Rapid Cycling of Autophosphorylation of a Ca<sup>2+</sup>-Calmodulin Regulated Plasma Membrane Located Protein Kinase from Pea

Blowers, David P.; Trewavas, Anthony

doi:10.1104/pp.90.4.1279

Cited by 21 publications

(5 citation statements)

References 18 publications

(27 reference statements)

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“…When purified pea plasma membrane was incubated in vitro with [32P]ATP, we observed one protein (in this example with a molecular mass of 18 kD) to complete its phosphorylation within 15 s, whereas other proteins required 5 to 10 min before phosphorylation was apparently completed. We were able to demonstrate subsequently (Blowers and Trewavas, 1989) that this very rapid phosphorylation was the result of autophosphorylation of a protein kinase. The 70-and the 60-kD peptides may simply be protein kinases that autophosphorylate in a Ca2+-dependent manner, a possibility we are testing at present (Grimm et al, 1989).…”

Section: Discussionmentioning

confidence: 80%

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

1993

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Etiolated wheat (Triticum aesfivum cv Mercia) leaf protoplasts respond to brief red-light irradiation by increasing in volume over a 10-min incubation period (M.E. Bossen, H.A. Dassen, R. E. Kendrick, W.J. Vredenberg [1988] Planta 174: 94-100). When the calcium-sensitive dye Fluo-3 was incorporated into these protoplasts, red-light irradiation initiated calcium transients lasting about 2 min (P.S. Shacklock, N.D. Read, A.J. Trewavas [1992] Nature 358 153-155). Release of calcium in the protoplasts by photolysis of incorporated 1 -(2-amino-5-[1 -hydroxy-l-(2-nitro-4, 5-methylenedioxyphenyl)-methyl]-phenoxy}-2-(2'-amino-5'-methy1phenoxy)-ethane-N,N,N',N'-tetraacetic acid, tetrasodium salt (caged calcium) or caged inositol trisphosphate frequently induced transient increases in intracellular calcium levels, although the kinetics of these changes showed variation between experiments. Upon exposure to red light, a pronounced increase in the phosphorylation of a 70-kD and to a lesser extent a 60-kD peptide was observed, commencing within 15 s and continuing for up to 2 min. Simultaneous far-red and red irradiation attenuated the response. Upon release of incorporated caged calcium by cage photolysis, the labeling of these two peptides was greatly increased. When incorporated caged inositol trisphosphate was photolyzed, only the labeling of the 70-kD peptide was enhanced. Phosphorylation of the 70-kD peptide was also increased when extracellular calcium was elevated, but it decreased with increasing extracellular ECTA. These data thus provide direct evidence for the operation of an in vivo transduction sequence involving red light-dependent, calcium-sensitive protein phosphorylation.Light initiates a diverse range of phenomena in plants (Smith, 19 75). Processes specifically controlled by R include the regulation of growth, reproduction, and time measurement. The effects of R are mediated by the family of photoreceptors designated phytochrome, and the biochemical mechanism of action of R has been of intense interest for many years (Smith, 1975; Kendrick and Kronenberg, 1986). A current hypothesis involves the mediation of R-induced changes in [Ca2+lC and was first introduced by Haupt and Weisenseel (1976). These authors pointed to the evidence that phytochrome regulated the passage of ions across the plasma membrane and noted that Caz+ had a key role in cell signaling in animal cells. With the detection of calmodulin in plant cells (Anderson and Cormier, 1978) and the discovery by Hetherington and Trewavas (1982) of Ca2+-regulated protein kinases in plants, Roux (1984) and Roux et al. (1986) ' Supported by the Science and Engineering Research Council.

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Section: Discussionmentioning

confidence: 80%

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

1993

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“…It is possible that they are the result of protein kinase activity in these membrane fractions. Rapidly tuming-over kinases have been demonstrated in other plant tissues (Blowers and Trewavas, 1989;Garbarino et al, 1991).…”

Section: Discussionmentioning

confidence: 99%

Investigation of the Calcium-Transporting ATPases at the Endoplasmic Reticulum and Plasma Membrane of Red Beet (Beta vulgaris)

et al. 1993

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Calcium-transporting ATPases were compared in endoplasmic reticulum (ER)-and plasma membrane-enriched fractions of red beet (Beta vulgaris 1.) storage tissue by measuring "Ca uptake and calcium-dependent phosphoenzyme formation. l h e plasma membrane fraction was prepared by aqueous two-phase partitioning of a microsomal fraction and collecting the upper phase. l h e ERenriched fraction was obtained by submitting a sucrose-gradient ER-enriched fradion to aqueous two-phase partitioning and collecting the lower phase; this reduced contaminating plasma membrane, which partitioned into the upper phase. l h e ATP-dependent calcium uptake observed in both fractions was released by the calcium ionophore A23187. Calcium uptake showed saturation kinetics for calcium with K, values of 0.92 mmol m-3 for the ER fraction and 1.24 mmol m-3 for the plasma membrane fraction. Uptake into both fractions was inhibited by vanadate and erythrosin 6, although the plasma membrane system was slightly more sensitive to both inhibitors. Cyclopiazonic acid and thapsigargin, at low concentrations, had no marked effect on uptake. The plasma membrane system was less substrate-specific for ATP than the ER system, since it was able to use CTP and ITP to drive calcium transport at up to 50% of the leve1 obtained with ATP. Following phosphorylation with [-p3'P]ATP, two high molecular mass, calcium-dependent phosphoproteins (119 and 124 kD) and a low molecular mass, calcium-independent phosphoprotein (1 7 kD) were observed in the plasma membrane fraction. The ER fraction showed one high molecular mass phosphoprotein (119 kD) in the presence of calcium and two low molecular mass phosphoproteins (17 and 20 kD) that showed no calcium dependence. l h e low molecular mass phosphoproteins were insensitive to hydroxylamine, but they did show turnover. l h e identity of these proteins is unknown, but they do not have the properties of phosphorylated intermediates of calcium-ATPases. In contrast, the high molecular mass phosphoproteins displayed properties consistent with their representing phosphorylated intermediates of E,Ez-type ATPases; they were hydroxylamine-sensitive, showed rapid turnover, and were inhibited by vanadate. Because they showed calcium-dependent phosphorylation and were sensitive to erythrosin B, the 119-and 124-kD phosphoproteins may be phosphorylated intermediates of the ER and plasma membrane calcium ATPases. These phosphoproteins were characterized further with respect to inhibitor sensitivity, responses to ions, and substrate specificity.

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“…A possible regulatory function remains conceivable given the abundant expression of NDPK/Nm23 in many tumor tissues [39, 401. Further evidence for the existence of a Nm23 protein phosphotransferase activity in the cell is provided by several reports on NDPK from plants, which all show a high degree of sequence similarity to human NDPK 1411. It was reported that NDPK may act as a kind of active subunit of protein kinase complexes in pea [42,43] and sugar-cane [44]. The NDPK preparations apparently exhibited a weak serine phosphotransferase activity toward membrane proteins, which could be stimulated by Ca' ' and calmodulin.…”

Section: Discussionmentioning

confidence: 99%

A Novel Serine/Threonine‐Specific Protein Phosphotransferase Activity of Nm23/Nucleoside‐Diphosphate Kinase

Engel

Véron

Theisinger

et al. 1995

European Journal of Biochemistry

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Two human nm23 genes have been identified, designated nm23-HI and nm23-H2, which encode the 88 % identical nucleoside-diphosphate kinase (NDPK) A and NDPK B polypeptides, respectively. The nm23-HI gene product has been shown to play a functional role in the suppression of tumor metastasis. The Nm23 proteins/NDPK are highly conserved throughout evolution and are implicated in controling cellular differentiation and development in various species, while the underlying mechanisms remain undefined. Neither the NDPK activity nor the DNA-binding activity, identified recently for NDPK B, can satisfactory explain the regulatory functions of Nm23. The present study provides evidence that purified Nm23 proteins are capable of transferring a phosphate group to other proteins when non-denaturing amounts of urea are present. This novel Nm23/NDPK activity was found to be specific for serine and threonine residues, and the transphosphorylation of substrate proteins occurred stoichiometrically. Because of the absence of a substrate turn-over, the novel function was termed protein phosphotransferase activity instead of protein kinase activity. It is demonstrated that urea stimulates the interaction of NDPK with other proteins. Identical phosphoprotein patterns were obtained using purified NDPK preparations from human, Drosophila, yeast and Dictyostelium in the presence of urea. Partially purified NDPK from human erythrocytes produced a similar phosphorylation pattern independent of urea addition and also acted stoichiometrically. In this preparation, a protein phosphotransferase activity of Nm23 species may possibly be generated andor stabilized by the interaction with copurified proteins. Using different mutants of Dictyostelium NDPK it was shown that the protein phosphotransferase activity depends on the same active site as the NDPK activity. A phosphotransfer mechanism analogous to that of protein-histidine kinases is proposed, involving a high-energy phosphohistidine intermediate. Furthermore, the novel Nm23 function is compared with an apparently similar protein phosphotransferase activity which was observed previously with partially purified NDPK from different plant species.Keywords: nucleoside-diphosphate kinase; Nm23 ; protein-histidine kinase ; serine phosphorylation Nucleoside-diphosphate kinase (NDPK) is an ubiquitous enzyme that catalyzes the transfer of the y-phosphate group from a nucleoside triphosphate to a nucleoside diphosphate involving a high-energy phosphoenzyme intermediate according to the following scheme [ I ] : N,TP + E * N,DP + E -P polypeptide chains, A and B, which are 88% identical. NDPK A is encoded by the putative tumor-metastasis-suppressor gene, nm23-HI, and NDPK B by the later-discovered nm23-H2 gene [2, 31. Expression of the murine homologue of nm23-HI has been reduced in highly metastatic tumor cell lines compared with cell lines of lower metastatic potential [4-61. Subsequently, the proposed metastasis suppressor function has been confirmed by transfection of nm23 cDNA into highly metasta...

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Rapid Cycling of Autophosphorylation of a Ca²⁺-Calmodulin Regulated Plasma Membrane Located Protein Kinase from Pea

Cited by 21 publications

References 18 publications

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

Investigation of the Calcium-Transporting ATPases at the Endoplasmic Reticulum and Plasma Membrane of Red Beet (Beta vulgaris)

A Novel Serine/Threonine‐Specific Protein Phosphotransferase Activity of Nm23/Nucleoside‐Diphosphate Kinase

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Rapid Cycling of Autophosphorylation of a Ca2+-Calmodulin Regulated Plasma Membrane Located Protein Kinase from Pea

Cited by 21 publications

References 18 publications

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

Detection in Vivo of Very Rapid Red Light-Induced Calcium-Sensitive Protein Phosphorylation in Etiolated Wheat (Triticum aestivum) Leaf Protoplasts

Investigation of the Calcium-Transporting ATPases at the Endoplasmic Reticulum and Plasma Membrane of Red Beet (Beta vulgaris)

A Novel Serine/Threonine‐Specific Protein Phosphotransferase Activity of Nm23/Nucleoside‐Diphosphate Kinase

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Rapid Cycling of Autophosphorylation of a Ca²⁺-Calmodulin Regulated Plasma Membrane Located Protein Kinase from Pea