Neutrophil elastase produces 52-kD and 30-kD glucocorticoid receptor fragments in the cytosol of human leukemia cells.
Characterization of glucocorticoid receptors in leukemia cells is important to understand mechanisms of glucocorticoid resistance but has been impeded by receptor fragmentation in cytosol extracts. We recently found that formation of 52- and 30-kilodalton (kD) glucocorticoid receptor fragments in cytosol of leukemia cells is due to proteolysis and is blocked by diisopropylfluorophosphate (DFP). In the present study, we identify a 28-kD serine protease in cytosol of leukemia cells that binds [3H]DFP and correlates with the formation of 52- and 30-kD receptor fragments. This protease is immunoprecipitated by antiserum to neutrophil elastase. Limited digestion of [3H]dexamethasone-21-mesylate-labeled receptors by purified neutrophil elastase produces 52- and 30-kD receptor fragments. Receptor fragmentation in the cytosol of leukemia cells in inhibited by methoxysuccinyl-alanyl-alanyl-prolyl-valyl-chloromethylketone, a highly specific inhibitor of neutrophil elastase. The addition of as few as 5% neutrophils to a lymphoid cell suspension provides sufficient elastase to produce receptor fragmentation. Our findings indicate that neutrophil elastase is responsible for receptor fragmentation in the cytosol of leukemia cells. The neutrophil elastase may be endogenous to the leukemia cells or may come from neutrophils that contaminate leukemia cell suspensions.
Distelhorst
C W
CW
Janiga
K E
KE
Howard
K J
KJ
Strandjord
S E
SE
Campbell
E J
EJ
D003600:Cytosol / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism; D006801:Humans; D007938:Leukemia / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D008214:Lymphocytes / Q000502:physiology; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology* / Q000502:physiology; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000097:blood*; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D012697:Serine Endopeptidases
D011965:Receptors, Glucocorticoid; D010450:Endopeptidases; D012697:Serine Endopeptidases; D010196:Pancreatic Elastase