Gain-of-function mutations of AtNHX1 suppress sos1 salt sensitivity and improve salt tolerance in Arabidopsis

Abstract

Soil salinity severely hampers agricultural productivity. Under salt stress, excess Na⁺ accumulation causes cellular damage and plant growth retardation, and membrane Na⁺ transporters play central roles in Na⁺ uptake and exclusion to mitigate these adverse effects. In this study, we performed sos1 suppressor mutant (named sup) screening to uncover potential genetic interactors of SOS1 and additional salt tolerance mechanisms. Map-based cloning and sequencing identified a group of mutants harboring dominant gain-of-function mutations in the vacuolar Na⁺/H⁺ antiporter gene AtNHX1. The gain-of-function variants of AtNHX1 showed enhanced transporter activities in yeast cells and increased salt tolerance in Arabidopsis wild type plants. Ion content measurements indicated that at the cellular level, these gain-of-function mutations resulted in increased cellular Na⁺ accumulation likely due to enhanced vacuolar Na⁺ sequestration. However, the gain-of-function suppressor mutants showed reduced shoot Na⁺ but increased root Na⁺ accumulation under salt stress, indicating a role of AtNHX1 in limiting Na⁺ translocation from root to shoot. We also identified another group of sos1 suppressors with loss-of-function mutations in the Na⁺ transporter gene AtHKT1. Loss-of-function mutations in AtHKT1 and gain-of-function mutations in AtNHX1 additively suppressed sos1 salt sensitivity, which indicates that the three transporters, SOS1, AtNHX1 and AtHKT1 function independently but coordinately in controlling Na⁺ homeostasis and salt tolerance in Arabidopsis. Our findings provide valuable information about the target amino acids in NHX1 for gene editing to improve salt tolerance in crops.

Keywords

sos1 suppressor / AtHKT1 / AtNHX1 / Gain-of-function / Salt tolerance

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