Association of Protein Kinase Cμ with Type II Phosphatidylinositol 4-Kinase and Type I Phosphatidylinositol-4-phosphate 5-Kinase

1. DEAE-cellulose chromatography of mouse brain cytosol indicated the presence of only the type II isoenzyme of cyclic AMP-dependent protein kinase. Mouse heart cytosol contained approximately equal amounts of the type I and type II isoenzymes. 2. Both brain and heart type II isoenzymes reassociated after a transient exposure to cyclic AMP, but the heart type I isoenzyme remained dissociated. 3. Elution of brain cytosol continuously exposed to cyclic AMP resolved multiple peaks of protein kinase and cyclic AMP-binding activities. A single peak of kinase and multiple peaks of cyclic AMP-binding activities were found under the same conditions with heart cytosol. Various control experiments suggested that the heterogeneity within the brain type II isoenzymic class had not been caused by proteolysis. 4. Kinetic experiments with unfractionated brain cytosol showed that the binding of cyclic AMP, the dissociation of cyclic AMP from protein and the rate of heat denaturation of the cyclic AMP-binding activity gave results consistent with the presence of multiple binding species. 5. It concluded that the type II isoenzymic peak obtained by DEAE-cellulose chromatography of mouse brain cytosol represents a class of enzymes containing multiple regulatory and catalytic subunits. The two heart cytosol isoenzymes contain a common catalytic subunit. The degree of protein kinase ‘microheterogeneity”, defined as the presence of multiple regulatory and/or catalytic subunits within a single isoenzymic class, appears to be tissue-specific.

Download Full-text

Elution of the regulatory subunit of cAMP-dependent protein kinase Type I isozyme derived from epididymal fat within the Type II isozyme chromatographic peak

Biochemical and Biophysical Research Communications ◽

10.1016/0006-291x(83)91818-1 ◽

1983 ◽

Vol 112 (1) ◽

pp. 214-220 ◽

Cited By ~ 8

Author(s):

Alvin M. Malkinson ◽

Deborah S. Beer ◽

Jeanne M. Wehner ◽

J.R. Sheppard

Keyword(s):

Protein Kinase ◽

Regulatory Subunit ◽

Type I ◽

Type Ii ◽

Chromatographic Peak ◽

Dependent Protein Kinase ◽

Camp Dependent Protein Kinase ◽

Epididymal Fat ◽

Dependent Protein

Download Full-text

Interleukin-1 alpha stimulates dopamine release by activating type II protein kinase A in PC-12 cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1995.269.6.e1083 ◽

1995 ◽

Vol 269 (6) ◽

pp. E1083-E1088

Author(s):

A. Joseph ◽

A. Kumar ◽

N. A. O'Connell ◽

R. K. Agarwal ◽

A. R. Gwosdow

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Dopamine Release ◽

Interleukin 1 ◽

Intracellular Camp ◽

Type I ◽

Camp Accumulation ◽

Type Ii ◽

Pc 12 Cells ◽

Kinase A

A recent study from this laboratory [A. R. Gwosdow, N. A. O'Connell, and A. B. Abou-Samra. Am. J. Physiol. 263 (Endocrinol. Metab. 26): E461-E466, 1992] showed that the inflammatory mediator interleukin-1 alpha (IL-1 alpha) stimulates catecholamine release from primary cultures of rat adrenal cells. The present studies were conducted to determine whether 1) IL-1 alpha stimulates catecholamine/dopamine release from the adrenal medullary cell line PC-12 and 2) the adenosine 3',5'-cyclic monophosphate (cAMP)-protein kinase A (PKA) pathway is involved in IL-1 alpha-induced dopamine release from PC-12 cells. The results indicate that IL-1 alpha significantly (P < 0.05) elevated dopamine release after a 24-h incubation period. IL-1 alpha did not stimulate cAMP accumulation at any time period between 5 min and 2 h. In contrast, forskolin-treated cells elevated (P < 0.05) intracellular cAMP levels and increased dopamine release. Because IL-1 alpha did not affect cAMP accumulation, the effect of IL-1 alpha on PKA activity was investigated. IL-1 alpha increased (P < 0.05) PKA activity at 15 and 30 min and returned to control levels by 1 h. Forskolin also increased (P < 0.05) PKA activity. The type of PKA activated (P < 0.05) by IL-1 alpha was type II PKA. In contrast, forskolin activated (P < 0.05) type I and type II PKA. Inhibition of PKA with the PKA inhibitor H-8 blocked PKA activity and dopamine secretion by both IL-1 alpha and forskolin in PC-12 cells. These observations demonstrate that 1) IL-1 alpha stimulated dopamine release from PC-12 cells by activating PKA, 2) the mechanism of IL-1 alpha activation of PKA does not involve detectable increases in intracellular cAMP accumulation, and 3) IL-1 alpha activates type II PKA, which is used by IL-1 alpha to stimulate dopamine secretion from PC-12 cells.

Download Full-text

Differential localization of protein kinase C isozymes in retinal neurons.

The Journal of Cell Biology ◽

10.1083/jcb.112.6.1241 ◽

1991 ◽

Vol 112 (6) ◽

pp. 1241-1247 ◽

Cited By ~ 48

Author(s):

N Usuda ◽

Y Kong ◽

M Hagiwara ◽

C Uchida ◽

M Terasawa ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Bipolar Cells ◽

Type I ◽

Rabbit Retina ◽

Type Ii ◽

Retinal Neurons ◽

Type Iii ◽

Kinase C ◽

Rod Bipolar Cells

We report the immunohistochemical localization of protein kinase C isozymes (types I, II, and III) in the rabbit retina using the monospecific monoclonal antibodies MC-1a, MC-2a, and MC-3a. Using immunoblot analysis of partially purified protein kinase C preparations of rabbit retina, types II and III isozymes alone were detected. The activity of type III was the stronger. By light microscopic immunohistochemical analysis, retinal neurons were negative for type I and positive for type II and type III isozymes. Type II was more diffusely distributed through the retinal layers, but was distinctive in ganglion cells, bipolar cells, and outer segments. The immunoreactivity was stronger for type III isozyme, and it was observed in mop (rod) bipolar cells and amacrine cells. By using immunoelectron microscopy, the cytoplasm of the cell body, the axon, and dendrites of the mop bipolar cells were strongly immunoreactive for type III. The so-called rod bipolar cells were for the first time seen to form synapses with rod photoreceptor cells. These differential localizations of respective isozymes in retinal neurons suggest that each isozyme has a different site of function in each neuron.

Download Full-text

Purification of PKC-I, an endogenous protein kinase C inhibitor, and types II and III protein kinase C isoenzymes from human neutrophils

Biochemical Journal ◽

10.1042/bj2840399 ◽

1992 ◽

Vol 284 (2) ◽

pp. 399-405 ◽

Cited By ~ 10

Author(s):

K J Balazovich ◽

E L McEwen ◽

M L Lutzke ◽

L A Boxer ◽

T White

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Human Neutrophils ◽

Type I ◽

Dependent Manner ◽

Type Ii ◽

Western Blots ◽

Type Iii ◽

Kinase C ◽

Endogenous Protein

Human neutrophil protein kinase C (PKC) activity is inhibited by an endogenous protein found primarily in the pellet fraction from homogenized specific granules, which was both heat- and proteinase-sensitive [Balazovich, Smolen & Boxer (1986) J. Immunol. 137, 1665-1673]. We now report that two PKC isoenzymes and the endogenous PKC inhibitor, which we named PKC-I, were purified from human neutrophils. A neutrophil soluble fraction that was subjected to DEAE-Sephacel chromatography yielded highly enriched PKC because, by definition, enzymic activity was strictly dependent on Ca2+ and phosphatidylserine. Hydroxyapatite chromatography resolved two peaks of PKC activity. Type II and Type III PKC isoenzymes were each identified on Western blots by using isoenzyme-specific monoclonal antibodies. Unlike rat brain, from which PKC isoenzymes were also purified, Type I PKC was not detected in human neutrophils. Western blots indicated that both Type II and Type III PKC isoenzymes had molecular masses near 80 kDa. In agreement with other reports, PKC was autophosphorylated in vitro. PKC-I, an endogenous neutrophil inhibitor of PKC, was purified to apparent homogeneity by DEAE-Sephacel and S-400 Sephacel chromatography. PKC-I had a molecular mass of 41 kDa. PKC-I inhibited purified PKC activity stimulated by 1,2-diacylglycerols in a concentration-dependent manner, and inhibited PKC-dependent phosphorylation of proteins present in neutrophil cytosol.

Download Full-text

Purification of a yeast protein kinase sharing properties with type I and type II casein kinases

Biochemistry ◽

10.1021/bi00388a005 ◽

1987 ◽

Vol 26 (14) ◽

pp. 4207-4212 ◽

Cited By ~ 12

Author(s):

Hans Sternbach ◽

Hans Kuentzel

Keyword(s):

Protein Kinase ◽

Type I ◽

Type Ii ◽

Yeast Protein ◽

Casein Kinases

Download Full-text

Activation of Both Protein Kinase A (PKA) Type I and PKA Type II Isozymes Is Required for Retinoid-Induced Maturation of Acute Promyelocytic Leukemia Cells

Molecular Pharmacology ◽

10.1124/mol.112.081034 ◽

2013 ◽

Vol 83 (5) ◽

pp. 1057-1065 ◽

Cited By ~ 10

Author(s):

Eric Nguyen ◽

Gro Gausdal ◽

Jacqueline Varennes ◽

Frédéric Pendino ◽

Michel Lanotte ◽

...

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Acute Promyelocytic Leukemia ◽

Promyelocytic Leukemia ◽

Leukemia Cells ◽

Type I ◽

Type Ii ◽

Kinase A

Download Full-text

Human T lymphocyte cAMP‐dependent protein kinase: subcellular distributions and activity ranges of type I and type II isozymes

The FASEB Journal ◽

10.1096/fasebj.6.9.1319361 ◽

1992 ◽

Vol 6 (9) ◽

pp. 2735-2741 ◽

Cited By ~ 17

Author(s):

Paul Hasler ◽

John J. Moore ◽

Gary M. Kammer

Keyword(s):

Protein Kinase ◽

T Lymphocyte ◽

Type I ◽

Type Ii ◽

Dependent Protein Kinase ◽

Camp Dependent Protein Kinase ◽

Human T Lymphocyte ◽

Dependent Protein

Download Full-text

Isolation of two myosin light-chain kinases from bovine carotid artery and their regulation by phosphorylation mediated by cyclic AMP-dependent protein kinase

Biochemical Journal ◽

10.1042/bj2030583 ◽

1982 ◽

Vol 203 (3) ◽

pp. 583-592 ◽

Cited By ~ 17

Author(s):

Ramesh C. Bhalla ◽

Ram V. Sharma ◽

Ramesh C. Gupta

Keyword(s):

Protein Kinase ◽

Carotid Artery ◽

Cyclic Amp ◽

Light Chain ◽

Myosin Light Chain ◽

Kinase Activity ◽

Type I ◽

Type Ii ◽

Dependent Protein Kinase ◽

Dependent Protein

Myosin light-chain kinase was purifed from bovine carotid artery. Approx. 90% of myosin kinase was extracted in the supernatant fraction with buffer containing EDTA during myofibril preparation. The soluble fraction yielded two distinct peaks on DEAE-Sephacel chromatography. Peak I was eluted at a conductance of 11–12mmho and was completely dependent on Ca2+–calmodulin for its activity. Peak II was eluted at a conductance of 13–14mmho and showed approx. 15% Ca2+-independent activity. The myosin kinases I and II were further purified by affinity chromatography by using calmodulin coupled to Sepharose 4B, which resulted in 960-and 650-fold purification of type I and type II kinases respectively. Myosin kinase II activity was completely Ca2+-dependent after affinity chromatography on the calmodulin–Sepharose column. Myosin kinases I and II were phosphorylated by cyclic AMP-dependent protein kinase. In the presence of bound calmodulin 0.5–0.7mol of phosphate was incorporated/mol of myosin kinases I and II. On the other hand, in the absence of bound calmodulin 1–1.4mol of phosphate was incorporated/mol of kinases I and II. Phosphorylation in the absence of calmodulin significantly decreased the myosin kinase activity of both enzymes, and the decrease in myosin kinase activity was due to a 3–5-fold increase in the amount of calmodulin required for half-maximal stimulation of both type I and type II kinases. The regulation of myosin kinase activity by cyclic AMP-dependent phosphorylation would suggest that β-adrenergic-mediated relaxation of vascular smooth muscle may be partly due to the direct interaction of cyclic AMP at the site of contractile proteins.

Download Full-text