The Independent Acquisition of Plant Root Nitrogen-Fixing Symbiosis in Fabids Recruited the Same Genetic Pathway for Nodule Organogenesis

Abstract

Only species belonging to the Fabid clade, limited to four classes and ten families of Angiosperms, are able to form nitrogen-fixing root nodule symbioses (RNS) with soil bacteria. This concerns plants of the legume family (Fabaceae) and Parasponia (Cannabaceae) associated with the Gram-negative proteobacteria collectively called rhizobia and actinorhizal plants associated with the Gram-positive actinomycetes of the genus Frankia. Calcium and calmodulin-dependent protein kinase (CCaMK) is a key component of the common signaling pathway leading to both rhizobial and arbuscular mycorrhizal symbioses (AM) and plays a central role in cross-signaling between root nodule organogenesis and infection processes. Here, we show that CCaMK is also needed for successful actinorhiza formation and interaction with AM fungi in the actinorhizal tree Casuarina glauca and is also able to restore both nodulation and AM symbioses in a Medicago truncatula ccamk mutant. Besides, we expressed auto-active CgCCaMK lacking the auto-inhibitory/CaM domain in two actinorhizal species: C. glauca (Casuarinaceae), which develops an intracellular infection pathway, and Discaria trinervis (Rhamnaceae) which is characterized by an ancestral intercellular infection mechanism. In both species, we found induction of nodulation independent of Frankia similar to response to the activation of CCaMK in the rhizobia-legume symbiosis and conclude that the regulation of actinorhiza organogenesis is conserved regardless of the infection mode. It has been suggested that rhizobial and actinorhizal symbioses originated from a common ancestor with several independent evolutionary origins. Our findings are consistent with the recruitment of a similar genetic pathway governing rhizobial and Frankia nodule organogenesis.

The Nod factor-independent nodulation in Aeschynomene evenia required the common plant-microbe symbiotic "toolkit"

“…Similar symbiotic cascade requirements were also found in the actinorhizal plant C. glauca, in which both SYMRK (Gherbi et al, 2008) and CCaMK (Svistoonoff et al, 2013) were shown to be required for nodulation. It is noteworthy that a microorganism lacking nod genes activates such a signaling pathway.…”

“…The symbiotic calcium-and calmodulin-dependent protein kinase CCaMK is required for the interpretation of this Ca 2+ signature (Oldroyd and Downie, 2006;Madsen et al, 2010;Singh et al, 2014). Mutations in CCaMK or CCaMK RNAi prevent root infections by microsymbionts through the inhibition of either the formation of ITs or the endocytosis of rhizobia inside the nodules, and they block or, in the case of RNAi lines, reduce nodule organogenesis in legumes (Catoira et al, 2000;Lévy et al, 2004;Mitra et al, 2004;Capoen et al, 2009;Sinharoy and DasGupta, 2009) and actinorhizal plants (Svistoonoff et al, 2013).…”

“…a kinase domain and an autoinhibition domain overlapping a calmodulinbinding domain, such that autoinhibition is released in the presence of calmodulin). Deletion of this autoinhibition domain or a phosphomimetic version of CCaMK leads to the formation of spontaneous nodules in the absence of symbionts in legumes (Gleason et al, 2006;Tirichine et al, 2006;Takeda et al, 2012;Miller et al, 2013), in C. glauca (Svistoonoff et al, 2013), and in Parasponia andersonii, the only nonlegume able to form a nitrogen-fixing symbiosis with rhizobia (Op den Camp et al, 2011).…”

“…Interestingly, two so-called common symbiotic genes, CgSYMRK/DgSYMRK and CgCCaMK, have been shown to be also necessary for the establishment of the symbiosis between actinorhizal plants (Casuarina glauca and Datisca glomerata) and Frankia spp. (Gherbi et al, 2008;Markmann et al, 2008;Svistoonoff et al, 2013).…”

Inhibition of Auxin Signaling in Frankia Species-Infected Cells in Casuarina glauca Nodules Leads to Increased Nodulation

“…The nature of the nodulation signal(s) emitted by Frankia spp. is still unknown, but it has been shown that its perception depends on the common Nod signaling pathway (Gherbi et al, 2008;Svistoonoff et al, 2013). In the actinorhizal tree Casuarina glauca, the involvement of the phytohormone auxin in symbiotic signaling in plant cells infected by Frankia spp.…”