Characterization of a cDNA coding for an extracellular calmodulin-binding protein from suspension-cultured cells of Angelica dahurica

Abstract

In order to characterize a specific extracellular 21-kDa calmodulin-binding protein (named: ECBP21) from Angelica dahurica L. suspension-cultured cells, the cDNA coding for the protein has been cloned. Here, Southern blot analysis shows that there are at least two copies of ECBP21 gene in Angelica genome. Using truncated versions of ECBP21 and synthetic peptide in CaM binding assays, we mapped the calmodulin-binding domain to a 16-amino acid stretch (residues 200-215) at the C-terminal region. The ECBP21 was localized in the cell wall area by the immunogold electron microscopy and by GFP labeling method. These results define ECBP21 as a kind of an extracellular calmodulin-binding protein (CaMBP). Furthermore, using Northern blot analysis, we examined the expression dynamics of ecbp21 during the incubation of Angelica suspension-cultured cells and the treatments with some growth regulators. The above studies further provide the molecular evidence for the existence of the gene coding for extracellular CaMBPs and imply a possible role for ECBP21.

Apoplastic Calmodulin Receptor-like Binding Proteins in Suspension-cultured Cells of Arabidopsis thaliana

“…7B), which is consistent with the previous findings of others and ours for ECBP21 in apoplasts of A. dahurica (21,37,45), ecto-apyrase in plant cells (38 -40), and CaMbinding proteins in the body fluids of animals (41). It is becoming clear that cell wall proteins or polypeptides play important roles in regulating plant development and growth, especially in sensing environment signals.…”

Calcium at the Cell Wall‐Cytoplast Interface

“…Taken together they suggest that heterotrimeric G‐proteins participate in the signal transduction pathway that involves extracellular CaM as a regulator of pollen tube growth. A possible downstream target has been identified in suspension‐culture cells of Angelica ; this is a 21 kDa CaM binding protein that is localized in the cell wall (Mao et al 2005).…”

Calmodulin Binds to Extracellular Sites on the Plasma Membrane of Plant Cells and Elicits a Rise in Intracellular Calcium Concentration

“…Although some 10 polypeptide hormones or putative polypeptide signals have been reported in plants, only a few have been well characterized at the genetic and biochemical level (34,35). It has also been proposed that extracellular CaM may serve as a polypeptide signal in plants (5,6,8,9). Experimental confirmation of the existence and localization of such CaM-binding sites on the cell surface was the subject of the present study.…”

“…Functional studies have established that exogenously applied CaM can stimulate the proliferation of suspension-cultured plant cells (6) as well as affect intracellular activities of heterotrimeric G proteins and phospholipases in protoplasts (7,8). Based on these findings, it has been proposed that, in plants, extracellular CaM may function as a signaling agent involved in the regulation of cell growth and development (4).…”

“…Although a number of studies have addressed the molecular mechanism(s) by which extracellular CaM might act as a signal (6,9) and attempts have been made to identify extracellular CaM-binding proteins (4,6), currently there is no direct evidence in support of the hypothesis that specific CaM-binding sites exist at the surface of plant cells.…”