Isolation of plasma-membrane-bound calcium/calmodulin-regulated protein kinase from pea using Western blotting

Planta ◽  
1985 ◽  
Vol 166 (2) ◽  
pp. 208-215 ◽  
Author(s):  
D. P. Blowers ◽  
A. Hetherington ◽  
A. Trewavas
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Western Blotting ◽  
Membrane Bound

scholarly journals Lipid-dependent Akt-ivity: where, when, and how

2019 ◽  
Vol 47 (3) ◽  
pp. 897-908 ◽  
Author(s):  
Katharina M. Siess ◽  
Thomas A. Leonard
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Kinase Domain ◽  
Target Of Rapamycin ◽  
Endomembrane System ◽  
Essential Protein ◽  
Mechanistic Target Of Rapamycin ◽  
Subcellular Compartments ◽  
Membrane Bound ◽  
Phosphoinositide 3 Kinase

Abstract Akt is an essential protein kinase activated downstream of phosphoinositide 3-kinase and frequently hyperactivated in cancer. Canonically, Akt is activated by phosphoinositide-dependent kinase 1 and mechanistic target of rapamycin complex 2, which phosphorylate it on two regulatory residues in its kinase domain upon targeting of Akt to the plasma membrane by PI(3,4,5)P3. Recent evidence, however, has shown that, in addition to phosphorylation, Akt activity is allosterically coupled to the engagement of PI(3,4,5)P3 or PI(3,4)P2 in cellular membranes. Furthermore, the active membrane-bound conformation of Akt is protected from dephosphorylation, and Akt inactivation by phosphatases is rate-limited by its dissociation. Thus, Akt activity is restricted to membranes containing either PI(3,4,5)P3 or PI(3,4)P2. While PI(3,4,5)P3 has long been associated with signaling at the plasma membrane, PI(3,4)P2 is gaining increasing traction as a signaling lipid and has been implicated in controlling Akt activity throughout the endomembrane system. This has clear implications for the phosphorylation of both freely diffusible substrates and those localized to discrete subcellular compartments.

A 67-kDa plasma-membrane-bound Ca 2+ -stimulated protein kinase active in sink tissue of higher plants

Planta ◽  
1998 ◽  
Vol 205 (2) ◽  
pp. 197-204 ◽  
Author(s):  
Laurence D. P. Barker ◽  
Matthew D. Templeton ◽  
Ian B. Ferguson
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Higher Plants ◽  
Membrane Bound ◽  
Sink Tissue

scholarly journals The plant calcium-dependent protein kinase CPK3 phosphorylates REM1.3 to restrict viral infection

2017 ◽  
Author(s):  
Artemis Perraki ◽  
Julien Gronnier ◽  
Paul Gouguet ◽  
Marie Boudsocq ◽  
Anne-Flore Deroubaix ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Cell Movement ◽  
Dependent Protein Kinase ◽  
Calcium Dependent ◽  
Intrinsically Disordered ◽  
Dependent Protein ◽  
Membrane Bound ◽  
Calcium Dependent Protein Kinase ◽  
Group 1

AbstractPlants respond to pathogens through dynamic regulation of plasma membrane-bound signaling pathways. To date, how the plant plasma membrane is involved in responses to viruses is mostly unknown. Here, we show that plant cells sense the Potato virus X (PVX) COAT PROTEIN and TRIPLE GENE BLOCK 1 proteins and subsequently trigger the activation of a membrane-bound calcium-dependent kinase. We show that the Arabidopsis thaliana CALCIUM-DEPENDENT PROTEIN KINASE 3-interacts with group 1 REMORINs in vivo, phosphorylates the intrinsically disordered N-terminal domain of the Group 1 REMORIN REM1.3, and restricts PVX cell-to-cell movement. REM1.3-s phospho-status defines its plasma membrane nanodomain organization and is crucial for REM1.3-dependent restriction of PVX cell-to-cell movement by regulation of callose deposition at plasmodesmata. This study unveils plasma membrane nanodomain-associated molecular events underlying the plant immune response to viruses.

Dematin Is Essential for Calcium Mobilization in Platelets

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 365-365
Author(s):  
Adam Joseph Wieschhaus ◽  
Guy Le Breton ◽  
Athar H Chishti
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Platelet Activation ◽  
Western Blotting ◽  
Conflicts Of Interest ◽  
Calcium Mobilization ◽  
Cytoskeletal Proteins ◽  
Actin Binding ◽  
Erythrocyte Membranes ◽  
Junctional Complex

Abstract Abstract 365 Dematin is an actin binding protein that was originally identified in the human erythrocyte membranes; however, dematin polypeptides are detectable in many non-erythroid cells. In erythrocytes, dematin is located at the spectrin-actin junctions thus linking the skeleton to the plasma membrane. This junctional complex serves to maintain both erythrocyte shape and membrane stability. Dematin is phosphorylated by multiple protein kinases, including the cAMP-dependent protein kinase and protein kinase C, and its actin bundling activity is regulated by phosphorylation. Dematin is also an excellent substrate of calpain-1, a calcium-dependent cysteine protease. Because of the functional similarities between the core components of the membrane skeleton in erythrocytes and platelets, we sought to investigate the physiological function of dematin in platelets. The remodeling of the actin cytoskeleton is known to regulate platelet activation and secretion of platelet granule contents. Western blotting demonstrated abundant expression of the ∼52 kDa polypeptide of dematin in both human and mouse platelets. To evaluate the functional role of dematin in platelets, we utilized the dematin headpiece knockout (HPKO) mouse model previously generated in our laboratory. Headpiece domain of dematin is an actin binding module sharing sequence similarity with the villin-family of cytoskeletal proteins. Importantly, the in vivo deletion of the headpiece domain of dematin resulted in substantial diminution of calcium mobilization in response to multiple agonists of platelet activation (Fig. 1). The reduced calcium mobilization in HPKO platelets was associated with significant inhibition of the platelet aggregation and granule secretion pathways. The HPKO platelets exhibit decreased activation of both the integrin αIIbβ3 receptor and RhoA upon platelet stimulation. Moreover, the HPKO platelets display aberrant morphology upon spreading on the fibrinogen and vWF-coated surfaces. Consistent with these findings, the HPKO mice show a significant clot retraction defect associated with a general tail bleeding phenotype. Mechanistically, the basal level of cAMP remained unaltered in the HPKO platelets suggesting the independence of the observed phenotype upon changes in the cAMP concentration. Immunofluorescence analysis indicated that dematin is associated with two platelet membrane compartments known to be involved in calcium fluxes, i.e., the dense tubular system (DTS) and the plasma membrane. Studies are currently in progress to identify the dematin-associated complex in platelets by Western blotting and proteomics approaches. Together, these results unveil dematin as a novel regulator of calcium homeostasis in platelets with functional implications for the development of new anti-platelet therapies. Disclosures: No relevant conflicts of interest to declare.

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