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Activation of a cGMP-stimulated cAMP phosphodiesterase by protein kinase C in a liver Golgi-endosomal fraction.
- Source :
-
European journal of biochemistry [Eur J Biochem] 1999 Feb; Vol. 259 (3), pp. 892-900. - Publication Year :
- 1999
-
Abstract
- The ability of Ca2+/phospholipid-dependent protein kinase (protein kinase C, PKC) to stimulate cAMP phosphodiesterase (PDE) activity in a liver Golgi-endosomal (GE) fraction was examined in vivo and in a cell-free system. Injection into rats of 4 beta-phorbol 12-myristate 13-acetate, a known activator of PKC, caused a rapid and marked increase in PKC activity (+325% at 10 min) in the GE fraction, along with an increase in the abundance of the PKC alpha-isoform as seen on Western immunoblots. Concurrently, 4 beta-phorbol 12-myristate 13-acetate treatment caused a time-dependent increase in cAMP PDE activity in the GE fraction (96% at 30 min). Addition of the catalytic subunit of protein kinase A (PKA) to GE fractions from control and 4 beta-phorbol 12-myristate 13-acetate-treated rats led to a comparable increase (130-150%) in PDE activity, suggesting that PKA is probably not involved in the in-vivo effect of 4 beta-phorbol 12-myristate 13-acetate. In contrast, addition of purified PKC increased (twofold) PDE activity in GE fractions from control rats but affected only slightly the activity in GE fractions from 4 beta-phorbol 12-myristate 13-acetate-treated rats. About 50% of the Triton-X-100-solubilized cAMP PDE activity in the GE fraction was immunoprecipitated with an anti-PDE3 antibody. On DEAE-Sephacel chromatography, three peaks of PDE were sequentially eluted: one early peak, which was stimulated by cGMP and inhibited by erythro-9 (2-hydroxy-3-nonyl) adenine (EHNA); a selective inhibitor of type 2 PDEs; and two retarded peaks of activity, which were potently inhibited by cGMP and cilostamide, an inhibitor of type 3 PDEs. Further characterization of peak I by HPLC resolved a major peak which was activated (threefold) by 5 microM cGMP and inhibited (87%) by 25 microM EHNA, and a minor peak which was insensitive to EHNA and cilostamide. 4 beta-Phorbol 12-myristate 13-acetate treatment caused a selective increase (2.5-fold) in the activity associated with DEAE-Sephacel peak I, without changing the K(m) value. These results suggest that PKC selectively activates a PDE2, cGMP-stimulated isoform in the GE fraction.
- Subjects :
- Adenine analogs & derivatives
Adenine pharmacology
Animals
Cyclic AMP-Dependent Protein Kinases metabolism
Enzyme Activation
Enzyme Inhibitors
Male
Naphthalenes pharmacology
Phosphodiesterase Inhibitors pharmacology
Quinolones pharmacology
Rats
Rats, Sprague-Dawley
Tetradecanoylphorbol Acetate pharmacology
Time Factors
3',5'-Cyclic-AMP Phosphodiesterases metabolism
Cyclic GMP pharmacology
Endosomes enzymology
Golgi Apparatus enzymology
Liver enzymology
Protein Kinase C metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0014-2956
- Volume :
- 259
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- European journal of biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 10092879
- Full Text :
- https://doi.org/10.1046/j.1432-1327.1999.00123.x