Light-controlled morphogenesis is crucial for the successful emergence, survival and trophic transition of seedlings. Over the past three decades, proteolysis mediated by the E3 ubiquitin ligase COP1 (CONSTITUTIVELY PHOTOMORPHOGENIC 1) has been established as a central regulatory mechanism of seedling photomorphogenesis. Recently, the research team led by Xing Wang Deng (Shandong Laboratory of Advanced Agricultural Sciences in Weifang/ Peking University Institute of Advanced Agricultural Sciences/State Key Laboratory of Wheat Improvement) in collaboration with Associate Professor Jian Li (College of Life Sciences, Nanjing Normal University), found that COP1-mediated lysine 63 (K63)-linked polyubiquitination inhibits the enzyme activity of GRETCHEN HAGEN 3.5 (GH3.5) in the dark, thereby promoting seedling hypocotyl elongation. This work, entitled “Inactivation of GH3.5 by COP1-mediated K63-linked ubiquitination promotes seedling hypocotyl elongation”, was published in Nature Communications on April 14, 2025.

 

 

Ubiquitination is a highly conserved post-translational modification in eukaryotes and is involved in numerous biological processes, including protein degradation, DNA damage response and cell cycle regulation. A ubiquitin protein contains seven lysine (K) residues. Ubiquitin chains linked via different lysines have different effects on substrate proteins. K48-linked ubiquitination is the most common ubiquitin modification type. Proteins labeled with K48-linked chains are usually directed to the 26S proteasome for degradation. K63 ubiquitination is the second most common type. Rather than triggering protein degradation, K63-linked chains usually regulate protein interaction, subcellular localization, and so on. It is worth noting that although there are more than 1000 E3s in higher plants, only a single-digit number of E3s have been found to catalyze the formation of K63 chains.

 

At the beginning of this study, the authors demonstrated through genetic and biochemical experiments that COP1 is involved in the formation of K63-linked ubiquitin chains in Arabidopsis. Then, two types of immunoprecipitation-mass spectrometry (IP-MS) were performed to screen potential substrates for COP1-mediated K63 ubiquitination. They used K63 TUBE (K63 tandem ubiquitin binding entity) beads to capture proteins whose K63 ubiquitination was dependent on COP1. At the same time, they used conventional IP-MS to screen COP1-interacting proteins. Using this strategy, the authors identified GH3.5 as one of the substrate candidates for COP1-mediated K63 ubiquitination. GH3.5 belongs to the auxin acyl amido synthetase family, whose members can inactivate IAA by conjugating IAA with aspartic acid (Asp), thereby inhibiting hypocotyl elongation. This study showed that COP1 attached K63 ubiquitin chains to GH3.5 in the dark and inhibited the enzymatic activity of GH3.5, instead of regulating its protein stability, thereby maintaining IAA activity and promoting hypocotyl elongation. In addition, light signaling was able to suppress the inhibitory effect of COP1 on the function of GH3.5 and its homologs, thereby inactivating IAA and inhibiting hypocotyl elongation (Fig. 1). In summary, the study reveals a new function of COP1-mediated ubiquitination beyond proteolysis, sheds light on the mechanism by which light regulates auxin activity, and provides a new insight into the regulation of plant morphogenesis by light-controlled auxin metabolism.

Fig. 1 COP1-mediated K63 ubiquitination suppresses GH3.5 activity and thus promotes seedling hypocotyl elongation

 

The first author is Dr. Yongting Liu (Assistant research scientist of Shandong Laboratory of Advanced Agricultural Sciences in Weifang/Peking University Institute of Advanced Agricultural Sciences). Professor Xing Wang Deng (Shandong Laboratory of Advanced Agricultural Sciences in Weifang/ Peking University Institute of Advanced Agricultural Sciences/State Key Laboratory of Wheat Improvement) and Associate Professor Jian Li (Nanjing Normal University) are the co-corresponding authors. Prof. Yinpeng Xie (Northwest A&F University) and Prof. Dongqing Xu (Nanjing Agricultural University) also contributed to the study. This work was supported by grants from the National Key R&D Program of China, the National Natural Science Foundation of China, the Shandong Provincial Natural Science Foundation, the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD), the Southern University of Science and Technology, and the Shenzhen Science and Technology Program.

 

Article link: https://doi.org/10.1038/s41467-025-58767-6