Site-specific phosphorylation and point mutations of telokin modulate its Ca2+-desensitizing effect in smooth muscle.
Forskolin and 8-bromoguanosine 3'-5'-cyclic monophosphate (8-Br-cGMP) induce phosphorylation of Ser-13 of telokin and relaxation of smooth muscle at constant calcium. Comparison with the effect of wild type with aspartate (D; to mimic phosphorylation) and alanine (A; non-phosphorylatable) mutants of telokin showed that the S13D mutant was more effective than wild type in relaxing smooth muscle at constant calcium. The efficacy of the Ser-13A, S12A, and S12D mutants was not significantly different from that of wild-type telokin. The effect of neither S13D nor Ser-13A was affected by 8-Br-cGMP, whereas the effect of wild type, S12A, and S12D was enhanced by 8-Br-cGMP, indicating the specificity of Ser-13 charge modification. Mutation of Ser-19 (a mitogen-activated protein kinase site) showed the S19A to be more effective than, and S19D to be not different from, wild-type telokin. The effect of both mutants was slightly enhanced by 8-Br-cGMP. A truncated (residues 1-142) form lacking the acidic C terminus had the same relaxant effect as wild-type telokin, whereas the C-terminal peptide (residues 142-155) had no effect. We conclude that site-specific modification of the N terminus modulates the Ca2+ -desensitizing effect of telokin on force.
Walker
L A
LA
Department of Molecular Physiology and Biological Physics, Health System, Charlottesville, VA 22908-0736, USA.
MacDonald
J A
JA
Liu
X
X
Nakamoto
R K
RK
Haystead
T A
TA
Somlyo
A V
AV
Somlyo
A P
AP
D019943:Amino Acid Substitution; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D046508:Culture Techniques; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D007082:Ileum / Q000187:drug effects / Q000378:metabolism; D052998:Microcystins; D008854:Microscopy, Electron; D009124:Muscle Proteins / Q000235:genetics* / Q000378:metabolism*; D009126:Muscle Relaxation / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D009219:Myosin-Light-Chain Kinase; D010446:Peptide Fragments; D010455:Peptides; D010456:Peptides, Cyclic / Q000494:pharmacology; D010766:Phosphorylation; D017354:Point Mutation; D011817:Rabbits; D012694:Serine / Q000378:metabolism
D052998:Microcystins; D009124:Muscle Proteins; D010446:Peptide Fragments; D010455:Peptides; D010456:Peptides, Cyclic; D005576:Colforsin; D001224:Aspartic Acid; C016276:8-bromocyclic GMP; C060640:telokin; D012694:Serine; C078588:microcystin; D009219:Myosin-Light-Chain Kinase; D006152:Cyclic GMP; D002118:Calcium