Summary
Cell-to-cell communication is critical for multicellular organisms. In plants, plasmodesmata—cytoplasmic channels—enable molecular transport between adjacent cells. In roots, this transport is predicted to be essential in nutrient acquisition and delivery to the vasculature. We demonstrate that plasmodesmatal transport persists in differentiated roots, despite apoplastic barriers such as Casparian strips and suberin lamellae in the endodermis, suggesting plasmodesmata as the sole pathway for water and nutrient flow at this stage. We also reveal a developmental switch in plasmodesmata function resulting in an unidirectional transport in differentiated roots. A genetic screen identified mutations that disrupt this directionality, leading to bidirectional transport. These mutations correlate with larger plasmodesmatal apertures, linked to defects in pectin composition and cell wall organization. This discovery underscores the role of plasmodesmatal aperture regulation and pectin in controlling directional transport. Our findings provide insights into plasmodesmata function and their regulation in roots.