TY - JOUR
T1 - MDR1 function is sensitive to the phosphorylation state of myosin regulatory light chain
AU - Bajaj, Gaurav
AU - Rodriguez-Proteau, Rosita
AU - Venkataraman, Anand
AU - Fan, Ying
AU - Kioussi, Chrissa
AU - Ishmael, Jane E.
N1 - Funding Information:
The American Association of Colleges of Pharmacy (New Investigator Program), and both the Cell Imaging and Culture Facilities ( P30-ES000210 ) and a Short-Term Training Grant ( T35-ES07316 ) from the National Institute of Environmental Health Sciences supported this work.
PY - 2010/7
Y1 - 2010/7
N2 - Multiple drug resistance protein 1 (MDR1) is composed of two homologous halves separated by an intracellular linker region. The linker has been reported to bind myosin regulatory light chain (RLC), but it is not clear how this can occur in the context of a myosin II complex. We characterized MDR1-RLC interactions and determined that binding occurs via the amino terminal of the RLC, a domain that typically binds myosin heavy chain. MDR1-RLC interactions were sensitive to the phosphorylation state of the light chain in that phosphorylation by myosin light chain kinase (MLCK) resulted in a loss of binding in vitro. We used ML-7, a specific inhibitor of MLCK, to study the functional consequences of disrupting RLC phosphorylation in intact cells. Pretreatment of polarized Madin-Darby canine kidney cells stably expressing MDR1 with ML-7 produced a significant increase in apical to basal permeability and a corresponding decrease in the efflux ratio (threefold; p<0.01) of [3H]-digoxin, a classic MDR1 substrate. Together these data show that MDR1-mediated transport of [3H]-digoxin can be modulated by pharmacological manipulation of myosin RLC, but direct MDR1-RLC interactions are atypical and not explained by the structure of the myosin II holoenzyme.
AB - Multiple drug resistance protein 1 (MDR1) is composed of two homologous halves separated by an intracellular linker region. The linker has been reported to bind myosin regulatory light chain (RLC), but it is not clear how this can occur in the context of a myosin II complex. We characterized MDR1-RLC interactions and determined that binding occurs via the amino terminal of the RLC, a domain that typically binds myosin heavy chain. MDR1-RLC interactions were sensitive to the phosphorylation state of the light chain in that phosphorylation by myosin light chain kinase (MLCK) resulted in a loss of binding in vitro. We used ML-7, a specific inhibitor of MLCK, to study the functional consequences of disrupting RLC phosphorylation in intact cells. Pretreatment of polarized Madin-Darby canine kidney cells stably expressing MDR1 with ML-7 produced a significant increase in apical to basal permeability and a corresponding decrease in the efflux ratio (threefold; p<0.01) of [3H]-digoxin, a classic MDR1 substrate. Together these data show that MDR1-mediated transport of [3H]-digoxin can be modulated by pharmacological manipulation of myosin RLC, but direct MDR1-RLC interactions are atypical and not explained by the structure of the myosin II holoenzyme.
KW - Digoxin
KW - Myosin light chain kinase
KW - NMDA receptor
KW - Nonmuscle myosin II
KW - Regulatory light chain
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U2 - 10.1016/j.bbrc.2010.05.084
DO - 10.1016/j.bbrc.2010.05.084
M3 - Article
C2 - 20510202
AN - SCOPUS:77954758415
SN - 0006-291X
VL - 398
SP - 7
EP - 12
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
IS - 1
ER -