Urocortin-Induced Decrease in Ca2+ Sensitivity of
Contraction in Mouse Tail Arteries Is Attributable to cAMPDependent Dephosphorylation of MYPT1 and Activation of
Myosin Light Chain Phosphatase
by Lubomir T. Lubomirov, Katrin Reimann, Doris Metzler, Veronika Hasse, Robert
Stehle, Masaaki Ito, David J. Hartshorne, Hristo Gagov, Gabriele Pfitzer, and Rudolf
Schubert
Circulation Research
Volume 98(9):1159-1167
May 12, 2006
Copyright © American Heart Association, Inc. All rights reserved.
Figure 1. Effect of urocortin (UCN-1) on tension and MYPT1 phosphorylation in intact arteries. a,
Concentration-dependent relaxation of arteries preconstricted by KCl (42 mmol/L, n=5) with or
without endothelium (endo).
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 2. Effect of urocortin (UCN-1) on tension and MLC20Ser19 phosphorylation in
permeabilized mouse tail arteries. a and b, Original tension recordings (a) and relaxation
calculated as percentage of submaximal force (b) (n=6). c, Representative Western blots from
exponentially diluted lysates of the arteries. d, Ratio of pMLC20Ser19 to total MLC20 of
densitometric scans of the chemiluminograms (n=9). ***P<0.001.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 3. CRF receptor antagonists inhibit UCN-1-induced relaxation and MLC20Ser19
phosphorylation.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 4. Effect of Rp-8-CPT-cAMPS (PKA-I) and Sp-5,6-DCl-cBIMPs (Sp) on urocortin-induced
relaxation and MLC20Ser19 phosphorylation. a and b, Original force recordings (a) and
summarized data (b) (n=4 to 5; **P<0.01, ***P<0.001). c, Representative Western blots of
exponentially diluted lysates. d, Ratio of pMLC20Ser19 (pMCL20) to total MLC20 of densitometric
scans of the chemiluminograms (n=4). *P<0.05. n.s. indicates not significant.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 5. Effect of urocortin (UCN-1), Sp-5,6-DCl-cBIMPs (Sp), Y-27632, and okadaic acid (OA) on
MYPT1Thr696/Thr850 phosphorylation in submaximally activated permeabilized arteries. a and
b, Representative Western blots (a) and summarized data (b) showing the effect of UCN-1 and
Sp-5,6-DCl-cBIMPs (SP) on pMYPT1Thr696/850 immunoreactivity.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 6. Effect of urocortin (UCN-1) on the time course of relaxation without MLCK activity. a
through d, Time course of relaxation (at 11.4°C) (a); tLIN: duration of the quasilinear phase (b);
kLIN: slope of the initial quasilinear phase (c); kEXP: rate constant of exponential phase (d).
*P<0.05.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 7. Effect of urocortin (UCN-1) and PKA on the time course of dephosphorylation of
MLC20Ser19 and MYPT1Thr696/Thr850 under conditions of inhibited MLCK. Experimental
protocol was as in Figure 6. a and b, Representative Western blots of pMLC20Ser19 (pMLC20)
and total light chains (MLC20 and MLC17). c, Summary of results.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.
Figure 8. Effect of urocortin (UCN-1) on the time course of contraction elicited by incubation with
pCa 6.95 under conditions of inhibited phosphatase activity.
Lubomir T. Lubomirov et al. Circ Res. 2006;98:1159-1167
Copyright © American Heart Association, Inc. All rights reserved.