OsSPL3, a SBP-Domain Protein, Regulates Crown Root Development in Rice.

The major root system of cereals consists of crown roots (or adventitious roots) which are important for anchoring plants to soil and for water and nutrient uptake. However, the molecular basis of crown root formation is largely unknown. Here, we isolated a rice mutant with fewer crown roots, named lower crown root number1 (lcrn1). Map-based cloning revealed that lcrn1 is caused by a mutation of a putative transcription factor coding gene OsSPL3 (SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE3). We demonstrate that the point mutation in lcrn1 perturbs the OsmiR156-directed cleavage of OsSPL3 transcripts, resulting in the mutant phenotype. Chromatin immunoprecipitation sequencing assays of OsSPL3-binding sites and RNA-sequencing of differentially expressed transcripts in lcrn1 further identified potential direct targets of OsSPL3 in basal nodes, including a MADS-box transcription factor OsMADS50. OsMADS50-overexpressing plants produced fewer crown roots, phenocopying lcrn1, while knocking out OsMADS50 in lcrn1 reversed this phenotype. We also show that OsSPL12, another OsmiR156 target gene, regulates OsMADS50 and the crown root development. Taken together, our findings suggest a hitherto unknown regulatory pathway in which the OsmiR156-OsSPL3/OsSPL12 module directly activates OsMADS50 in the node to regulate crown root development in rice.