Aquaporins are important transmembrane water transport proteins which transport water and several neutral molecules. However, how aquaporins are involved in the synergistic transport of Mg²⁺ and water remains poorly understood. Here, we found that the cassava aquaporin MePIP2;7 was involved in Mg²⁺ transport through interaction with MeMGT9, a lower affinity magnesium transporter protein. Knockdown of MePIP2;7 in cassava led to magnesium deficiency in basal mature leaves with chlorosis and necrotic spots on their edges and starch over-accumulation. Mg²⁺ content was significantly decreased in leaves and roots of MePIP2;7-RNA interference (PIP-Ri) plants grown in both field and Mg²⁺-free hydroponic solution. Xenopus oocyte injection analysis verified that MePIP2;7 possessed the ability to transport water only and MeMGT9 was responsible for Mg²⁺ efflux. More importantly, MePIP2;7 improved the transportability of Mg²⁺ via MeMGT9 as verified using the CM66 mutant complementation assay and Xenopus oocytes expressing system. Yeast two-hybrid, bimolecular fluorescence complementation, co-localization, and co-immunoprecipitation assays demonstrated the direct protein–protein interaction between MePIP2;7 and MeMGT9 in vivo. Mg²⁺ flux was significantly elevated in MePIP2;7-overexpressing lines in hydroponic solution through non-invasive micro-test technique analysis. Under Mg²⁺-free condition, the retarded growth of PIP-Ri transgenic plants could be recovered with Mg²⁺ supplementation. Taken together, our results demonstrated the synergistic effect of the MePIP2;7 and MeMGT9 interaction in regulating water and Mg²⁺ absorption and transport in cassava.

Pollen hydration on dry stigmas is strictly regulated by pollen–stigma interactions in Brassicaceae. Although several related molecular events have been described, the molecular mechanism underlying pollen hydration remains elusive. Multiple B-class pollen coat proteins (PCP-Bs) are involved in pollen hydration. Here, by analyzing the interactions of two PCP-Bs with three Arabidopsis thaliana stigmas strongly expressing S-domain receptor kinase (SD-RLK), we determined that SD-RLK28 directly interacts with PCP-Bβ. We investigated pollen hydration, pollen germination, pollen tube growth, and stigma receptivity in the sd-rlk28 and pcp-bβ mutants. PCP-Bβ acts in the pollen to regulate pollen hydration on stigmas. Loss of SD-RLK28 had no effect on pollen viability, and sd-rlk28 plants had normal life cycles without obvious defects. However, pollen hydration on sd-rlk28 stigmas was impaired and pollen tube growth in sd-rlk28 pistils was delayed. The defect in pollen hydration on sd-rlk28 stigmas was independent of changes in reactive oxygen species levels in stigmas. These results indicate that SD-RLK28 functions in the stigma as a PCP-Bβ receptor to positively regulate pollen hydration on dry stigmas.