The phosphorylation motif at serine 225 governs the localization and function of sphingosine kinase 1 in resistance arteries.
OBJECTIVE
The purpose of this study was to characterize a phosphorylation motif at serine 225 as a molecular switch that regulates the pressure-dependent activation of sphingosine kinase 1 (Sk1) in resistance artery smooth muscle cells.


RESULTS
In isolated hamster gracilis muscle resistance arteries, pressure-dependent activation/translocation of Sk1 by ERK1/2 was critically dependent on its serine 225 phosphorylation site. Specifically, expression of Sk1(S225A) reduced resting and myogenic tone, resting Ca(2+), pressure-induced Ca(2+) elevations, and Ca(2+) sensitivity. The lack of function of the Sk1(S225A) mutant could not be entirely overcome by forced localization to the plasma membrane via a myristoylation/palmitylation motif; the membrane anchor also significantly inhibited the function of the wild-type Sk1 enzyme. In both cases, Ca(2+) sensitivity and myogenic tone were attenuated, whereas Ca(2+) handling was normalized/enhanced. These discrete effects are consistent with cell surface receptor-mediated effects (Ca(2+) sensitivity) and intracellular effects of S1P (Ca(2+) handling). Accordingly, S1P(2) receptor inhibition (1 micromol/L JTE013) attenuated myogenic tone without effect on Ca(2+).


CONCLUSIONS
Translocation and precise subcellular positioning of Sk1 is essential for full Sk1 function; and two distinct S1P pools, proposed to be intra- and extracellular, contribute to the maintenance of vascular tone.
Lidington
Darcy
D
Department of Physiology, University of Toronto, Toronto, Ontario, Canada.
Peter
Bernhard Friedrich
BF
Meissner
Anja
A
Kroetsch
Jeffrey T
JT
Pitson
Stuart M
SM
Pohl
Ulrich
U
Bolz
Steffen-Sebastian
SS
D000704:Analysis of Variance; D000818:Animals; D001158:Arteries / Q000201:enzymology*; D020013:Calcium Signaling / Q000502:physiology; D002463:Cell Membrane Permeability / Q000502:physiology; D002478:Cells, Cultured; D006224:Cricetinae; D009131:Muscle, Smooth, Vascular / Q000098:blood supply* / Q000166:cytology / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011312:Pressure; D011336:Probability; D012694:Serine / Q000378:metabolism*; D015398:Signal Transduction; D014655:Vascular Resistance / Q000502:physiology*; D014661:Vasoconstriction / Q000502:physiology; D000099157:Sphingosine Kinase
D017853:Phosphotransferases (Alcohol Group Acceptor); D012694:Serine; D000099157:Sphingosine Kinase