BACTERIAL-INDUCED CHANGES IN PLANT FORM AND FUNCTION

Numerous steps occur between the mutual recognition of host and Rhizobium and the establishment of a nitrogen-fixing indeterminate nodule: root hair curling and infection thread formation, cortical cell divisions and initiation of a nodule primordium, nodule meristem formation, differentiation of specific cell types and tissues, and induction of nitrogen fixation. Rhizobia with defined mutations or Agrobacterium tumefaciens transconjugants carrying R. meliloti symbiotic genes have been used to interrupt these steps. Using this approach, we have detected transcripts of the early nodulin MsENOD2 in alfalfa nodules induced by A. tumefaciens transconjugants carrying one or both symbiotic plasmids of R. meliloti. We have also specifically localized MsENOD2 transcripts by in situ hybridization to nodule parenchyma cells in nodules that form spontaneously on alfalfa roots. These bacteria-free nodules differentiate a discrete nodule meristem and other tissues that are typical of normal nodules. Transcripts for MsENOD12, a proline-rich protein normally found in the invasion zone of wildtype R. meliloti-induced nodules, were not detected in nodules induced by the Agrobacterium transconjugants or in the spontaneously formed nodules. However, we have detected MsENOD12 mRNAs in nodules formed in response to R. meliloti exoH mutants. The exoH mutants, which induce nodules without a persistent nodule meristem, infect host cells at a reduced frequency compared with wildtype or with fix or nif mutant R. meliloti. These results indicate that infection rather than the formation of a persistent nodule meristem is correlated with MsENOD12 gene expression.