CALMODULIN-LIKE16 and PIN-LIKES7a cooperatively regulate rice seedling primary root elongation under chilling

Low-temperature sensitivity at the germination stage is a challenge for direct seeding of rice in Asian countries. How Ca2+ and auxin (IAA) signaling regulate primary root growth under chilling remains unexplored. Here, we showed that OsCML16 interacted specifically with OsPILS7a to improve primary root elongation of early rice seedlings under chilling. OsCML16, a subgroup 6c member of the OsCML family, interacted with multiple cytosolic loop regions of OsPILS7a in a Ca2+-dependent manner. OsPILS7a localized to the endoplasmic reticulum membranes and functioned as an auxin efflux carrier in a yeast growth assay. Transgenics showed that presence of OsCML16 enhanced primary root elongation under chilling, whereas the ospils7a knockout mutant lines showed the opposite phenotype. Moreover, under chilling conditions, OsCML16 and OsPILS7a-mediated Ca2+ and IAA signaling and regulated the transcription of IAA signaling-associated genes (OsIAA11, OsIAA23, and OsARF16) and cell division marker genes (OsRAN1, OsRAN2, and OsLTG1) in primary roots. These results show that OsCML16 and OsPILS7a cooperatively regulate primary root elongation of early rice seedlings under chilling. These findings enhance our understanding of the crosstalk between Ca2+ and IAA signaling and reveal insights into the mechanisms underlying cold-stress response during rice germination.