Irreversible inhibition of human natural killer cell natural cytotoxicity by modification of the extracellular membrane by the adenine nucleotide analog 5'-p-(fluorosulfonyl)benzoyl adenosine.
Extracellular adenine nucleotides are inhibitors of the human natural killer cell line NK3.3 natural cytotoxicity activity. Natural cytotoxicity was inhibited approximately 26% by 1 mM ATP and 21% by 1 mM ADP. 5'-Adenylyl imidodiphosphate, a nonhydrolyzable ATP analog, inhibited natural cytotoxicity by 41% at a concentration of 1 mM and > 97% at a concentration of 10 mM. In contrast, AMP was not inhibitory. Adenosine was a weak inhibitor of natural cytotoxicity and may represent an alternate regulatory pathway. Removal of the nucleotides resulted in the restoration of control levels of natural cytotoxicity activity. The affinity label 5'-p-(fluorosulfonyl)benzoyladenosine (5'-FSBA) is a synthetic analog of ATP or ADP containing an electrophilic fluorosulfonyl group capable of covalently modifying proteins at adenine di- and triphosphate nucleotide-binding sites. Natural cytotoxicity was irreversibly inhibited by modification of the extracellular membrane of NK3.3 cells by 5'-FSBA. This inhibition was concentration dependent with an I50 approximately 100 microM and complete inhibition at 1 mM. Modification of NK3.3 by 5'-FSBA did not affect the formation of effector-target cell conjugates; however, granule release was inhibited. This targets the site of inhibition by 5'-FSBA modification to a pathway preceding granule release. Irreversible, covalent modification of surface adenine nucleotide-binding proteins by 5'-FSBA provides a probe to study the role of specific adenine nucleotide-binding proteins in the extracellular regulation of natural killer cytolytic activity by adenine nucleotides.
Dombrowski
K E
KE
Department of Veterans Affairs Medical Center, Amarillo, Texas 79106, USA.
Cone
J C
JC
Bjorndahl
J M
JM
Phillips
C A
CA
D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D001565:Benzoates / Q000494:pharmacology; D002051:Burkitt Lymphoma; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects*; D003594:Cytoplasmic Granules / Q000378:metabolism; D003602:Cytotoxicity, Immunologic / Q000187:drug effects*; D005110:Extracellular Space; D053804:Granzymes; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000187:drug effects* / Q000276:immunology; D004915:Leukemia, Erythroblastic, Acute; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D012697:Serine Endopeptidases / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D001565:Benzoates; D007166:Immunosuppressive Agents; D000266:Adenylyl Imidodiphosphate; D010664:Phenylmethylsulfonyl Fluoride; D000244:Adenosine Diphosphate; C010959:5'-(4-fluorosulfonylbenzoyl)adenosine; D000255:Adenosine Triphosphate; D053804:Granzymes; D012697:Serine Endopeptidases; C509212:GZMA protein, human; D000241:Adenosine