename: Kristin H
authors.initials: KH
authors.lastname: Long
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Gomez
authors.forename: Randall E
authors.initials: RE
authors.lastname: Morris
authors.forename: Simon L
authors.initials: SL
authors.lastname: Newman
mesh_terms: D000818:Animals; D001667:Binding, Competitive / Q000276:immunology; D018845:CD11 Antigens / Q000378:metabolism; D018821:CD18 Antigens / Q000378:metabolism*; D016466:CHO Cells; D002448:Cell Adhesion / Q000276:immunology; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000032:analysis / Q000378:metabolism / Q000502:physiology*; D006224:Cricetinae; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism / Q000382:microbiology; D005656:Fungal Proteins / Q000378:metabolism; D006658:Histoplasma / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D016177:Macrophage-1 Antigen / Q000378:metabolism*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism* / Q000382:microbiology*; D008565:Membrane Proteins / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism; D011485:Protein Binding / Q000276:immunology
chemical_list: D018845:CD11 Antigens; D018821:CD18 Antigens; D018834:Chaperonin 60; D005656:Fungal Proteins; D016177:Macrophage-1 Antigen; D008565:Membrane Proteins