ROLES OF PLANT HOMOLOGS OF RAB1P AND RAB7P IN THE BIOGENESIS OF THE PERIBACTEROID MEMBRANE, A SUBCELLULAR COMPARTMENT FORMED DE-NOVO DURINGROOT-NODULE SYMBIOSIS

The peribacteroid membrane (PBM) in legume root nodules is derived fro m plasma membrane following endocytosis of Rhizobium by fusion of newl y synthesized vesicles. We studied the roles of plant Rab1p and Rab7p homologs, the small GTP-binding proteins involved in vesicular transpo rt, in the biogenesis of the PBM. Three cDNAs encoding legume homologs of mammalian Rab1p and Rab7p were isolated from soybean (sRab1p, sRab 7p) and Vigna aconitifolia (vRab7p). sRab1p was confirmed to be a func tional counterpart of yeast Ypt1p (Rab1p) by complementation of a yeas t ypt1-1 mutant. Both srab1 and vrab7 genes are induced during nodulat ion with the level of vrab7 mRNA being 12 times higher than that in ro ot meristem and leaves. This induction directly correlates with membra ne proliferation in nodules. Antisense constructs of srab1 and vrab7, under a nodule-specific promoter (leghemoglobin, Lbc3), were made in a binary vector and transgenic nodules were developed on soybean hairy roots obtained through Agrobacterium rhizogenes-mediated transformatio n. Both antisense srab1 and vrab7 nodules were smaller in size and sho wed tower nitrogenase activity than controls. The antisense srab1 nodu les showed lack of expansion of infected cells, fewer bacteroids per c ell and their frequent release into vacuoles. In contrast, antisense v rab7 expressing nodules showed accumulation of late endosomal structur es and multivesicular bodies in the perinuclear region. These data sug gest that both Rab1p and Rab7p are essential for the development of th e PBM compartment in effective symbiosis.