A point mutation in the IAA14 promoter enhances callus formation and regeneration

Callus formation induced by auxin accumulation is considered the first step of in vitro plant regeneration. In Arabidopsis, degradation of the Aux/IAA protein, IAA14, in response to auxin signaling, which activates the AUXIN RESPONSE FACTOR 7 (ARF7) and ARF19 along with a series of downstream transcription factors, also plays a critical role in this process. However, the specific mechanism by which auxin regulates callus formation remains unclear. By screening mutant library in the solitary root 1 (iaa14/slr) Arabidopsis background we obtained the callus formation related 2 (cfr2) mutant. The cfr2 mutant exhibited a stronger capacity for callus formation, as well as lateral root and adventitious root regeneration from leaf explants than wild type (WT) seedlings, but did not recover gravitropism capability. The auxin signal in cfr2 was significantly enhanced, and the expression of some downstream transcription factors was increased. Map-based cloning, whole genome resequencing, and phenotypic complementation experiments showed that the phenotypes observed in the cfr2 mutant were caused by a point mutation in the IAA14 promoter region. This mutation, which is predicted to disrupt the binding of LBD16, LBD19, and LBD30 to the IAA14 promoter, changed the expression pattern of IAA14 in cfr2. Taken together, our results identified a new mutation in the IAA14 promoter region, which affects the expression pattern of IAA14 and in turn its ability to control plant regeneration.