Analysis of ENOD40 expression in alb1, a symbiotic mutant of Lotus japonicus that forms empty nodules with incompletely developed nodule vascular bundles

The alb1 mutant of Lotus japonicus (Ljsym74) forms empty nodules in which m ost of the bacteria remain in abnormally enlarged infection threads and fai l to enter the host plant cells. The alb1 mutant was also found to be defec tive in differentiation of ramified nodule vascular bundles; only a single vascular bundle differentiates at the proximal end of the alb1 nodules and it fails to differentiate further. Histochemical analysis using fluorescein -conjugated wheat-germ agglutinin (F-WGA) indicated that the mutation in th e ALB1 gene specifically affects the differentiation of vascular bundles in nodules. Analysis of nodulin gene expression revealed that the expression of an early nodulin gene, ENOD40, was very low in alb1 nodules. At early de velopmental stages of alb1 nodules, the pattern of ENOD40 transcription was essentially the same as that in wild-type nodules; transcripts were locali zed in dividing cortical cells and in the pericycle of the root stele oppos ite nodule primordia, as in wild-type nodules, However, mature alb1 nodules exhibited very weak or no expression of ENOD40 ill the peripheral cells of the undeveloped nodule vascular bundle. The ENOD40 expression pattern in a lb1 nodules is distinct from that in another ineffective mutant, fen1 (Ljsy m76), in which ENOD40 expression persists prior to premature senescence. Th ese findings lead us to speculate that ENOD40 may play a role in the differ entiation of nodule vascular bundles.