The key role of indole-3-acetic acid biosynthesis by Bacillus thuringiensis RZ2MS9 in promoting maize growth revealed by the ipdC gene knockout mediated by the CRISPR-Cas9 system.

ABSTRACT

The bacterial biosynthesis of indole-3-acetic acid (IAA) is often related to the beneficial effects of plant growth-promoting rhizobacteria (PGPR) on plant development. In PGPR belonging to the Bacillus genus, the synthesis of IAA may occur through different metabolic pathways that are still poorly understood. B. thuringiensis (Bt) is well known for its insecticidal properties; however, its beneficial features are not limited to pest control. Our group has been studed the beneficial effects of Bt strain RZ2MS9 as growth promoter in a range of plant crops, including soybean, tomato, and maize. We recently demonstrated that bacterial IAA biosynthesis plays an important role in the ability of RZ2MS9 to benefit plant development. However, the molecular involved mechanisms in the IAA biosynthesis by this bacterium in the beneficial interaction with plants remain unclear. Here, we investigated the genetic basis of IAA biosynthesis by RZ2MS9. We knocked out the ipdC gene, involved in IAA biosynthesis via the tryptophan-dependent IPyA pathway, using the CRISPR-Cas9 system. Our results showed that, by disrupting the IPyA pathway, the amount of IAA synthesized by the mutant RZ2MS9 (ΔipdC) in the presence of tryptophan drops 57%. The gene knockout did not affect the bacterial growth, but it did affect its ability to colonize maize. Moreover, deactivating the ipdC gene in RZ2MS9 significantly reduces its ability to promote maize growth. ΔipdC performed worse than RZ2MS9 in almost all evaluated plant parameters, including total root length, projected root area, lateral roots, aerial part dry matter, and germination speed index. Therefore, we demonstrated that tryptophan-dependent IAA biosynthesis via the IPyA pathway by RZ2MS9 is strongly influenced by the ipdC gene. Furthermore, IAA biosynthesis by RZ2MS9 is a major mechanism used by this PGPR to promote maize growth.