Pj. Mansfield et al., Regulation of polymorphonuclear leukocyte phagocytosis by myosin light chain kinase after activation of mitogen-activated protein kinase, BLOOD, 95(7), 2000, pp. 2407-2412
Polymorphonuclear leukocyte (PMNL) phagocytosis mediated by Fc gamma RII pr
oceeds in concert with activation of the mitogen-activated protein (MAP) ki
nase, extracellular signal-regulated kinase ERK2, We hypothesized that myos
in light chain kinase (MLCK) could be phosphorylated and activated by ERK,
thereby linking the MAP kinase pathway to the activation of cytoskeletal co
mponents required for pseudopod formation. To explore this potential linkag
e, PMNLs were challenged with antibody-coated erythrocytes (EIgG). Peak MLC
K activity, 3-fold increased over controls, occurred at 4 to 6 minutes, cor
responding with the peak rate of target ingestion and ERK2 activity. The ML
CK inhibitor ML-7 (10 mu mol/L) inhibited both phagocytosis and MLCK activi
ty to basal values, thereby providing further support for the linkage betwe
en the functional response and the requirement for MLCK activation. The MAP
K kinase (MEK) inhibitor PD098059 inhibited phagocytosis, MLCK activity, an
d ERK2 activity by 80% to 90%, To directly link ERK activation to MLCK acti
vation, ERK2 was immunoprecipitated from PMNLs after EIgG ingestion. The is
olated ERK2 was incubated with PMNL cytosol as a source of unactivated MLCK
and with MLCK substrate; under these conditions ERK2 activated MLCK, resul
ting in phosphorylation of the MLCK substrate or of the myosin light chain
itself. Because MLCK activates myosin, we evaluated the effect of directly
inhibiting myosin adenosine triphosphatase using 2,3-butanedione monoxime (
BDM) and found that phagocytosis was inhibited by more than 90% but MLCK ac
tivity remained unaffected. These results are consistent with the interpret
ation that MEK activates ERK, ERK2 then activates MLCK, and MLCK activates
myosin, MLCK activation is a critical step in the cytoskeletal changes resu
lting in pseudopod formation, (Blood, 2000;95:2407-2412) (C) 2000 by The Am
erican Society of Hematology.