Transgenic tobacco plants that express an antisense construct derived from a Medicago sativa cDNA encoding a Rac-related small GTP-binding protein fail to develop necrotic lesions upon elicitor infiltration.

Abstract

Using an RT-PCR approach a rac-related cDNA clone, designated Ms-rac1, was isolated from Medicago sativa (alfalfa). Ms-rac1 encodes a putative protein of 197 amino acids, which is closely related to known Rac-related GTP-binding proteins from Pisum sativum and Arabidopsis thaliana. RT-PCR analysis demonstrated that Ms-rac1 is ubiquitously expressed in various tissues in alfalfa. Expression of Ms-rac1 in suspension cultures occurred independently of treatment with elicitor, indicating that it is constitutively expressed. Heterologous expression of an antisense Ms-rac1 cDNA construct in transgenic tobacco plants was associated with poor growth and retarded flowering. Following infiltration with yeast elicitor, transgenic tobacco plants transformed with either the empty vector or Ms-rac1 in sense orientation developed brown necrotic lesions and subsequently cell death was observed within the infiltrated tissues. In contrast, the majority of the antisense transformants neither formed necrotic lesions nor showed any other visible defence reactions, demonstrating that Rac-related GTPases play an important role in the establishment of plant defence reactions.