The miR156u/v-MhDIV3 Module Modulates Cadmium Uptake and Damage via Enhancing MhNRAMP1 Expression in Malus hupehensis

Cadmium (Cd), a toxic heavy metal, threatens crop production and human health, and its uptake by Malus hupehensis is regulated by MhNRAMP1. The role of the DIVARICATA (DIV) transcription factor (TF) in Cd stress remains largely unclear. Here, nine DIVs were isolated from M. hupehensis based on CDS sequences of identified DIV members in Malus. Among them, MhDIV3 exhibited the earliest and strongest response to Cd. Its encoded protein, MhDIV3, possesses the characteristics of R2R3-MYB TF. Suppression of MhDIV3 in M. hupehensis roots and apple calli led to a higher fresh weight and lower levels of reactive oxygen species (ROS) and malondialdehyde (MDA) under Cd stress, while overexpression of MhDIV3 in M. hupehensis roots and tomato exacerbated Cd-caused oxidative damage by accelerating Cd²⁺ uptake. Mechanistically, MhDIV3 bound to an enhancer in the intron of MhNRAMP1 to positively regulate its expression under Cd stress. Additionally, miR156u/v, an upstream regulator of MhDIV3, suppressed MhDIV3 expression by complementing its 3'UTR. Overexpression of miR156u/v reduced Cd²⁺ uptake and stress damage, similar to MhDIV3-suppression. Overall, our results suggested that miR156u/v-MhDIV3 module positively modulates Cd uptake and damage by triggering MhNRAMP1 expression in M. hupehensis.