Expression of srab7 and SCaM genes required for endocytosis of Rhizobium in root nodules

Symbiotic nitrogen fixation requires the internalization of rhizobia into root cells of legumes and subsequent formation of symbiosomes, rhizobia-containing organelles inside the plant cells. A small GTP-binding protein from soybean, sRab7, was shown to be essential for nodule development. When yeast ypt7 (Rab7) null mutant was transformed with the srab7 gene, highly fragmented vacuoles seen in the mutant were replaced with a prominent central vacuole, as in the wild-type cells. These results confirm that sRab7 is an authentic homolog of Ypt7p. When nodules obtained at different stages of development were hybridized with srab7 cDNA, 7-day-old nodules showed the most intense hybridization signal. Hybridization was only detected in the infection zone of forming nodules, implying that sRab7 may be involved in endocytosis of rhizobia. SCaM-1 and SCaM-4, soybean calmodulin genes, were also expressed in the same region at this stage of nodulation as shown by in situ hybridization. RT-PCR analysis confirmed the expression of these genes. These results indicate that both the srab7 and SCaM genes were expressed at an early stage of nodulation. Since membrane fusion of vesicles has been shown to be dependent on calmodulin and Ypt7p, the similarity of the expression patterns between the two genes in soybean nodules may indicate that sRab7 and SCaM are essential for vesicular fusion during rhizobial endocytosis and symbiosome formation.