Abstract:Nuclear–mitochondrial communication is crucial for plant growth, particularly in the context of cytoplasmic male sterility (CMS) repair mechanisms linked to mitochondrial genome mutations. The restorer of fertility-like (RFL) genes, known for their role in CMS restoration, remain largely unexplored in plant development. In this study, we focused on the evolutionary relationship of RFL family genes in poplar specifically within the dioecious Salicaceae plants. PtoRFL30 was identified to be preferentially expressed in stem vasculature, suggesting a distinct correlation with vascular cambium development. Transgenic poplar plants overexpressing PtoRFL30 exhibited a profound inhibition of vascular cambial activity and xylem development. Conversely, RNA interference-mediated knockdown of PtoRFL30 led to increased wood formation. Importantly, we revealed that PtoRFL30 plays a crucial role in maintaining mitochondrial functional homeostasis. Treatment with mitochondrial activity inhibitors delayed wood development in PtoRFL30-RNAi transgenic plants. Further investigations unveiled significant variations in auxin accumulation levels within vascular tissues of PtoRFL30-transgenic plants. Wood development anomalies resulting from PtoRFL30 overexpression and knockdown were rectified by NAA and NPA treatments, respectively. Our findings underscore the essential role of the PtoRFL30-mediated mitochondrion-auxin signaling module in wood formation, shedding light on the intricate nucleus–organelle communication during secondary vascular development.