Nengwen Huang, Yang Li, Jie Lu, Yifeng Xing, Weiping Chen, Kaidi Chen, Geyuan Zheng, Pengyuan Hu, Kaixun He, Hanyu Lin, Wenxiu Yuan, Yuwei Zhou, Zhenzhu Xue, Junjin Lin, Wen Li, Sihui Zhang, Yanjing Ou, Jiang Chen
Osteogenic differentiation requires sophisticated mitochondrial adaptation to meet bioenergetic demands, yet regulatory checkpoints governing this organelle reorganization remain poorly defined. Through single-cell RNA sequencing reanalysis and metabolic intervention, this study unveils a non-canonical signaling role for PFKM, traditionally recognized solely as a glycolytic enzyme, in orchestrating mitochondrial remodeling during bone formation. Beyond its established metabolic function, Pfkm suppression triggers distinctive donut-shaped mitochondria through a novel signaling cascade. Mechanistically, Pfkm knockdown expands mitochondria-endoplasmic reticulum contacts (MERCs), facilitating mitochondrial calcium influx. Concomitantly, elevated CD38 suppresses protein kinase A (PKA) activity, inducing DRP1 dephosphorylation at Serine 656. This signaling integration promotes DRP1 mitochondrial translocation, driving the characteristic donut architecture. This structural transformation initiates comprehensive mitochondrial quality control (MQC) encompassing enhanced biogenesis, selective mitophagy, and mitochondrial-derived vesicles (MDVs) secretion, collectively optimizing the osteogenic microenvironment and cellular mineralization capacity. In vivo validation demonstrates that AAV-mediated Pfkm knockdown accelerates bone repair in rat calvarial and femoral defect models. This work establishes PFKM as a dual-function regulator bridging metabolism and mitochondrial signaling, offering a potent therapeutic avenue for bone regeneration.