On basis of the preliminary results obtained with the above mentioned reporter lines we can conclude that L. japonicus responses to Nod factors display various differences with other legumes tested previously. For instance, the response of GH3 promoter activity in the vascular bundle is different from the results obtained in the clover and pea system (Mathesius et al. 1998; Boot et al. 1999). Whereas clover and pea respond to the local application of Nod factor with a local decrease of GH3 promoter activity in the vascular bundle, such a decrease was not observed in L. japonicus. The results indicate that in L. japonicus Nod factors do not redirect auxin to the nodule primordia by influencing auxin flow in the vascular tissue. Rather the results indicate that Nod factors might induce the local production of auxin in the nodule primordia. Also in the response of the ENOD40 promoters clear differences are found when compared with the results for other plants, such as Medicago sativa (Fang et al. 1998). For instance, a very poor response could be detected towards cytokinin in transgenic L. japonicus plants containing the soybean or L. japonicus promoter fused to GUS-GFP. These lines responded specifically to M. loti Nod factors at nanomolar concentrations in the outer cortex and the root hairs. Interestingly, transgenic lines that contain multiple integrations of the soybean ENOD40 reporter construct showed no constitutive GUS expression in the vascular tissue as was detected in the single integration lines. In none of the tested lines a response to the compound mastoporan was detected. In contrast, mastoporan is a strong inducer of the ENOD12 promoter in M. sativa (Pingret et al. 1998). We are currently involved in a pharmacological approach to test the effect of many other compounds on the expression of the ENOD40 promoter. For this purpose, the application of microtargeting techniques for direct introduction of the tested compounds in the plant tissue will be very valuable (Sautter et al. 1991).
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