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Breeding of stress-tolerant plants is able to improve crop yield under stress conditions, whereas CRISPR/Cas9 genome editing has been shown to be an efficient way for molecular breeding to improve agronomic traits including stress tolerance in crops. However, genes can be targeted for genome editing to enhance crop abiotic stress tolerance remained largely unidentified. We have previously identified abscisic acid (ABA)-induced transcription repressors ( AITRs ) as a novel family of transcription factors that are involved in the regulation of ABA signaling, and we found that knockout of the entire family of  AITR  genes in Arabidopsis enhanced drought and salinity tolerance without fitness costs. Considering that AITRs are conserved in angiosperms,  AITRs  in crops may be targeted for genome editing to improve abiotic stress tolerance. We report here that mutation of  GmAITR  genes by CRISPR/Cas9 genome editing leads to enhanced salinity tolerance in soybean. By using quantitative RT-PCR analysis, we found that the expression levels of  GmAITRs  were increased in response to ABA and salt treatments. Transfection assays in soybean protoplasts show that GmAITRs are nucleus proteins, and have transcriptional repression activities. By using CRISPR/Cas9 to target the six  GmAITRs  simultaneously, we successfully generated Cas9-free  gmaitr36  double and  gmaitr23456  quintuple mutants. We found that ABA sensitivity in these mutants was increased. Consistent with this, ABA responses of some ABA signaling key regulator genes in the  gmaitr  mutants were altered. In both seed germination and seedling growth assays, the  gmaitr  mutants showed enhanced salt tolerance. Most importantly, enhanced salinity tolerance in the mutant plants was also observed in the field experiments. These results suggest that mutation of  GmAITR  genes by CRISPR/Cas9 is an efficient way to improve salinity tolerance in soybean.

Mutation of GmAITR Genes by CRISPR/Cas9 Genome Editing Results in Enhanced Salinity Stress Tolerance in Soybean