IBA induces adventitious rooting in Arabidopsis thaliana thin cell layers by conversion into IAA, involving nitric oxide formation, IAA transport, and IAA biosynthesis

Adventitious roots (AR) are essential for plant survival and propagation via cuttings. Indole-3-acetic acid (IAA), and its precursor indole-3-butyric acid (IBA), control the AR-process. In cuttings of numerous species, exogenous IBA is more AR-inductive than exogenous IAA, but the reason needs investigation. In Arabidopsis thaliana thin cell layers (TCLs), IBA induces ARs when combined with Kinetin (Falasca et al., Plant Cell Rep, 2004), but in dark-grown seedlings IBA is able alone to induce AR-formation (Veloccia et al., J Exp Bot, 2016). In cuttings as in planta, the endogenous IAA/IBA contents are determinant for the AR-process, and differences in response to exogenous IAA/IBA may depend on differences in endogenous contents. It is unknown whether Arabidopsis TCLs contain endogenous IAA/IBA at culture onset. Results showed that IAA and IBA were at undetectable levels at culture onset, and this was an optimal premise to investigate AR-formation under the total control of exogenous auxin, revealing possible differences between IAA and IBA. The AR-response of TCLs from various ecotypes, transgenic lines and knockout mutants showed that IBA was an AR-inducer better than IAA. IBA positively affected IAA cellular efflux and influx, and expression of ANTHRANILATE SYNTHASE-alpha1 (ASA1), a gene of the tryptophan-dependent IAA biosynthesis. Moreover, ASA1 and ANTHRANILATE SYNTHASE-beta1 (ASB1), the other subunit of the enzyme, positively affected AR-formation in the presence of exogenous IBA. The AR-response of IBA-treated TCLs from ech2ibr10 mutant, blocked into IBA-to-IAA conversion, was strongly reduced, showing that IBA acted mainly by conversion into IAA. Nitric oxide (NO), a downstream signal of IAA, but also a by-product of the conversion process, was early detected in IAA- and IBA-treated TCLs, but at higher levels in the latter ones. Altogether results showed that exogenous IBA induced AR-formation in TCLs by conversion into IAA involving NO, IAA transport, and ASA1/ASB1-mediated IAA biosynthesis.