Regulation of the exocytotic machinery by cAMP-dependent protein kinase: implications for presynaptic plasticity

2003 ◽  
Vol 31 (4) ◽  
pp. 824-827 ◽  
Author(s):  
G.J.O. Evans ◽  
A. Morgan
Keyword(s):  
Protein Kinase ◽  
Protein Interactions ◽  
Cell Types ◽  
Protein Protein Interactions ◽  
Dependent Protein Kinase ◽  
Camp Dependent Protein Kinase ◽  
Cysteine String Protein ◽  
Presynaptic Plasticity ◽  
Dependent Protein

For over a decade, the enhancement of regulated exocytosis by cAMP-dependent protein kinase (PKA) has remained unexplained at the molecular level. The fact that this phenomenon has been observed in such a wide variety of secretory cell types, from pancreatic β-cells to neurons, suggests that it is an important and fundamental mechanism. Extensive analysis of the phosphorylation of exocytotic proteins has yielded few substrates of PKA in vitro, and fewer still have had physiological effects attributed to their phosphorylation. Here we review two proteins that do fulfil these criteria: the synaptic vesicle proteins cysteine string protein (CSP) and Snapin. Phosphorylation of these proteins by PKA produces changes in their respective protein–protein interactions, and has been attributed to modulation of the vesicle priming (Snapin) and vesicle fusion (CSP) stages of exocytosis. We also discuss how the function of CSP and Snapin phosphorylation might fit into an interesting aspect of the PKA-dependent enhancement of exocytosis: presynaptic plasticity in the brain.

Subcellular Localization of the Type II cAMP-Dependent Protein Kinase

Physiology ◽  
1992 ◽  
Vol 7 (4) ◽  
pp. 143-148 ◽  
Author(s):  
JD Scott ◽  
DW Carr
Keyword(s):  
Protein Kinase ◽  
Protein Interactions ◽  
Regulatory Subunit ◽  
Type Ii ◽  
Protein Protein Interactions ◽  
Dependent Protein Kinase ◽  
Biochemical Effects ◽  
Camp Dependent Protein Kinase ◽  
Anchoring Proteins ◽  
Dependent Protein

Diverse biochemical effects of different neurotransmitters or hormones that stimulate cAMP production may occur through activation of compartmentalized pools of cAMP-dependent protein kinase (PKA). Evidence suggests that compartmentalization of type II PKA is maintained through protein-protein interactions between the regulatory subunit and specific anchoring proteins.

scholarly journals High-resolution crystal structure of cAMP-dependent protein kinase fromCricetulus griseus

2015 ◽  
Vol 71 (8) ◽  
pp. 1088-1093 ◽  
Author(s):  
Denis Kudlinzki ◽  
Verena L. Linhard ◽  
Krishna Saxena ◽  
Sridhar Sreeramulu ◽  
Santosh Gande ◽  
...  
Keyword(s):  
Protein Kinase ◽  
High Resolution ◽  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Dependent Protein Kinase ◽  
Cellular Processes ◽  
Camp Dependent Protein Kinase ◽  
Dependent Protein ◽  
Functional Regulation ◽  
Very High

Protein kinases (PKs) are dynamic regulators of numerous cellular processes. Their phosphorylation activity is determined by the conserved kinase core structure, which is maintained by the interaction and dynamics with associated domains or interacting proteins. The prototype enzyme for investigations to understand the activity and regulation of PKs is the catalytic subunit of cAMP-dependent protein kinase (PKAc). Major effects of functional regulation and ligand binding are driven by only minor structural modulations in protein–protein interactions. In order to resolve such minor structural differences, very high resolution structures are required. Here, the high-resolution X-ray structure of PKAc fromCricetulus griseusis reported.

scholarly journals Regulation of Microtubule Dynamics by Extracellular Signals: cAMP-dependent Protein Kinase Switches Off the Activity of Oncoprotein 18 in Intact Cells

1998 ◽  
Vol 140 (1) ◽  
pp. 131-141 ◽  
Author(s):  
Helena Melander Gradin ◽  
Niklas Larsson ◽  
Ulrica Marklund ◽  
Martin Gullberg
Keyword(s):  
Protein Kinase ◽  
Genetic System ◽  
Cell Surface Receptor ◽  
In Vitro Assays ◽  
Dependent Protein Kinase ◽  
Intact Cells ◽  
Camp Dependent Protein Kinase ◽  
Oncoprotein 18 ◽  
Dependent Protein

Oncoprotein 18 (Op18, also termed p19, 19K, metablastin, stathmin, and prosolin) is a recently identified regulator of microtubule (MT) dynamics. Op18 is a target for both cell cycle and cell surface receptor-coupled kinase systems, and phosphorylation of Op18 on specific combinations of sites has been shown to switch off its MT-destabilizing activity. Here we show that induced expression of the catalytic subunit of cAMP-dependent protein kinase (PKA) results in a dramatic increase in cellular MT polymer content concomitant with phosphorylation and partial degradation of Op18. That PKA may regulate the MT system by downregulation of Op18 activity was evaluated by a genetic system allowing conditional co-expression of PKA and a series of kinase target site–deficient mutants of Op18. The results show that phosphorylation of Op18 on two specific sites, Ser-16 and Ser-63, is necessary and sufficient for PKA to switch off Op18 activity in intact cells. The regulatory importance of dual phosphorylation on Ser-16 and Ser-63 of Op18 was reproduced by in vitro assays. These results suggest a simple model where PKA phosphorylation downregulates the MT-destabilizing activity of Op18, which in turn promotes increased tubulin polymerization. Hence, the present study shows that Op18 has the potential to regulate the MT system in response to external signals such as cAMP-linked agonists.

scholarly journals Adrenocorticotrophic hormone stimulates phosphotyrosine phosphatase SHP2 in bovine adrenocortical cells: phosphorylation and activation by cAMP-dependent protein kinase

2000 ◽  
Vol 352 (2) ◽  
pp. 483-490 ◽  
Author(s):  
Stéphane ROCCHI ◽  
Isabelle GAILLARD ◽  
Emmanuel VAN OBBERGHEN ◽  
Edmond M. CHAMBAZ ◽  
Isabelle VILGRAIN
Keyword(s):  
Protein Kinase ◽  
Kinase Assay ◽  
Dependent Manner ◽  
Adrenocorticotrophic Hormone ◽  
Adrenocortical Cells ◽  
Dependent Protein Kinase ◽  
Phosphotyrosine Phosphatase ◽  
Camp Dependent Protein Kinase ◽  
Dependent Protein

During activation of adrenocortical cells by adrenocorticotrophic hormone (ACTH), tyrosine dephosphorylation of paxillin is stimulated and this correlates with protrusion of filopodial structures and a decreased number of focal adhesions. These effects are inhibited by Na3VO4, a phosphotyrosine phosphatase inhibitor [Vilgrain, Chinn, Gaillard, Chambaz and Feige (1998) Biochem. J. 332, 533–540]. However, the tyrosine phosphatases involved in these processes remain to be identified. In this study, we provide evidence that the Src homology domain (SH)2-containing phosphotyrosine phosphatase (SHP)2, but not SHP1, is expressed in adrenocortical cells and is phosphorylated upon ACTH challenge. ACTH (10-8M) treatment of 32P-labelled adrenocortical cells resulted in an increase in phosphorylated SHP2. By probing SHP2-containing immunoprecipitates with an antibody to phosphoserine we found that SHP2 was phosphorylated on serine in ACTH-treated cells in a dose- and time-dependent manner. Furthermore, using an in vitro kinase assay, we showed that SHP2 was a target for cAMP-dependent protein kinase (PKA). Serine was identified as the only target amino acid phosphorylated in SHP2. Phosphorylation of SHP2 by PKA resulted in a dramatic stimulation of phosphatase activity measured either with insulin receptor substrate-1 or with the synthetic peptide [32P]poly(Glu/Tyr) as substrate. In an in-gel assay of SHP2-containing immunoprecipitates, phosphorylated in vitro by PKA or isolated from adrenocortical cells treated with 10nM ACTH, a pronounced activation of SHP2 activity was shown. These observations clearly support the idea that a PKA-mediated signal transduction pathway contributes to SHP2 regulation in adrenocortical cells and point to SHP2 as a possible mediator of the effects of ACTH.

scholarly journals cAMP-dependent protein kinase (PKA) complexes probed by complementary differential scanning fluorimetry and ion mobility–mass spectrometry

2016 ◽  
Vol 473 (19) ◽  
pp. 3159-3175 ◽  
Author(s):  
Dominic P. Byrne ◽  
Matthias Vonderach ◽  
Samantha Ferries ◽  
Philip J. Brownridge ◽  
Claire E. Eyers ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Kinase ◽  
Ion Mobility ◽  
Catalytic Domain ◽  
Ion Mobility Mass Spectrometry ◽  
Dependent Protein Kinase ◽  
Camp Dependent Protein Kinase ◽  
Differential Scanning Fluorimetry ◽  
Dependent Protein

cAMP-dependent protein kinase (PKA) is an archetypal biological signaling module and a model for understanding the regulation of protein kinases. In the present study, we combine biochemistry with differential scanning fluorimetry (DSF) and ion mobility–mass spectrometry (IM–MS) to evaluate effects of phosphorylation and structure on the ligand binding, dynamics and stability of components of heteromeric PKA protein complexes in vitro. We uncover dynamic, conformationally distinct populations of the PKA catalytic subunit with distinct structural stability and susceptibility to the physiological protein inhibitor PKI. Native MS of reconstituted PKA R2C2 holoenzymes reveals variable subunit stoichiometry and holoenzyme ablation by PKI binding. Finally, we find that although a ‘kinase-dead’ PKA catalytic domain cannot bind to ATP in solution, it interacts with several prominent chemical kinase inhibitors. These data demonstrate the combined power of IM–MS and DSF to probe PKA dynamics and regulation, techniques that can be employed to evaluate other protein-ligand complexes, with broad implications for cellular signaling.

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