Abstract
The formation of nitrogen-fixing nodules in legumes involves the initiation of synchronized programs in the root epidermis and cortex to allow rhizobial infection and nodule development. 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 rhizobia promote a reduction in callose levels in inner tissues where nodules initiate. This downregulation coincides with the localized expression of M.truncatula β-1,3-glucanase 2 (MtBG2), encoding a novel PD-associated callose-degrading enzyme. Spatiotemporal analyses revealed that MtBG2 expression expands from dividing nodule initials to rhizobia-colonized cortical and epidermal tissues. As shown by the transport of fluorescent molecules invivo, symplastic-connected domains are created in rhizobia-colonized tissues and enhanced in roots constitutively expressing MtBG2. MtBG2-overexpressing roots additionally displayed reduced levels of PD-associated callose. Together, these findings suggest an active role for MtBG2 in callose degradation and in the formation of symplastic domains during sequential nodule developmental stages. Interfering with symplastic connectivity led to drastic nodulation phenotypes. Roots ectopically expressing β-1,3-glucanases (including MtBG2) exhibited increased nodule number, and those expressing MtBG2 RNAi constructs or a hyperactive callose synthase (under symbiotic promoters) showed defective nodulation phenotypes. Obstructing symplastic connectivity appears to block a signaling pathway required for the expression of NODULE INCEPTION (NIN) and its target NUCLEAR FACTOR-YA1 (NF-YA1) in the cortex. We conclude that symplastic intercellular communication is proactively enhanced by rhizobia, and this is necessary for appropriate coordination of bacterial infection and nodule development.
Highlights
Legumes establish symbiotic associations with nitrogen-fixing rhizobia, which, hosted within newly formed root nodules, have the optimized environment for fixing atmospheric dinitrogen for the plant [1]
We show that callose is reduced in root tissues forming the primordia and that this correlates with the progressive expression of a rhizobia-induced, PD-located, callose-degrading enzyme
Rhizobial Inoculation Modulates Callose Levels and the Early Expression of Callose-Degrading b-1,3Glucanases in M. truncatula Roots Past research indicates that symplastic connectivity increases between phloem and nodule initials as early as 48 hr upon inoculation with rhizobia [34], raising the question of whether callose regulation is involved in this process
Summary
Legumes establish symbiotic associations with nitrogen-fixing rhizobia, which, hosted within newly formed root nodules, have the optimized environment for fixing atmospheric dinitrogen for the plant [1]. Endosymbiotic entry occurs via newly formed tubular structures, initiated from bacterial-entrapped root hairs, called infection threads (ITs) [2]. This host-constructed transcellular compartment guides rhizobia to the underlying nodule primordium in the cortex, where the bacteria are released and differentiate into nitrogen-fixing bacteroids. Nodule development engages local hormonal changes and is tightly controlled by a systemic autoregulatory pathway (AON) that restricts nodule number [4] Diffusible hormones, such as cytokinin and auxin [5, 6], are important for initiating nodule organogenesis, little is known of how signal transmission is regulated from the infection site to underlying root tissues to coordinate nodule development
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