The maize aquaporin ZmPIP1;6 enhances stomatal opening and CO2- and ABA-induced stomatal closure

The plasma membrane aquaporin ZmPIP1;6 is expressed in maize stomatal complexes, with higher expression during the day than at night. To elucidate the role of ZmPIP1;6 in gas exchange and stomatal movement, it was expressed in maize (inbred line B104) under the control of the p35S promoter (overexpression) or its native promoter fused with mYFP (monomeric yellow fluorescent protein) cDNA (mYFP-ZmPIP1;6). In stomatal complexes of the mature zone of the leaf, mYFP-ZmPIP1;6 showed higher expression in subsidiary cells than in guard cells, and light and dark treatments influenced its subcellular localization. Notably, ZmPIP1;6 internalization increased in dark conditions compared with light. Stomatal opening was greater in ZmPIP1;6 overexpression plants than in the wild type, while closure exhibited greater sensitivity to elevated CO2 concentration or abscisic acid (ABA) treatment. Our findings revealed that reactive oxygen species (H2O2) were involved in ABA-induced stomatal closure, while ZmPIP1;6 was unable to facilitate H2O2 diffusion when expressed in yeast. Finally, ZmPIP1;6 overexpression and mYFP-ZmPIP1;6 transgenic plants exhibited higher abaxial stomatal density than the wild type. Overall, these results indicate that ZmPIP1;6 plays important roles in stomatal opening and CO2- and ABA-induced stomatal closure.