A Ca²⁺/Calmodulin-Binding Peroxidase from Euphorbia Latex:  Novel Aspects of Calcium−Hydrogen Peroxide Cross-Talk in the Regulation of Plant Defenses^,

Abstract

Calmodulin (CaM) is a ubiquitous Ca(2+) sensor found in all eukaryotes, where it participates in the regulation of diverse calcium-dependent physiological processes. In response to fluctuations of the intracellular concentration of Ca(2+), CaM binds to a set of unrelated target proteins and modulates their activity. In plants, a growing number of CaM-binding proteins have been identified that apparently do not have a counterpart in animals. Some of these plant-specific Ca(2+)/CaM-activated proteins are known to tune the interaction between calcium and H(2)O(2) in orchestrating plant defenses against biotic and abiotic stresses. We previously characterized a calcium-dependent peroxidase isolated from the latex of the Mediterranean shrub Euphorbia characias (ELP) [Medda et al. (2003) Biochemistry 42, 8909-8918]. Here we report the cDNA nucleotide sequence of Euphorbia latex peroxidase, showing that the derived protein has two distinct amino acid sequences recognized as CaM-binding sites. The cDNA encoding for an E. characias CaM was also found and sequenced, and its protein product was detected in the latex. Results obtained from different CaM-binding assays and the determination of steady-state parameters showed unequivocally that ELP is a CaM-binding protein activated by the Ca(2+)/CaM system. To the best of our knowledge, this is the first example of a peroxidase regulated by this classic signal transduction mechanism. These findings suggest that peroxidase might be another node in the Ca(2+)/H(2)O(2)-mediated plant defense system, having both positive and negative effects in regulating H(2)O(2) homeostasis.

Specific functions of individual class III peroxidase genes

“…Among plant Ca 2+ -sensing proteins, calmodulin (CaM) is increasingly appreciated as a critical player, but identifying its downstream target proteins and understanding their functions is still a work in progress. In this study, the authors isolated and sequenced the ELP cDNA and showed that the mature protein harbours two distinct CaM-binding sites (Mura et al, 2005). ELP-CaM interaction was also determined experimentally and by computer-assisted prediction methods.…”

Calcium promotes activity and confers heat stability on plant peroxidases

“…Calcium is essential for horseraddish peroxidase (HRP) activity 16,17 and other peroxidases have been shown to bind Ca 2+ ions. 14,18 Some peroxidases even have a calmodulin binding domain 14,19 and calcium may switch a peroxidase between different modes of action. 20 In addition, apoplastic peroxidases are anchored in the cell wall matrix by calcium-dependent binding to pectate (i.e., polygalacturonic acid).…”

“…Since these peroxidases are dependent on calcium ions as well as on H 2 O 2 , they may function as indispensable nodes of a signaling network which employs these two known signaling molecules for stress defense. 19,30 In this study we have extracted peroxidases from a range of different plant sources and investigated their heat stability and dependency on calcium using a previously established in vitro assay. 31 We demonstrate that enzymes of similar function in plant species from natural habitats with varying maximum day temperatures exhibit different thermostability and different calcium responsiveness.…”

A probable crosstalk between Ca+2, reactive oxygen species accumulation and scavenging mechanisms and modulation of protein kinase C activity during seed development in sunflower

“…[19][20][21] Protein kinase C activity exhibits development-dependent modulation during seed maturation in sunflower. This has been observed both during early stages of embryo development and during the development of seeds by the use of PKC specific fluorescent probe Fim-1, a membrane permeant derivative of bisindoylmaleamide, as well as PKC inhibitors, namely staurosporine and bisindoylmaleamide.…”