Periodontal ligament stem cells (PDLSCs) are a key cell type for restoring/regenerating lost/damaged periodontal tissues, including alveolar bone, periodontal ligament and root cementum, the latter of which is important for regaining tooth function. However, PDLSCs residing in an inflammatory environment generally exhibit compromised functions, as demonstrated by an impaired ability to differentiate into cementoblasts, which are responsible for regrowing the cementum. This study investigated the role of mitochondrial function and downstream long noncoding RNAs (lncRNAs) in regulating inflammation-induced changes in the cementogenesis of PDLSCs. We found that the inflammatory cytokine-induced impairment of the cementogenesis of PDLSCs was closely correlated with their mitochondrial function, and lncRNA microarray analysis and gain/loss-of-function studies identified GACAT2 as a regulator of the cellular events involved in inflammation-mediated mitochondrial function and cementogenesis. Subsequently, a comprehensive identification of RNA-binding proteins by mass spectrometry (ChIRP-MS) and parallel reaction monitoring (PRM) assays revealed that GACAT2 could directly bind to pyruvate kinase M1/2 (PKM1/2), a protein correlated with mitochondrial function. Further functional studies demonstrated that GACAT2 overexpression increased the cellular protein expression of PKM1/2, the PKM2 tetramer and phosphorylated PKM2, which led to enhanced pyruvate kinase (PK) activity and increased translocation of PKM2 into mitochondria. We then found that GACAT2 overexpression could reverse the damage to mitochondrial function and cementoblastic differentiation of PDLSCs induced by inflammation and that this effect could be abolished by PKM1/2 knockdown. Our data indicated that by binding to PKM1/2 proteins, the lncRNA GACAT2 plays a critical role in regulating mitochondrial function and cementogenesis in an inflammatory environment.

牙周膜干细胞（PDLSCs）是修复/再生丢失/受损牙周组织的关键细胞类型，包括牙槽骨、牙周膜和牙骨质，后者对于恢复牙齿功能很重要。然而，存在于炎症环境中的 PDLSCs 通常表现出受损的功能，正如其分化为负责牙骨质再生的成牙骨质细胞的能力受损所证明的那样。本研究调查了线粒体功能和下游长链非编码 RNA (lncRNA) 在调节炎症诱导的 PDLSCs 牙骨质发生变化中的作用。我们发现炎症细胞因子诱导的 PDLSCs 牙骨质形成障碍与其线粒体功能密切相关，GACAT2作为参与炎症介导的线粒体功能和牙骨质形成的细胞事件的调节剂。随后，通过质谱 (ChIRP-MS) 和平行反应监测 (PRM) 分析对 RNA 结合蛋白的全面鉴定表明GACAT2可以直接与丙酮酸激酶 M1/2 (PKM1/2) 结合，这是一种与线粒体功能相关的蛋白质. 进一步的功能研究表明，GACAT2过表达增加了 PKM1/2、PKM2 四聚体和磷酸化 PKM2 的细胞蛋白表达，这导致丙酮酸激酶 (PK) 活性增强并增加了 PKM2 易位到线粒体中。然后我们发现GACAT2过表达可以逆转炎症诱导的PDLSCs对线粒体功能和成牙骨质细胞分化的损害，并且这种作用可以通过PKM1 / 2敲低而消除。我们的数据表明，通过与 PKM1/2 蛋白结合，lncRNA GACAT2在炎症环境中调节线粒体功能和牙骨质形成中起关键作用。