PMID- 10601309
OWN - NLM
STAT- MEDLINE
DCOM- 20000113
LR  - 20210209
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 274
IP  - 52
DP  - 1999 Dec 24
TI  - Inhibitory phosphorylation site for Rho-associated kinase on smooth muscle myosin
      phosphatase.
PG  - 37385-90
AB  - It is clear from several studies that myosin phosphatase (MP) can be inhibited
      via a pathway that involves RhoA. However, the mechanism of inhibition is not
      established. These studies were carried out to test the hypothesis that
      Rho-kinase (Rho-associated kinase) via phosphorylation of the myosin phosphatase 
      target subunit 1 (MYPT1) inhibited MP activity and to identify relevant sites of 
      phosphorylation. Phosphorylation by Rho-kinase inhibited MP activity and this
      reflected a decrease in V(max). Activity of MP with different substrates also was
      inhibited by phosphorylation. Two major sites of phosphorylation on MYPT1 were
      Thr(695) and Thr(850). Various point mutations were designed for these
      phosphorylation sites. Following thiophosphorylation by Rho-kinase and assays of 
      phosphatase activity it was determined that Thr(695) was responsible for
      inhibition. A site- and phosphorylation-specific antibody was developed for the
      sequence flanking Thr(695) and this recognized only phosphorylated Thr(695) in
      both native and recombinant MYPT1. Using this antibody it was shown that
      stimulation of serum-starved Swiss 3T3 cells by lysophosphatidic acid, thought to
      activate RhoA pathways, induced an increase in Thr(695) phosphorylation on MYPT1 
      and this effect was blocked by a Rho-kinase inhibitor, Y-27632. In summary, these
      results offer strong support for a physiological role of Rho-kinase in regulation
      of MP activity.
FAU - Feng, J
AU  - Feng J
AD  - First Department of Internal Medicine, Mie University School of Medicine, Tsu,
      Mie 514-8507, Japan.
FAU - Ito, M
AU  - Ito M
FAU - Ichikawa, K
AU  - Ichikawa K
FAU - Isaka, N
AU  - Isaka N
FAU - Nishikawa, M
AU  - Nishikawa M
FAU - Hartshorne, D J
AU  - Hartshorne DJ
FAU - Nakano, T
AU  - Nakano T
LA  - eng
GR  - HL23615/HL/NHLBI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (Amides)
RN  - 0 (Intracellular Signaling Peptides and Proteins)
RN  - 0 (Lysophospholipids)
RN  - 0 (Pyridines)
RN  - 138381-45-0 (Y 27632)
RN  - 2ZD004190S (Threonine)
RN  - EC 2.7.11.1 (Protein-Serine-Threonine Kinases)
RN  - EC 2.7.11.1 (rho-Associated Kinases)
RN  - EC 3.1.3.16 (Phosphoprotein Phosphatases)
RN  - EC 3.1.3.53 (Myosin-Light-Chain Phosphatase)
SB  - IM
MH  - 3T3 Cells
MH  - Amides/pharmacology
MH  - Animals
MH  - Antibody Specificity
MH  - Intracellular Signaling Peptides and Proteins
MH  - Lysophospholipids/pharmacology
MH  - Mice
MH  - Muscle, Smooth/*enzymology
MH  - Mutagenesis, Site-Directed
MH  - Myosin-Light-Chain Phosphatase
MH  - Phosphoprotein Phosphatases/chemistry/immunology/*metabolism
MH  - Phosphorylation
MH  - Protein-Serine-Threonine Kinases/*physiology
MH  - Pyridines/pharmacology
MH  - Threonine
MH  - rho-Associated Kinases
EDAT- 1999/12/22 00:00
MHDA- 1999/12/22 00:01
CRDT- 1999/12/22 00:00
PHST- 1999/12/22 00:00 [pubmed]
PHST- 1999/12/22 00:01 [medline]
PHST- 1999/12/22 00:00 [entrez]
AID - 10.1074/jbc.274.52.37385 [doi]
AID - S0021-9258(19)53105-2 [pii]
PST - ppublish
SO  - J Biol Chem. 1999 Dec 24;274(52):37385-90. doi: 10.1074/jbc.274.52.37385.