scholarly journals Mechanism of A-kinase-anchoring protein 79 (AKAP79) and protein kinase C interaction

1999 ◽  
Vol 343 (2) ◽  
pp. 443-452 ◽  
Author(s):  
Maree C. FAUX ◽  
Emily N. ROLLINS ◽  
Amelia S. EDWARDS ◽  
Lorene K. LANGEBERG ◽  
Alexandra C. NEWTON ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Protein Interactions ◽  
Hippocampal Neurons ◽  
Kinase Activity ◽  
Catalytic Core ◽  
Kinase C ◽  
Anchoring Protein ◽  
Arginine Residues

The A-kinase-anchoring protein AKAP79 co-ordinates the location of cAMP-dependent protein kinase, phosphatase 2B (PP2B/calcineurin) and protein kinase C (PKC) at postsynaptic sites in neurons. In this report we focus on the mechanism of interaction between AKAP79 and PKC. We show that neither lipid activators nor kinase activation are required for association with AKAP79. The anchoring protein binds and inhibits the conserved catalytic core of PKCβII. AKAP79 also associates with conventional, novel and atypical isoforms of PKC in vitro andin vivo, and immunofluorescence staining of rat hippocampal neurons demonstrates that the murine anchoring-protein homologue AKAP150 is co-distributed with PKCα/β, PKCε or PKCℓ. Binding of the AKAP79(31-52) peptide, which inhibits kinase activity, exposes the pseudosubstrate domain of PKCβII, allowing endoproteinase Arg-C proteolysis in the absence of kinase activators. Reciprocal experiments have identified two arginine residues at positions 39 and 40 that are essential for AKAP79(31-52) peptide inhibition of PKCβII. Likewise, the same mutations in the full-length anchoring protein reduced inhibition of PKCβII. Thus AKAP79 associates with multiple PKC isoforms through a mechanism involving protein-protein interactions at the catalytic core where binding of the anchoring protein inhibits kinase activity through displacement of the pseudosubstrate.

RACK1, a Protein Kinase C Anchoring Protein, Coordinates the Binding of Activated Protein Kinase C and Select Pleckstrin Homology Domains in Vitro†

Biochemistry ◽  
1999 ◽  
Vol 38 (42) ◽  
pp. 13787-13794 ◽  
Author(s):  
Manuel M. Rodriguez ◽  
Dorit Ron ◽  
Kazushige Touhara ◽  
Che-Hong Chen ◽  
Daria Mochly-Rosen
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Pleckstrin Homology ◽  
Kinase C ◽  
Anchoring Protein ◽  
Homology Domains

scholarly journals 6,6′-Dihydroxythiobinupharidine (DTBN) Purified from Nuphar lutea Leaves Is an Inhibitor of Protein Kinase C Catalytic Activity

Molecules ◽  
2021 ◽  
Vol 26 (9) ◽  
pp. 2785
Author(s):  
Kamran Waidha ◽  
Nikhil Ponnoor Anto ◽  
Divya Ram Jayaram ◽  
Avi Golan-Goldhirsh ◽  
Saravanakumar Rajendran ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Kinase Domain ◽  
Water Lily ◽  
Activity Data ◽  
Nuphar Lutea ◽  
Kinase C ◽  
Disease Biology

Water lily (Nuphar) bioactive extracts have been widely used in traditional medicine owing to their multiple applications against human ailments. Phyto-active Nuphar extracts and their purified and synthetic derivatives have attracted the attention of ethnobotanists and biochemists. Here, we report that 6,6′-dihydroxythiobinupharidine (DTBN), purified from extracts of Nuphar lutea (L.) Sm. leaves, is an effective inhibitor of the kinase activity of members of the protein kinase C (PKC) family using in vitro and in silico approaches. We demonstrate that members of the conventional subfamily of PKCs, PKCα and PKCγ, were more sensitive to DTBN inhibition as compared to novel or atypical PKCs. Molecular docking analysis demonstrated the interaction of DTBN, with the kinase domain of PKCs depicting the best affinity towards conventional PKCs, in accordance with our in vitro kinase activity data. The current study reveals novel targets for DTBN activity, functioning as an inhibitor for PKCs kinase activity. Thus, this and other data indicate that DTBN modulates key cellular signal transduction pathways relevant to disease biology, including cancer.

scholarly journals Three Distinct Mechanisms for Translocation and Activation of the δ Subspecies of Protein Kinase C

1998 ◽  
Vol 18 (9) ◽  
pp. 5263-5271 ◽  
Author(s):  
Shiho Ohmori ◽  
Yasuhito Shirai ◽  
Norio Sakai ◽  
Motoko Fujii ◽  
Hiroaki Konishi ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Fusion Protein ◽  
Kinase Activity ◽  
Fluorescent Protein ◽  
Simultaneous Treatment ◽  
Kinase C ◽  
Green Fluorescent ◽  
Number Of Cells

ABSTRACT We expressed δ subspecies of protein kinase C (δ-PKC) fused with green fluorescent protein (GFP) in CHO-K1 cells and observed the movement of this fusion protein in living cells after three different stimulations. The δ-PKC–GFP fusion protein had enzymological characteristics very similar to those of the native δ-PKC and was present throughout the cytoplasm in CHO-K1 cells. ATP at 1 mM caused a transient translocation of δ-PKC–GFP to the plasma membrane approximately 30 s after the stimulation and a sequent retranslocation to the cytoplasm within 3 min. A tumor-promoting phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (TPA; 1 μM), induced a slower translocation of δ-PKC–GFP, and the translocation was unidirectional. Concomitantly, the kinase activity of δ-PKC–GFP was increased by these two stimulations, when the kinase activity of the immunoprecipitated δ-PKC–GFP was measured in vitro in the absence of PKC activators such as phosphatidylserine and diacylglycerol. Hydrogen peroxide (H2O2; 5 mM) failed to translocate δ-PKC–GFP but increased its kinase activity more than threefold. δ-PKC–GFP was strongly tyrosine phosphorylated when treated with H2O2 but was tyrosine phosphorylated not at all by ATP stimulation and only slightly by TPA treatment. Both TPA and ATP induced the translocation of δ-PKC–GFP even after treatment with H2O2. Simultaneous treatment with TPA and H2O2 further activated δ-PKC–GFP up to more than fivefold. TPA treatment of cells overexpressing δ-PKC–GFP led to an increase in the number of cells in G2/M phase and of dikaryons, while stimulation with H2O2 increased the number of cells in S phase and induced no significant change in cell morphology. These results indicate that at least three different mechanisms are involved in the translocation and activation of δ-PKC.

scholarly journals Caspase processing activates atypical protein kinase C ζ by relieving autoinhibition and destabilizes the protein

2003 ◽  
Vol 375 (3) ◽  
pp. 663-671 ◽  
Author(s):  
Lucinda SMITH ◽  
Zhi WANG ◽  
Jeffrey B. SMITH
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Hela Cells ◽  
Half Life ◽  
Kinase Activity ◽  
Ubiquitin Proteasome System ◽  
Kinase Domain ◽  
Kinase C ◽  
Pkc Ζ

Treatment of HeLa cells with tumour necrosis factor α (TNFα) induced caspase processing of ectopic PKC (protein kinase C) ζ, which converted most of the holoenzyme into the freed kinase domain and increased immune-complex kinase activity. The goal of the present study was to determine the basis for the increased kinase activity that is associated with caspase processing of PKC ζ. Atypical PKC ι is largely identical with PKC ζ, except for a 60-amino-acid segment that lacks the caspase-processing sites of the ζ isoform. Replacement of this segment of PKC ζ with the corresponding segment of PKC ι prevented caspase processing and activation of the kinase function. Processing of purified recombinant PKC ζ by caspase 3 in vitro markedly increased its kinase activity. Caspase processing activated PKC ζ in vitro or intracellularly without increasing the phosphorylation of Thr410 of PKC ζ, which is required for catalytic competency. The freed kinase domain of PKC ζ had a much shorter half-life than the holoenzyme in transfected HeLa cells and in non-transfected kidney epithelial cells. Treatment with TNF-α shortened the half-life of the kinase domain protein, and proteasome blockade stabilized the protein. Studies of kinase-domain mutants indicate that a lack of negative charge at Thr410 can shorten the half-life of the freed kinase domain. The present findings indicate that the freed kinase domain has substantially higher kinase activity and a much shorter half-life than the holoenzyme because of accelerated degradation by the ubiquitin–proteasome system.

Coordinate regulation of pp90rsk and a distinct protein-serine/threonine kinase activity that phosphorylates recombinant pp90rsk in vitro

1991 ◽  
Vol 11 (4) ◽  
pp. 1868-1874
Author(s):  
J Chung ◽  
R H Chen ◽  
J Blenis
Keyword(s):  
Protein Kinase C ◽  
Growth Factor ◽  
Protein Kinase ◽  
Protein Kinases ◽  
Kinase Activity ◽  
Cultured Cells ◽  
Protein Kinase Activity ◽  
Kinase C ◽  
Dependent Protein

Protein kinase assays that use recombinant pp90rsk as a substrate were developed in an attempt to identify growth-regulated enzymes responsible for the phosphorylation and activation of pp90rsk S6 phosphotransferase activity. With this assay we have ientified a pp60v-src-, growth factor-, phorbol ester-, and vanadate-regulated serine/threonine protein kinase activity that is not related to two other cofactor-independent, growth-regulated protein kinases, pp70-S6 protein kinase and pp90rsk. The pp90rsk-protein kinase activity (referred to as rsk-kinase) is also not related to cofactor-dependent signal transducing protein kinases such as the cyclic AMP-dependent protein kinases, members of the protein kinase C family, or other Ca2(+)-dependent protein kinases. In vitro, partially purified rsk-kinase phosphorylates several of the sites (serine and threonine) that are phosphorylated in growth-stimulated cultured cells. A detailed examination of the mitogen-regulated activation kinetics of rsk-kinase and pp90rsk activities demonstrated that they are coordinately regulated. In addition, protein kinase C is not absolutely required for epidermal and fibroblast growth factor-stimulated activation of rsk-kinase, whereas, like pp90rsk, platelet-derived growth factor- and vanadate-stimulated rsk-kinase activity exhibits a greater dependence on protein kinase C-mediated signal transduction. The characterization and future purification of the rsk-kinase(s) will improve our understanding of the early signaling events regulating cell growth.

scholarly journals GAK and PRKCD are positive regulators of PRKN-independent mitophagy

2020 ◽  
Author(s):  
Michael J. Munson ◽  
Benan J. Mathai ◽  
Laura Trachsel ◽  
Matthew Yoke Wui Ng ◽  
Laura Rodriguez de la Ballina ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Significant Inhibition ◽  
Kinase C ◽  
Positive Regulators ◽  
Sirna Library ◽  
Cyclin G

ABSTRACTThe mechanisms involved in programmed or damage-induced removal of mitochondria by mitophagy in response to different stimuli remains elusive. Here, we have screened for regulators of PRKN-independent mitophagy using an siRNA library targeting 197 proteins containing lipid interacting domains. We identify Cyclin G-associated kinase (GAK) and Protein Kinase C Delta (PRKCD) as novel regulators of PRKN-independent mitophagy, with both being dispensable for PRKN-dependent mitophagy and starvation-induced autophagy. We demonstrate that the kinase activity of both GAK and PRKCD are required for efficient mitophagy in vitro, that PRKCD is present on mitochondria, and that PRKCD is required for ULK1/ATG13 recruitment to early autophagic structures. Importantly, we demonstrate in vivo relevance for both kinases in the regulation of basal mitophagy. Knockdown of GAK homologue (gakh-1) in C.elegans or PRKCD homologues in zebrafish led to significant inhibition of basal mitophagy, highlighting the evolutionary relevance of these kinases in mitophagy.

scholarly journals Coordinate regulation of pp90rsk and a distinct protein-serine/threonine kinase activity that phosphorylates recombinant pp90rsk in vitro.

1991 ◽  
Vol 11 (4) ◽  
pp. 1868-1874 ◽  
Author(s):  
J Chung ◽  
R H Chen ◽  
J Blenis
Keyword(s):  
Protein Kinase C ◽  
Growth Factor ◽  
Protein Kinase ◽  
Protein Kinases ◽  
Kinase Activity ◽  
Cultured Cells ◽  
Protein Kinase Activity ◽  
Kinase C ◽  
Dependent Protein

Protein kinase assays that use recombinant pp90rsk as a substrate were developed in an attempt to identify growth-regulated enzymes responsible for the phosphorylation and activation of pp90rsk S6 phosphotransferase activity. With this assay we have ientified a pp60v-src-, growth factor-, phorbol ester-, and vanadate-regulated serine/threonine protein kinase activity that is not related to two other cofactor-independent, growth-regulated protein kinases, pp70-S6 protein kinase and pp90rsk. The pp90rsk-protein kinase activity (referred to as rsk-kinase) is also not related to cofactor-dependent signal transducing protein kinases such as the cyclic AMP-dependent protein kinases, members of the protein kinase C family, or other Ca2(+)-dependent protein kinases. In vitro, partially purified rsk-kinase phosphorylates several of the sites (serine and threonine) that are phosphorylated in growth-stimulated cultured cells. A detailed examination of the mitogen-regulated activation kinetics of rsk-kinase and pp90rsk activities demonstrated that they are coordinately regulated. In addition, protein kinase C is not absolutely required for epidermal and fibroblast growth factor-stimulated activation of rsk-kinase, whereas, like pp90rsk, platelet-derived growth factor- and vanadate-stimulated rsk-kinase activity exhibits a greater dependence on protein kinase C-mediated signal transduction. The characterization and future purification of the rsk-kinase(s) will improve our understanding of the early signaling events regulating cell growth.

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