ATXR2 as a core regulator of de novo root organogenesis (2025)

Differentiated plant somatic cells can form unorganized pluripotent cell mass callus. Callus formation can be stimulated mainly by two different methods, mechanical wounding and incubation on high auxin-containing CIM medium.1^,2 In particular, CIM-induced callus is analogous to lateral root primordial tissues.1 In support, CIM-induced callus formation requires many molecular components involved in lateral root initiation.1^,3 Further, several root meristem genes are subsequently activated during callus formation.1 Based on these observations, callus formation from leaf explants looks similar to de novo root organogenesis.4

We recently investigated a role of the ATXR2 trithorax group (TrxG) protein in CIM-induced callus formation.5 ATXR2 promotes global accumulation of H3K36me3 during callus formation on CIM,5 raising the possibility that massive transcriptional reprogramming is facilitated by ATXR2 during cell fate changes. Consistent with the fact that the CIM-induced callus resembles lateral root primordium,6 ATXR2 mainly controls molecular components implicated in lateral root formation. ATXR2 physically interacts with AUXIN RESPONSE FACTOR 7 (ARF7) and ARF19, and binds directly to the promoters of LATERAL ORGAN BOUNDARIES DOMAIN 16 (LBD16) and LBD29,5 which are involved in both lateral root development and callus formation.3^,7 The ATXR2-ARF complex catalyzes H3K36me3 deposition at the LBD promoters and stimulates their expression in order to stimulate callus formation on CIM. Consistently, atxr2 leaf explants displayed reduced callus formation, whereas overexpression of ATXR2 results in enhanced callus formation capability.5

To further support the role of ATXR2 in establishing lateral root identity, we also measured lateral root numbers in seedlings grown under normal growth conditions. Consistent with the strong implication of LBDs in lateral root formation,7 lateral root development was significantly reduced in atxr2-deficient mutants (Fig.1A), suggesting that ATXR2 is involved in lateral root initiation and development during normal plant growth.

Our results also raised the possibility that ATXR2 might be a general regulator of de novo root organogenesis. To test whether ATXR2 is indeed implicated in de novo root organogenesis, we employed a conventional hormone-free method that usually monitors adventitious root formation from leaf explants.8 Leaf explants were placed on hormone-free medium, and we counted frequency of adventitious roots emerged from wound sites. As expected, adventitious root formation was significantly reduced in atxr2 mutant leaf explants compared with wild-type leaf explants on hormone-free medium (Fig.1B).

Callus formation and adventitious roots from leaf explants share in part genetic pathways. Auxin plays a fundamental role in both processes,4^,9 and molecular components are also conserved, including ABERRANT LATERAL ROOT FORMATION 4 (ALF4), CURLY LEAF (CLF), LBD16, LBD29, and SWINGER (SWN), WUSCHEL RELATED HOMEOBOX 11 (WOX11), and WOX12.4^,10 ATXR2 may be another member accounting for conservation of the processes. Complex formation of ATXR2 with ARFs or WOXs would be flexible, depending on given environments, and ensures diverse repertories of root development. Taken together, ATXR2 might be an integrative regulator of de novo root organogenesis, which plays a role in the cell fate transition possibly during the initial stages (Fig.1C).