LncRNA GACAT2 binding protein PKM1/2 reverses inflammation-compromised mitochondrial function and cementoblastic differentiation of periodontal ligament stem cells

This study investigated the role of mitochondrial function and the downstream long noncoding RNAs (lncRNAs) in regulating inflammation-induced changes to the cementoblastic differentiation of PDLSCs. We found that inflammatory cytokine-caused impairment to cementoblastic differentiation of PDLSCs was closely correlated to their mitochondrial function, and in terms of lncRNA microarray analysis and gain/loss-of-function studies, gastric cancer associated transcript 2 (GACAT2) was identified as a regulator across the cellular events of both inflammation-mediated mitochondrial function and cementoblastic differentiation. Next, comprehensive identification of RNA-binding proteins by mass spectrometry (ChIRP-MS) and parallel reaction monitoring (PRM) assay revealed that GACAT2 was directly binded to protein pyruvate kinase M1/2 (PKM1/2). Further functional studies demonstrated that GACAT2 overexpression increased cellular protein expressions of PKM1/2, PKM2 tetramer and phosphorylated PKM2, led to an enhanced pyruvate kinase (PK) activity along with increased translocation of PKM2 into mitochondria. Finally, we found that GACAT2 overexpression could reverse inflammation-compromised mitochondrial function and cementoblastic differentiation of PDLSCs, and this function could be abolished by PKM1/2 knockdown. Our data suggest that lncRNA GACAT2 plays a critical role in inflammation-compromised mitochondrial function and cementoblastic differentiation by binding protein PKM1/2. PDLSCs were incubated in normal α-MEM medium (Nor), medium with the presence of inflammatory cytokines TNF-α plus IL-1β (Infla), medium with the presence of cementoblastic inducer EMD (EMD), or medium with the presence of both inflammatory cytokines and EMD (Infla-EMD)