title: Protein I, a translocatable ion channel from Neisseria gonorrhoeae, selectively inhibits exocytosis from human neutrophils without inhibiting O2- generation.
abstract: Protein I, the major outer membrane protein of Neisseria gonorrhoeae, is a voltage-dependent anion channel which can translocate from the gonococcus into human cells. Since granule exocytosis from neutrophils is regulated by ion fluxes, we examined the effect of protein I on neutrophil activation. Pretreatment with protein I (250 nM) impaired degranulation from neutrophils: beta-glucuronidase release decreased to 27 +/- 6% S.E. of cells treated with N-f-Met-Leu-Phe (fMLP, 0.1 microM) and to 13 +/- 4% of cells treated with leukotriene B4 (LTB4, 0.1 microM); lysozyme release decreased to 52 +/- 17% of fMLP-treated cells and 22 +/- 9% of LTB4-treated cells. Morphometric analysis was consistent: control neutrophils increased their surface membrane after fMLP (43.3 +/- 5.6 microns relative perimeter versus 71.4 +/- 3.7 microns) while protein I-treated neutrophils did not (29.4 +/- 2 (S.E.) microns relative perimeter versus 34 +/- 4 microns). Enzyme release after exposure to phorbol myristate acetate was not affected (lysozyme: 86 +/- 27% of control). Cell/cell aggregation in response to fMLP was inhibited by treatment with protein I. However, generation of O2 was not affected. Protein I altered the surface membrane potential (Oxonol V): protein I evoked a transient membrane hyperpolarization which was not inhibited by furosemide. After exposure to fMLP, protein I-treated neutrophils underwent a furosemide-sensitive hyperpolarization rather than the usual depolarization. Protein I did not alter increments in [Ca]i (Fura-2) stimulated by fMLP (460 +/- 99 nM (S.E.) versus 377 +/- 44 nM) nor decrements in [pH]i (7.22 +/- 0.04 S.E. versus 7.22 +/- 0.02, bis-(carboxy-ethyl)carboxyfluorescein). The results suggest that degranulation and O2 generation have separate ionic requirements and that protein I interrupts the activation sequence proximal to activation of protein kinase C.
authors.affiliation: Department of Medicine, New York University Medical Center, New York 10016.
authors.forename: K A
authors.initials: KA
authors.lastname: Haines
authors.forename: L
authors.initials: L
authors.lastname: Yeh
authors.forename: M S
authors.initials: MS
authors.lastname: Blake
authors.forename: P
authors.initials: P
authors.lastname: Cristello
authors.forename: H
authors.initials: H
authors.lastname: Korchak
authors.forename: G
authors.initials: G
authors.lastname: Weissmann
mesh_terms: D001572:Benzofurans; D002118:Calcium / Q000097:blood; D003571:Cytochalasin B / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis / Q000187:drug effects; D016257:Fura-2; D005966:Glucuronidase / Q000097:blood; D006801:Humans; D007975:Leukotriene B4 / Q000494:pharmacology; D009113:Muramidase / Q000097:blood; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009344:Neisseria gonorrhoeae / Q000032:analysis*; D009419:Nerve Tissue Proteins / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects*; D013481:Superoxides / Q000097:blood*; D016704:Synapsins
chemical_list: D001572:Benzofurans; D009419:Nerve Tissue Proteins; D016704:Synapsins; D013481:Superoxides; D007975:Leukotriene B4; D003571:Cytochalasin B; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009113:Muramidase; D005966:Glucuronidase; D002118:Calcium; D016257:Fura-2