# Unveiling the GA4-Ferulic Acid Regulatory Axis: Redox-Mediated Suberization Governs Adventitious Rooting Recalcitrance in Pinus massoniana

## TL;DR

The study identifies a new regulatory pathway involving GA4 and ferulic acid that affects root development in Pinus massoniana, offering a solution to improve rooting success in conifers.

## Contribution

The discovery of the GA4-ferulic acid regulatory axis and its role in controlling suberization and root development in conifers is novel.

## Key findings

- PBZ treatment significantly improved rooting success and reduced rooting time in Pinus massoniana.
- GA4 accumulation triggered suberization via ferulic acid-mediated redox imbalance, impairing root development.
- The combination of PBZ and FA synergistically enhanced rooting success and field survival in conifers.

## Abstract

Pinus massoniana, a critically important afforestation species in subtropical China, shows severe adventitious rooting recalcitrance linked to endogenous gibberellin (GA) dysregulation. Our study reveals a GA4-mediated regulatory network that coordinates hormonal crosstalk, redox homeostasis, and cell wall remodeling. Treatment with the GA biosynthesis inhibitor paclobutrazol (PBZ, 100 mg·L−1) shortened rooting time by 32.5% and increased rooting success by 79.5%. We found that PBZ redirected GA flux by upregulating GA3-oxidase (GA3OX), leading to GA4 accumulation. However, elevated GA4 levels impaired root development by triggering suberization through ferulic acid (FA)-mediated redox imbalance. Application of GA4 (100 mg·L−1) reduced caffeoyl alcohol content by 54.4% but increased FA and caffeic acid levels 2.4–3.9-fold, shifting lignin precursors toward suberin biosynthesis. FA modulated H2O2 flux in a dose-dependent manner: 200 mg·L−1 optimized redox homeostasis (93.7% lower H2O2 influx), whereas 1000 mg·L−1 suppressed mitosis. The combination of PBZ (100 mg·L−1) and FA (200 mg·L−1) synergistically enhanced rooting success by 34.4% and achieved 95.8% field survival after two years (vs. 68.5% in controls), challenging the traditional view that lignification alone limits rooting in woody plants. This work provides the first evidence that the GA4-FA axis controls adventitious root formation in conifers via a Reactive oxygen species (ROS)-dependent switch between suberin and lignin metabolism, offering new strategies to overcome rooting barriers. The PBZ + FA protocol enables scalable clonal propagation of recalcitrant conifers, with potential applications in molecular breeding and forest restoration.