MiR156-dependent and miR156-independent effects of the RNA-binding protein AtGRP7 on vegetative phase change in Arabidopsis

Abstract Background The life cycle of higher plants consists of a vegetative and a reproductive phase. The vegetative phase is further divided into a juvenile and an adult phase. Precise timing of the vegetative phase change, the transition from juvenile to adult vegetative phase, is crucial for plant fitness. A master regulator of vegetative phase change is the highly-conserved microRNA miR156. RNA-binding proteins (RBPs) are central regulators of post-transcriptional gene regulation and developmental plasticity, yet for many RBPs their direct RNA targets and the developmental relevance of these interactions remain incompletely understood. Results Here, we identify the glycine-rich RBP At GRP7 as a regulator of vegetative phase change in Arabidopsis thaliana . Plants overexpressing At GRP7 ( At GRP7ox) exhibited a prolonged juvenile vegetative phase with a reduced leaf emergence rate. Nevertheless, they flowered earlier than wild-type plants. In contrast, the grp7-1 loss-of-function mutant transitioned earlier to the adult vegetative phase without changes in leaf emergence rate but displayed delayed flowering. Additional loss of the paralog At GRP8 in grp7-1 grp8-2 plants did not further accelerate phase change, indicating a predominant role of At GRP7 in this process. To systematically identify pri-miRNA targets of At GRP7, we applied an adapted iCLIP2 workflow and detected in vivo binding of At GRP7 to 44 pri-miRNAs, including MIR156A and MIR156C . At GRP7 altered the abundance of MIR156A and MIR156C precursors, and expression of a miR156 target mimic (MIM156) partially suppressed the prolonged juvenile phase of At GRP7ox plants, indicating that altered miR156 activity contributes to the observed developmental phenotype. Expression of miR156-responsive SPL transcription factors was affected only modestly, suggesting that the developmental effects of At GRP7 cannot be explained solely by regulation of the canonical miR156-SPL pathway. Conclusion Together, our findings identified At GRP7 as a regulator of vegetative phase change and established an adapted iCLIP2 workflow for identifying At GRP7-bound pri-miRNAs in vivo. Our data indicate that altered miR156 activity contributes to the developmental phenotype of At GRP7ox, while additional post-transcriptional mechanisms are likely involved. These findings extend the established role of At GRP7 in flowering time regulation and reveal a novel function in coordinating vegetative and reproductive development.