Guanine nucleotide-induced polymerization of actin in electropermeabilized human neutrophils.
The effects of exogenous guanine nucleotides on the polymerization of actin in human neutrophils were tested in an electropermeabilized cell preparation. Close to 40% permeabilization was achieved with a single electric discharge as measured by nucleic acid staining with ethidium bromide or propidium iodide with minimal (less than 2%) release of the cytoplasmic marker lactate dehydrogenase. In addition, electropermeabilized neutrophils retained their capacity to produce superoxide anions and to sustain a polymerization of actin in response to surface-receptor dependent stimuli such as chemotactic factors. Electropermeabilization produced a rapid and transient permeabilization that allowed the entry of guanine nucleotides into the cells. GTP and, to a larger extent, its nonhydrolyzable analog guanosine 5'-O-2-thiotriphosphate (GTP[S]), induced a time- and concentration-dependent polymerization of actin, as determined by increased staining with 7-nitrobenz-2-oxa-1,3-diazolylphallacidin. The effects of the aforementioned guanine nucleotides were antagonized by GDP[S], but were insensitive to pertussis toxin. Cholera toxin potentiated to a small degree the amount of actin polymerization induced by GTP[S]. These results provided direct evidence for the involvement of GTP-binding proteins in the regulation of the organization of the cytoskeleton of neutrophils, an event that is of crucial importance to the performance of the defense-oriented functions of these cells.
Therrien
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Département de Médecine, Université Laval, Ste Foy, Québec, Canada.
Naccache
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PH
D000199:Actins / Q000097:blood*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D002463:Cell Membrane Permeability; D004558:Electric Stimulation / Q000379:methods; D006150:Guanine Nucleotides / Q000097:blood / Q000494:pharmacology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000031:analogs & derivatives*; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D046911:Macromolecular Substances; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D013189:SRS-A / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology
D000199:Actins; D006150:Guanine Nucleotides; D046911:Macromolecular Substances; D013189:SRS-A; D013873:Thionucleotides; C034883:guanosine 5'-O-(2-thiotriphosphate); D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009240:N-Formylmethionine Leucyl-Phenylalanine; C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate