Plants have the unique ability to regenerate completely from a somatic cell, that is, an ordinary cell that is not normally involved in reproduction. This process includes: new It hits the apical meristem (SAM) formation, which gives rise to the lateral organs that are key in remodeling the (or new) plant.

At the cellular scale, SAM formation is carefully controlled by positive or negative regulators (genes/protein molecules) that can trigger or limit shoot regeneration, respectively. So which molecules are we talking about? Are there other regulatory levels not yet covered?

To find answers to the above questions, a research team led by Japan’s Nara Institute of Science and Technology (NAIST) studied the process. ArabidopsisA plant widely used in genetic research.

Their work has been published in the journal: Science Advancesidentified and characterized an important negative regulator of shoot regeneration. They showed how WUSCHEL related gene HOMEOBOX 13 (WOX13) and its protein may contribute to the non-meristematic (non-dividing) function of callus cells, acting as a transcriptional repressor (at the RNA level) and thereby affecting regeneration efficiency.

“The search for strategies to increase the efficiency of shoot regeneration in plants has been ongoing for a long time. However, progress has been hampered by a lack of clarity regarding the regulatory mechanisms involved. Our study fills this gap by identifying a new pathway for cell fate determination,” explains Momoko Ikeuchi, principal investigator of the study.

His team’s previous research had already established a role WOX13 Tissue healing and organ adhesion after transplantation. Thus, they tested for the first time the potential role of this gene in the control of shoot regeneration in the mutant. wox13 Arabidopsis (non-functional facility WOX13 ), using a two-stage tissue culture system.

Phenotypic and image analysis showed that shoot regeneration was accelerated (3 days faster) in plants. WOX13 and the expression is slower when stimulated WOX13. In addition, in normal plants WOX13 It showed locally decreased expression levels in SAM. These results show that WOX13 It can negatively regulate shoot regeneration.

To confirm their findings, the researchers compared the mutants. wox13 and wild-type (normal) plants by RNA sequencing at multiple time points. Absence WOX13 did not significantly alter gene expression Arabidopsis in callus-forming conditions. But the conditions that triggered the exile greatly increased the changes caused by the mutation. wox13this led to increased regulation of shoot meristem regulatory genes.

Interestingly, these genes were repressed within 24 hours after overexpression. WOX13 in mutant plants. In general, they found WOX13 It directly activates cell wall-modifying genes involved in cell expansion and differentiation, while suppressing a subset of shoot meristem regulators. Next Quartz-Seq2-based single-cell RNA sequencing (scRNA-seq) confirms key role WOX13 in determining the fate of pluripotent callus cells.

This study showed that unlike other known negative regulators of shoot regeneration, which only inhibits the transition from callus to SAM, WOX13 It suppresses the SAM specification, promoting the acquisition of alternative fates. It achieves this inhibition through a mutually suppressive regulatory circuit with the regulator. WUSpromoting a non-meristematic cell fate through transcriptional inhibition WUS and induction of other SAM modifiers and cell wall modifiers.

For this reason, WOX13 It acts as the main regulator of regeneration efficiency. “Our results show that there are knockouts WOX13 It can promote the exile fate and increase the efficiency of the exile arrangement. That means knockout WOX13 It can serve as a tool in agriculture and horticulture and promote tissue culture-mediated de novo shoot regeneration in crops,” concludes Ikeuchi.