Transcriptomic and Functional Characterization Reveals Cshak5;3 as Key Player in K+ Homeostasis of Grafted Cucumbers Under Salinity

Abstract

Grafting can improve the salt tolerance of many crops. However, whereas the critical genes in scions responsive to rootstock under salt stress remain a mystery. We found that pumpkin rootstock decreased the content of Na+ by 70.24%, increased the content of K+ by 25.9%, and increased the K+/Na+ ratio by 366.0% in cucumber scion leaves. Furthermore, RNA-seq analysis showed that ion transport-related genes were key genes responding to salt stress tolerance in grafted cucumbers. The identification and expression analysis of K+ channel proteins in cucumber and pumpkin reveals 6 and 5 HAK5 members in cucumber and pumpkin, respectively. The expression of CsHAK5;3 in cucumber was elevated in different graft combinations under salt stress, but most notably in C/P (cucumber scion/pumpkin rootstock). CsHAK5;3 was localized to the plasma membrane and a yeast complementation assay revealed that it could transport K+. Knockout of CsHAK5;3 in hairy root mutants decreased the K+ content of leaves (45.6%) and roots (50.3%), increased the Na+ content of leaves (29.3%) and roots (34.8%), and decreased the K+/Na+ ratio of the leaves (57.9%) and roots (62.9%) in cucumber. However, CsHAK5;3 overexpression in hairy roots increased the K+ content of the leaves (31.2%) and roots (38.3%), decreased the Na+ content of leaf (17.2%) and root (14.3%), and increased the K+/Na+ ratio of the leaf (58.9%) and root (61.6%) in cucumber. In conclusion, CsHAK5;3 of cucumber can mediate K+ transport, which is a key target gene of pumpkin rootstock to enhance grafted cucumber salt tolerance.

Keywords: grafted cucumber, Salinity, K+ homeostasis, HAK5, transcriptome