Abstract

Cold stress limits the entire growth cycle of plants, affecting crop yield, quality, and geographic distribution. The calcium ion (Ca2+) signal, generated by rapid and dynamic changes in cytoplasmic free Ca2+ concentration ([Ca2+]cyt), plays a key role in plant cold responses, but the mechanisms governing the initiation and termination of cold-induced Ca2+ signaling remain unclear. Here, a cyclic nucleotide-gated channel (CNGC) family member, MdCNGC1C, was found to positively regulate apple (Malus domestica) freezing tolerance by mediating cold-induced Ca2+ influx. MdOST1, a SnRK2 family kinase, interacts with and phosphorylates MdCNGC1C at Ser47 in response to cold stress, activating its channel activity and enhancing MdCNGC1C-mediated Ca2+ influx and cold tolerance. The Ca2+ sensor MdCaM7.1 also interacts with MdCNGC1C but inhibits its Ca2+ influx activity, thereby negatively regulating cold tolerance. MdOST1 competes with MdCaM7.1 for binding to the N-terminus of MdCNGC1C, disrupting the MdCaM7.1-MdCNGC1C interaction. However, Ca2+ treatment promotes the interaction with MdCaM7.1 and reduces MdOST1's inhibitory effect, indicating a potential [Ca2+]cyt-dependent feedback regulation of MdCNGC1C activity and cold-induced Ca2+ signaling mediated by MdOST1 and MdCaM7.1. Therefore, this study reveals a competitive interaction module of MdOST1-MdCNGC1C-MdCaM7.1 that fine-tunes the cold response in apple via Ca2+ signaling, offering insights into the dynamic mechanisms underlying plant cold responses.