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Abstract: Plants possess multiple genes encoding calcium sensor proteins that are members of the penta-EF-hand (PEF) family. Characterized PEF proteins such as ALG-2 (apoptosis-linked gene 2 product) and the calpain small subunit function in diverse cellular processes in a calcium-dependent manner by interacting with their target proteins at either their N-terminal extension or Ca2+ binding domains. We have identified a previously unreported class of PEF proteins in plants that are notable because they do not possess the hydrophobic amino acid rich N-terminal extension that is typical of these PEF proteins. We demonstrate that the maize PEF protein without the N-terminal extension has the characteristics of known PEF proteins; the protein binds calcium in the 100 nM range and, as a result of calcium binding, displays an increase in hydrophobicity. Characterization of the truncated maize PEF protein provides insights into the role of the N-terminal extension in PEF protein signaling. In the context of the current model of how PEF proteins are activated by calcium binding, these results demonstrate that this distinctive class of PEF proteins could function as calcium sensor proteins in plants even in the absence of the N-terminal extension. [Copyright &y& Elsevier]

Abstract: The phospholipase D (PLD) family of Arabidopsis thaliana has 12 identified members, including three highly homologous PLDγs. The enzymatic and molecular properties of PLDγ2 were characterized and compared with those of PLDγ1. Two variants of PLDγ2 cDNAs, designated PLDγ2a and PLDγ2b, were isolated, and they differ in the presence of a 96-nucleotide fragment at the beginning of the open reading frame. Catalytically active PLDγ2a was expressed in E. coli. PLDγ2a and γ1 both require phosphatidylinositol 4,5-bisphosphate (PIP2) and calcium for activity, but they differ in the effect of PIP2 and Triton X-100 on their activities. While Triton X-100 could greatly activate PLDγ1 activity and served only as a neutral diluent in the substrate vesicles, it totally abolished PLDγ2a activity and prohibited any stimulation effect from PIP2. PLDγ2a misses one of the basic, PIP2-interacting residues, which may weaken the binding of PIP2 to PLDγ2a. In addition, PLDγ2 and PLDγ1 displayed different patterns of expression in different tissues, and the transcript of PLDγ2a differs from that of PLDγ1 by having a longer 5′-UTR. These differences in biochemical and molecular properties suggest that the highly homologous PLDγs are subjected to unique regulations and might have distinguishable functions. [Copyright &y& Elsevier]