Amiloride inhibits phorbol ester-stimulated Na+/H+ exchange and protein kinase C. An amiloride analog selectively inhibits Na+/H+ exchange.
The human leukemic cell line, HL-60, differentiates in response to tumor-promoting phorbol esters. Recently, we have reported that one of the first events evoked by phorbol esters in HL-60 cells is the stimulation of Na+-dependent H+ efflux. In efforts to determine whether stimulation of Na+/H+ exchange by phorbol esters is coupled to induction of cellular differentiation, we found that 1) amiloride, a frequently used inhibitor of Na+/H+ exchange, rapidly inhibits phorbol ester-stimulated protein phosphorylation in vivo and protein kinase C-mediated phosphorylation in vitro, both with potency similar to that with which amiloride inhibits Na+/H+ exchange; 2) an amiloride analog, dimethylamiloride, is a far more potent inhibitor of Na+/H+ exchange than is amiloride, while being no more potent than amiloride in inhibiting phorbol ester/protein kinase C-mediated phosphorylation; and 3) at concentrations sufficient to completely inhibit Na+/H+ exchange, amiloride blocked phorbol ester-induced adhesion of HL-60 cells (adhesion being a property indicative of the differentiated state), but dimethylamiloride (as well as ethylisopropylamiloride, another very potent amiloride analog) did not. Thus, dimethylamiloride represents a potential tool for distinguishing protein kinase C-coupled from Na+/H+ exchange-coupled events in phorbol ester-stimulated cells.
Besterman
J M
JM
May
W S
WS
LeVine
H
H
Cragoe
E J
EJ
Cuatrecasas
P
P
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology*; D002352:Carrier Proteins / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate; D010703:Phorbol Esters / Q000494:pharmacology*; D010704:Phorbols / Q000494:pharmacology*; D010766:Phosphorylation; D011493:Protein Kinase C; D047428:Protein Kinase Inhibitors; D011719:Pyrazines / Q000494:pharmacology*; D011956:Receptors, Cell Surface / Q000378:metabolism; D011990:Receptors, Transferrin; D017923:Sodium-Hydrogen Exchangers
D002352:Carrier Proteins; D010703:Phorbol Esters; D010704:Phorbols; D047428:Protein Kinase Inhibitors; D011719:Pyrazines; D011956:Receptors, Cell Surface; D011990:Receptors, Transferrin; D017923:Sodium-Hydrogen Exchangers; D015240:Phorbol 12,13-Dibutyrate; C039465:5-dimethylamiloride; D000584:Amiloride; D011493:Protein Kinase C