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Stimulation by insulin of a serine kinase in human platelets that phosphorylates and activates the cGMP-inhibited cAMP phosphodiesterase

Lopez-Aparicio, Pilar; Belfrage, Per LU ; Manganiello, Vincent C; Kono, Tetsuro and Degerman, Eva LU (1993) In Biochemical and Biophysical Research Communications 193(3). p.1137-1144
Abstract
We previously reported that insulin stimulation of human platelets induces serine phosphorylation and activation of the cGMP-inhibited cAMP phosphodiesterase (cGI-PDE). Here, we describe methods to detect and partially purify an insulin-stimulated cGI-PDE kinase (cGI-PDE ISK) from lysates of platelets incubated with insulin. Incubation of human platelets with 10(-8) M insulin increased cGI-PDE ISK activity two-fold. The DEAE-Sephacel-purified cGI-PDE ISK phosphorylated the cGI-PDE on serine in a time- and concentration-dependent manner resulting in an increased incorporation of about 0.2 mol of [32P]/mol of cGI-PDE and 15-20% increase in cGI-PDE activity. The phosphorylation of cGI-PDE was not affected by 10 microM PKI, 1 microgram/ml of... (More)
We previously reported that insulin stimulation of human platelets induces serine phosphorylation and activation of the cGMP-inhibited cAMP phosphodiesterase (cGI-PDE). Here, we describe methods to detect and partially purify an insulin-stimulated cGI-PDE kinase (cGI-PDE ISK) from lysates of platelets incubated with insulin. Incubation of human platelets with 10(-8) M insulin increased cGI-PDE ISK activity two-fold. The DEAE-Sephacel-purified cGI-PDE ISK phosphorylated the cGI-PDE on serine in a time- and concentration-dependent manner resulting in an increased incorporation of about 0.2 mol of [32P]/mol of cGI-PDE and 15-20% increase in cGI-PDE activity. The phosphorylation of cGI-PDE was not affected by 10 microM PKI, 1 microgram/ml of heparin, 3 mM CaCl2 or 1 mM MnCl2. cGI-PDE ISK did not adsorb to antiphosphotyrosine antibodies. To maintain its activation it was necessary to add protein phosphatase inhibitors to the lysate-buffers. All of these findings are consistent with the conclusion that a serine/threonine phosphorylation of the cGI-PDE ISK is involved in its activation by insulin. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biochemical and Biophysical Research Communications
volume
193
issue
3
pages
1137 - 1144
publisher
Elsevier
external identifiers
  • pmid:7686745
  • scopus:0027326931
ISSN
1090-2104
DOI
10.1006/bbrc.1993.1744
language
English
LU publication?
yes
id
26f93135-24c5-465f-83e1-1aae3e693850 (old id 1107504)
date added to LUP
2008-07-31 10:41:19
date last changed
2017-01-01 06:52:52
@article{26f93135-24c5-465f-83e1-1aae3e693850,
  abstract     = {We previously reported that insulin stimulation of human platelets induces serine phosphorylation and activation of the cGMP-inhibited cAMP phosphodiesterase (cGI-PDE). Here, we describe methods to detect and partially purify an insulin-stimulated cGI-PDE kinase (cGI-PDE ISK) from lysates of platelets incubated with insulin. Incubation of human platelets with 10(-8) M insulin increased cGI-PDE ISK activity two-fold. The DEAE-Sephacel-purified cGI-PDE ISK phosphorylated the cGI-PDE on serine in a time- and concentration-dependent manner resulting in an increased incorporation of about 0.2 mol of [32P]/mol of cGI-PDE and 15-20% increase in cGI-PDE activity. The phosphorylation of cGI-PDE was not affected by 10 microM PKI, 1 microgram/ml of heparin, 3 mM CaCl2 or 1 mM MnCl2. cGI-PDE ISK did not adsorb to antiphosphotyrosine antibodies. To maintain its activation it was necessary to add protein phosphatase inhibitors to the lysate-buffers. All of these findings are consistent with the conclusion that a serine/threonine phosphorylation of the cGI-PDE ISK is involved in its activation by insulin.},
  author       = {Lopez-Aparicio, Pilar and Belfrage, Per and Manganiello, Vincent C and Kono, Tetsuro and Degerman, Eva},
  issn         = {1090-2104},
  language     = {eng},
  number       = {3},
  pages        = {1137--1144},
  publisher    = {Elsevier},
  series       = {Biochemical and Biophysical Research Communications},
  title        = {Stimulation by insulin of a serine kinase in human platelets that phosphorylates and activates the cGMP-inhibited cAMP phosphodiesterase},
  url          = {http://dx.doi.org/10.1006/bbrc.1993.1744},
  volume       = {193},
  year         = {1993},
}