Single Amino Acid Divergence Drives Functional Specialisation of Homologous GRFs in Populus
Haofei Wang, Jifan Zhang, Ruoting Wang, Shuxian Tan, Jiadong Wu, Zhen Cheng, Yuxin Du, Lin Liu, Deqiang Zhang, Yuchao Ma, Jianbo XieABSTRACT
Gene duplication with sub‐ and/or neo‐functionalization is a key pathway in plant adaptive evolution. The duplication‐degeneration‐complementation (DDC) model explains sub‐functionalization of paralogs, yet the relative contributions of coding sequence versus cis‐regulatory changes remain contested. In this study, we show that a single amino acid substitution (Met30/Ile30) in the WRC domain drives sub‐functionalization of poplar paralogs PagGRF29 and PagGRF10, demonstrating a key role for coding changes. Met30 confers high DNA‐binding capacity and represents the ancestral state, whereas Ile30 severely impairs DNA binding. Evolutionary analysis reveals residue 30 as a conserved tuning site, with the degenerative Ile30 variant emerging during plant terrestrialization. Despite impaired DNA binding, PagGRF10 physically interacts with PagGRF29 to form a complementary module: PagGRF29 alone enhances drought tolerance at growth expense, while the PagGRF29‐PagGRF10 complex promotes biomass accumulation with reduced stress resilience. Thus, the Met30/Ile30 switch drives the resolution of duplicated genes through DNA‐binding‐based functional specialisation and cooperative regulatory module formation, orchestrating the growth‐stress trade‐off. This pattern represents an extended manifestation of the DDC framework, combining asymmetric sub‐functionalization with interaction‐mediated functional innovation. Our study demonstrates how a single‐amino‐acid substitution can drive paralog specialisation, offering mechanistic insight into the evolutionary fates of duplicated genes in plants.