Callose-Regulated Symplastic Communication Coordinates Symbiotic Root Nodule Development

Abstract

The formation of nitrogen-fixing nodules in legumes involves the initiation of nodule organogenesis in the root cortex, which is tightly synchronized, with symbiotic rhizobial infection in the epidermis. In this study, we provide evidence that symplastic communication, regulated by callose turnover at plasmodesmata (PD), is important for coordinating nodule development and infection in Medicago truncatula. Here we show that nodule organogenesis involves degradation of callose and the induced expression of a novel PD-associated callose-degrading enzyme (MtBG2) in inner root tissues where nodule initiates. MtBG2 is PD-localized and reduces PDcallose levels when constitutively expressed in roots. Spatio-temporal analyses revealed that MtBG2 expression expands from dividing nodule initials to rhizobial-colonized cortical and epidermal tissues. As shown by the transport of fluorescent molecules in vivo, symplastic-connected domains are created in these rhizobial-colonized tissues. These findings highlight that progressive degradation of PD-callose, driven in part by the upregulation of MtBG2, creates symplastic domains during sequential nodule developmental stages. Interfering with symplastic connectivity in rhizobial-infected tissues led to drastic nodulation phenotypes. Indeed, enhancing or blocking it by the controlled expression of PD-localized β- 1,3-glucanases (including MtBG2) or a hyperactive callose synthase, led to enhanced or reduced numbers of colonized nodules respectively. Obstructing symplastic connectivity appears to block the transmission of a diffusible signal required for the expression of Nodule Inception (NIN) in the cortex, while epidermal NIN expression is maintained. We conclude that symplastic communication between cells is proactively enhanced by rhizobia and this is necessary to allow appropriate coordination of bacterial infection and nodule development.