In nature, mechanical stress caused by various biotic and abiotic factors often leads to partial or even complete loss of organs and tissues in living organisms. Both plants and animals have the ability to repair tissues and regenerate organs in response to damage. Compared to animals, plants with fixed growth are more susceptible to mechanical damage. However, plants have developed an unparalleled ability to cope with damage in animals over a long period of evolution. That is to say, in the face of constantly present and unpredictable mechanical damage, plants can quickly activate defense responses to avoid pest infestation and wound infections; Faced with varying degrees of mechanical damage, plants are capable of tissue repair and organ and even whole life regeneration.

Tomatoes are a classic model plant for studying plant injury responses. In this mode system, Clarence Ryan discovered the systematic defense of plants against mechanical damage and insect invasion in 1972. In the early 1990s, Ryan's research team discovered that systemic hormones and jasmonic acid regulate plant systemic defense responses through a common signaling pathway, and established a basic working model in the field of plant systemic defense.

Compared to others, little is known about the primary signals and transduction mechanisms of plant regeneration caused by damage. As is well known, cell damage is the primary physical trigger that triggers the regeneration process of living organisms. Scientists speculate that signaling molecules induced by cell damage should play a regulatory role in tissue repair and organ regeneration processes. However, since scientists discovered the phenomenon of regeneration in 1740, the chemical nature of signaling molecules has been unclear. In addition, the regenerative ability of plants varies greatly and varies depending on genotype, which limits the potential application of transgenic and gene editing technologies in biological breeding.

On May 22, the research team of Li Chuanyou, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, published a research paper entitled Peptide REF1 is a local wound signal promoting plant regeneration in Cell, which identified the original injury signal molecule inducing plant regeneration - regeneration factor REF1 for the first time, revealed the signal transduction network of REF1 regulating tissue repair and organ regeneration, and proved the application value of REF1 in plant transgenic and gene editing fields.

The Li Chuanyou team has been using tomatoes as a model plant for a long time and using genetic methods to analyze the plant's systemic defense signaling pathway regulated by both systemic hormones and jasmonic acid. In the early stage, the team obtained a series of SPR mutants with changes in the systematic signaling pathway through large-scale genetic screening. In order to use genetic methods to analyze the process of plant regeneration, the team creatively proposed the scientific concept that plant injury response actually includes two inseparable and interactive physiological processes, defense and regeneration. Based on this concept, researchers infer that tomato mutants related to regeneration should exhibit some degree of deficiency in defense response. Therefore, this study started with analyzing the defense deficient mutants accumulated in the laboratory and identified the tomato mutant spr9 with both defense and regeneration defects simultaneously. Of particular importance, The local defense response and systemic defense response of SPR9 are both defective, and the ability to form callus and regenerate organs is lost, indicating that SPR9 plays a role in the defense response and regeneration response induced by injury.



The gene cloning results indicate that, SPR9 encodes the precursor protein of the small peptide SlPep. Knockout of SPR9 results in the loss of injury induced callus formation and organ regeneration ability in tomatoes, while overexpression of SPR9 can enhance tomato regeneration ability. In addition, exogenous application of small peptides encoded by SPR9 can also improve the regeneration ability of tomatoes. Therefore, researchers renamed the small peptide as the regeneration factor REF1.



This study confirms that the receptor kinase PORK1, which is rich in leucine repeat sequences, is a receptor for REF1. When plant cells are damaged, REF1, as a primary injury signaling molecule, is recognized by receptor PORK1 and activates the expression of downstream cell reprogramming key regulatory factor SlWIND1, thereby initiating tissue repair and organ regeneration processes. Meanwhile, SlWIND1 binds to the promoter region of the REF1 precursor gene to activate its expression, thereby producing more REF1 peptides and amplifying the REF1 signal. Therefore, REF1 regulates the regeneration process through the action of plant cytokines.



Of particular importance, the role of the regeneration factor REF1 is conserved in the plant kingdom. Corresponding REF1 peptides and their receptors can be found in almost all dicotyledonous and monocotyledonous plants. External application of REF1 can improve the regeneration ability and genetic transformation efficiency of tomatoes, as well as enhance the regeneration ability and genetic transformation efficiency of crops recognized as difficult to transform, such as soybeans, wheat, and corn. The relevant method has been applied for an international PCT patent.



The above achievements are the development of research on the mechanism of plant injury response. This study identified the primary injury signaling molecule REF1 that induces plant regeneration, solving the challenges that have plagued the scientific community and providing a convenient and universal solution to the problems of low genetic transformation efficiency and severe dependence on species and genotypes in crop breeding practices.



The research work has received support from the National Natural Science Foundation of China and the National Key Research and Development Program.





REF1 regulates the regeneration process through the action of plant cytokines