CNGCs in Marchantia paleacea uncouple arbuscular mycorrhizal symbiosis and rhizoid development.

In Marchantia paleacea, MpaDMI1-dependent nuclear Ca2+ oscillations are essential for arbuscular mycorrhizal (AM) fungal colonisation, indicating that endosymbiosis-mediated nuclear Ca2+ signalling is a conserved feature of land plant-AM symbiosis. Despite this conservation, DOES NOT MAKE INFECTION (DMI)1 regulatory properties have diverged between bryophytes and angiosperms, suggesting lineage-specific adaptation and incomplete conservation of the Ca2+ oscillation machinery. In angiosperms, DMI1-dependent Ca2+ release requires CYCLIC NUCLEOTIDE-GATED CHANNELS (CNGC)15, but whether a comparable CNGC module operates in bryophytes, whose CNGC gene family is greatly reduced, has remained unknown. Here, we combined phylogenetic, genetic and cell biology approaches to investigate diverging land plant CNGCs function. Phylogenetic analyses across streptophytes reveal that CNGCs diversified into three ancient superclades before the terrestrialisation of plants. Functional analyses reveal that in M. paleacea, the combined activity of three MpaCNGCs spanning two superclades is required for endosymbiosis-associated nuclear Ca2+ oscillations and AM fungal colonisation. Although two of these MpaCNGCs redundantly regulate rhizoid elongation, AM fungi activate Ca2+ signalling and penetrate ventral cells lacking rhizoid growth, indicating that tip-growing cells are not strictly required for fungal entry in M. paleacea. Together, these findings link MpaCNGC function in rhizoid development and AM symbiosis to nutrient acquisition, supporting both soil exploration and AM fungal colonisation.