Calcium‐dependent protein kinase from maize seedlings activated by phospholipids

Abstract

A calcium-and phospholipid-dependent protein kinase of apparent molecular mass 54 kDa (designated ZmCPKp54) was partially purified from etiolated maize seedlings. Activity of ZmCPKp54 is stimulated by phosphatidylserine and phosphatidylinositol, but is not essentially affected by diolein and phorbol esters. The enzyme cross-reacts with polyclonal antibodies against the calmodulin like-domain of the calcium-dependent protein kinase, but not with antibodies against catalytic or regulatory domains of protein kinase C. ZmCPKp54 is not able to phosphorylate the specific substrates of protein kinase C (MARCKS peptide and protein kinase C substrate peptide derived from pseudosubstrate sequence) and its activity is not inhibited by specific PKC inhibitors (bisindolylmaleimide, protein kinase C pseudosubstrate inhibitory peptide). The substrate specificity and sensitivity to the inhibitors of the maize enzyme resembles calcium-dependent protein kinase. The biochemical and immunological properties indicate that ZmCPKp54 belongs to the calcium-dependent protein kinase family.Keywords: plant protein kinase; protein kinase specificity; calcium, phosphatidylserine; phosphatidylinositol. In addition, the rapid turnover of phosphatidylinositol (PtdIns) metabolites has been demonstrated in a variety of plant signal responses (reviewed in [2]). The phosphatidylinositol pathway involves stimulation of a phospholipase C and release of two messengers: diacylglycerol and inositol(1,4,5)-trisphosphate. Diacylglycerol and inositol(1,4,5)trisphosphate are widespread in plant cells, and the levels of diacylglycerol and inositol(1,4,5)trisphosphate are changed during leaf movement [3], when the plants are exposed to the light [4] or during osmotic stress [5,6]. In the animal kingdom the activity of protein kinase C (PKC) is dependent on calcium and phospholipids. It is involved in sensing the cellular changes of diacylglycerol and calcium ion concentrations, transducing the signal from the cell membrane to the other signalling components, and finally to the cell nucleus (reviewed in [7]).The presence of elements required for inositol phospholipidbased signalling in plant cells has prompted investigators to look for a calcium-and phospholipid-dependent protein kinase resembling mammalian PKC. The presence of a Ca 12 -and lipid-dependent kinase activity in plants was first shown in cytosolic fraction of zucchini (courgettes) [8] Recently, a 70-kDa protein kinase possessing activity that is stimulated by diacylglycerol or phorbol esters in the presence of calcium was purified to homogenity from etiolated maize leaves [16]. Moreover, a functional homolog of mammalian PKC was identified in potato as a key element in the elicitorinduced defence [18]. The above biochemical and immunological data suggest the existence of PKC counterpart in plants.A large family of calcium-dependent protein kinases (CDPKs), also called calmodulin-like domain protein kinases, is present in plants. These enzymes control many physiological responses and are involved ...

Molecular aspects of the early stages of elicitation of secondary metabolites in plants

“…Although the use of these synthetic compounds could be questioned since they have been scarcely tested in plants and in general are considered specific for animals, due to the structural differences between PKC and CDPK and in cofactor requirements, their use allow to distinguish between both enzymes. For example, calphostin C, which interacts with the PKC regulatory domain by competing at the binding site of diacylglycerol (DAG) and phorbol esters, is devoid of inhibitory activity on CDPK as it does not require DAG for its activation [114,115]. Likewise, it has been shown that diolein and PMA are unable to affect the activity of the CDPK ZmCPKp54.…”

Evolutionary and functional study of the CDPK gene family in wheat (Triticum aestivum L.)

“…Activated CDPKs participate in different signal transduction pathways through phosphorylation of specific substrates (Zhang and Chollet 1997;Hernandez et al 2004). In addition to calcium, CDPKs are also regulated by phospholipids (Szczegielniak et al 2000(Szczegielniak et al , 2005 and 14-3-3 proteins (Camoni et al 1998;Cheng et al 2002).…”

StCDPK2 expression and activity reveal a highly responsive potato calcium-dependent protein kinase involved in light signalling

“…The activity of CDPKs from oat (Schaller et al 1992), Arabidopsis thaliana (Harper et al 1993), and maize (Szczegielniak et al 2000) and of a recombinant CDPK from Daucus carota (Farmer and Choi 1999) can be stimulated by different lipids (phosphatidylinositol, lysophosphatidylcholine, phosphatidylserine, and/or phosphatidic acid (PA), suggesting that membrane association is a factor that contributes to CDPK activity.…”