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Glutamate decarboxylase (GAD) catalyzes the decarboxylation of glutamate to CO2 and gamma‐aminobutyrate (GABA). GAD is ubiquitous in prokaryotes and eukaryotes, but only plant GAD has been shown to bind calmodulin (CaM). Here, we assess the role of the GAD CaM‐binding domain in vivo. Transgenic tobacco plants expressing a mutant petunia GAD lacking the CaM‐binding domain (GADdeltaC plants) exhibit severe morphological abnormalities, such as short stems, in which cortex parenchyma cells fail to elongate, associated with extremely high GABA and low glutamate levels. The morphology of transgenic plants expressing the full‐length GAD (GAD plants) is indistinguishable from that of wild‐type (WT) plants. In WT and GAD plant extracts, GAD activity is inhibited by EGTA and by the CaM antagonist trifluoperazine, and is associated with a CaM‐containing protein complex of approximately 500 kDa. In contrast, GADdeltaC plants lack normal GAD complexes, and GAD activity in their extracts is not affected by EGTA and trifluoperazine. We conclude that CaM binding to GAD is essential for the regulation of GABA and glutamate metabolism, and that regulation of GAD activity is necessary for normal plant development. This study is the first to demonstrate an in vivo function for CaM binding to a target protein in plants.

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Pathogenesis-related (PR) proteins form a heterogeneous group of host-encoded, low-molecular-mass proteins that are secreted through the exocytic pathway. They are synthesized by the plant in response to various stimuli, including pathogen attack or exposure to certain chemicals. The PRB-1b gene of Nicotiana tabacum codes for a basic-type PR-1 protein whose transcription is regulated by ethylene. A minimal ethylene-responsive promoter element was defined by deletion analysis in transgenic tobacco plants. Promoter sequences containing 213 bp or more were sufficient to enhance a 20-fold increase of beta-glucuronidase reporter gene expression in transgenic tobacco leaves exposed to 20 microliters l-1 of ethylene, while 67 bp were not sufficient to trigger ethylene responsiveness. All the constructs that retained ethylene inducibility exhibited phloem-specific activity, which was constitutive in petiole and pedicel abscission zones. This functional study was correlated to an in vitro screening of the major nuclear proteins' binding sites present on the promoter. Gel-shift analysis using nuclear extracts from ethylene-treated and non-treated plants revealed five sequence-specific protein-DNA complexes on promoter sequences spanning -863 to -142 bp. Constitutive expression of the basic-type PR-1 genes at the leaf and petiole or flower and pedicel interfaces may represent pre-emption of plant defenses against potential pathogens, suggesting a functional similarity to pathogen-induced expression in the leaf.

Abstract. In addition to their well-documented beneficial effects on plant physiological processes, anthocyanins have also been proposed to function in a diverse array of plant/animal interactions. These include the attraction of pollinators and frugivores, as well as the repellence of herbivores and parasites. The optical properties of anthocyanins may serve as visual signals to potential herbivores, indicating a strong metabolic investment in toxic or unpalatable chemicals. Anthocyanins have also been implicated in the camouflage of plant parts against their backgrounds, in the undermining of insect crypsis, and in the mimicry of defensive structures. These hypotheses have in recent years attracted strong theoretical support and increasing experimental evidence. We emphasize that both the defensive and the physiological functions of anthocyanins may operate in plants simultaneously.

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