Fig. 5

circCacna1c binds to Hnrnpf and inhibits nuclear translocation of Hnrnpf. A Gene Ontology (GO) analysis of proteins captured by the circCacna1c probe. B Prediction of binding of circCacna1c to Hnrnpf. C The Hnrnpf antibody was utilized in the RIP assay to determine its affinity for circCacna1c. Subsequently, western blotting analysis was performed to confirm the binding specificity of the Hnrnpf antibody, and qRT-PCR was employed to assess the level of circCacna1c enrichment facilitated by the Hnrnpf antibody in H9c2 cells. **P < 0.01. n = 3. D RNA pulldown was used to detect the binding of Hnrnpf to circCacna1c. n = 3. E H9c2 cells were transfected with the circCacna1c expression vector, followed by the determination of Hnrnpf protein levels. n = 3. F The circCacna1c expression vector was transfected into H9c2 cells, which were then exposed to 500 μM H₂O₂ for a period of 6 h. Following this, the cytoplasmic and nuclear fractions were separated and the protein quantity of Hnrnpf was measured. Lamin B and β-tubulin acted as controls for nuclear and cytoplasmic compartments respectively, with their relative protein levels being determined. *P < 0.05, **P < 0.01. n = 3. G The H9c2 cells were transfected with a plasmid that overexpressed Hnrnpf, followed by the determination of Hnrnpf protein levels. GAPDH was selected as a reference. n = 3. H, I H9c2 cells were transfected with circCacna1c expression vectors and Hnrnpf expression vectors, then exposed to 500 μM H₂O₂ for a period of 6 h. The PI assay (H) and LDH activity assay (I) were employed to assess cell necroptosis in H9c2 cells. ***P < 0.001. n = 3