RHIZOBIUM-MELILOTI NOD FACTORS ELICIT CELL-SPECIFIC TRANSCRIPTION OF THE ENOD12 GENE IN TRANSGENIC ALFALFA

Extracellular lipo oligosaccharides of Rhizobium, known as Nod factors , play a key role in the molecular signal exchange which leads to the specific nitrogen-fixing symbiotic association between the soil microb e and its host legume. The biological activity of Nod factors and thei r perception by the host plant during the earliest stages of the Rhizo bium/legume interaction have been studied using transgenic alfalfa car rying a fusion between the promoter of the early nodulin gene MtENOD12 and the beta-glucuronidase (GUS) reporter gene. Histochemical stainin g has shown that GUS accumulates specifically in the differentiating r oot epidermis, prior to and during root hair emergence, within 2-3 h f ollowing the addition of purified Rhizobium meliloti Nod factors. This precocious transcriptional activation of the MfENOD12 gene, reminisce nt of that observed after inoculation with intact Rhizobium, implies t hat the Nod factor signal can be perceived at a developmental stage pr eceding root hair formation. GUS activity can be detected following tr eatment with a wide range of R. meliloti Nod factor concentrations dow n to 10(-13) M, and furthermore, this rapid response to the bacterial elicitor appears to be non-systemic. Significantly, MtENOD12-GUS expre ssion is not observed after inoculation with a R. meliloti nodH mutant which synthesizes exclusively non-sulphated Nod factors. Indeed purif ied Nod factors which lack the sulphate substituent are approximately 1000-fold less active than their sulphated counterparts. Thus, the tri ggering of ENOD12 transcription in the alfalfa root epidermis is a rap id molecular response which is subject to the same host-specificity de terminant (Nod factor sulphation) that governs the interaction between alfalfa and its bacterial symbiont.