N6-Methyladenosine-Modified circSMAD4 Prevents Lumbar Instability Induced Cartilage Endplate Ossification

Lumbar instability causes cartilage endplate ossification and intervertebral disc degeneration. In this study, it is determined that circSMAD4, a Yap1-related circRNA, is stably downregulated under abnormal stress. In vitro, circSMAD4 knockdown resulted in Yap1 mRNA degradation, whereas circSMAD4 overexpression increased Yap1 mRNA expression and nuclear translocation. Hence, the stabilization of circSMAD4 is essential for maintaining the homeostasis of endplate cartilage under abnormal stress. Furthermore, transcriptome sequencing and mass spectrometry analysis revealed that METTL14-mediated N⁶-methyladenosine (m⁶A) modification can stabilize circSMAD4 expression. Moreover, circSMAD4 is shown to regulate Yap1 mRNA through the m6A reader IGF2BP1. The IGF2BP1 functions to translocate Yap1 mRNA into the nucleus, which protects endplate chondrocytes from degeneration. Finally, local injection of an AAV5-containing circSMAD4 overexpression plasmid successfully rescued LSI-induced cartilage endplate degeneration, which wasn't observed in Yap1 knockout mice. These findings suggest that m6A-modified circSMAD4 can stabilize Yap1 mRNA expression and translocation, thus preventing degeneration of the cartilage endplate under abnormal stress. Hence, circSMAD4 may become a potential therapeutic tool for managing instability-induced intervertebral disc degeneration.