Two homologous INDOLE-3-ACETAMIDE (IAM) HYDROLASE genes are required for the auxin effects of IAM in Arabidopsis

Indole-3-acetamide (IAM) is the first confirmed auxin biosynthetic intermediate in some plant pathogenic bacteria. Exogenously applied IAM or production of IAM by overexpressing the bacterial iaaM gene in Arabidopsis causes auxin overproduction phenotypes. However, it is still inconclusive whether plants use IAM as a key precursor for auxin biosynthesis. Herein, we reported the isolation IAMHYDROLASE1 (IAMH1) gene in Arabidopsis from a forward genetic screen for IAM-insensitive mutants that display normal auxin sensitivities. IAMH1 has a close homolog named IAMH2 that is located right next to IAMH1 on chromosome IV in Arabidopsis. We generated iamh1 iamh2 double mutants using our CRISPR/Cas9 gene editing technology. We showed that disruption of the IAMH genes rendered Arabidopsis plants resistant to IAM treatments and also suppressed the iaaM overexpression phenotypes, suggesting that IAMH1 and IAMH2 are the main enzymes responsible for converting IAM into indole-3-acetic acid (IAA) in Arabidopsis. The iamh double mutants did not display obvious developmental defects, indicating that IAM does not play a major role in auxin biosynthesis under normal growth conditions. Our findings provide a solid foundation for clarifying the roles of IAM in auxin biosynthesis and plant development.