A novel GRAS protein gene MtSymSCL1 plays a role in regulating nodule number in Medicago truncatula

Legume-rhizobium symbiosis requires the expression of a series of plant genes playing roles in nodulation signaling that leads to the formation of functional nodules. A few plant-specific GRAS proteins have been shown to be involved in early nodulation signaling. Here, we isolated a novel SCARECROW -like GRAS protein gene, MtSymSCL1 , from the model legume Medicago truncatula . MtSymSCL1 has unique gene and protein domain structures, and its homologs are identified from various plant species. Phylogenetic analysis showed that MtSymSCL1 could be classified into a novel subclass in the GRAS protein family. MtSymSCL1 transcript accumulated highly in nodulated roots at 14 days after inoculation of Sinorhizobium meliloti . Translationally fused MtSymSCL1-GFP expressed from a viral or the native promoter was co-localized with nuclear localization signal-fused RFP in leaf mesophyll protoplasts , Agrobacterium rhizogenes -transformed hairy root cells, and epidermal cells of nodules that developed on the transformed roots. Unlike SCARECROW of Arabidopsis , however, RNA silencing of MtsymSCL1 did not affect root growth, nor root hair curling and infection thread growth. Nevertheless, MtSymSCL1 -silenced roots developed significantly fewer nodules than GUS -silenced roots, as shown by replicated nodulation assay with pot- or agar plate-grown plants. The structure of MtSymSCL1 -silenced root nodules appeared similar to that of the wild type, as shown by sectioned nodules stained with green fluorescent dye. Together, these results suggest that MtSymSCL1 plays a role in regulating nodule number during legume-rhizobium symbiosis.