Fig. 4

FEACR interacts with NAMPT and maintains its stability. a Pull-down assay was performed by using biotinylated FEACR (Bio-FEACR) or negative control RNA (Bio-NC) in cardiomyocytes, and then separated protein molecules by electrophoresis and developed by Coomassie blue staining, proteins from the specific band was used for protein mass spectrometry (MS) identification. b NAMPT was identified by LC–MS/MS in the group of biotinylated FEACR. c RNA pull-down assay was performed to confirm the NAMPT binding to FEACR by using bio-FEACR or bio-NC. d RIP assay was performed by using antibodies against NAMPT to confirm the binding of FEACR to NAMPT by qPCR. e 100 μg/mL CHX was used to treat FEACR overexpressed cardiomyocytes at the indicated time. The expression of NAMPT was measured by Western blot. f Relative NAMPT protein levels were quantified with Image J. g Expression levels of NAMPT mRNA in cardiomyocytes transfected with si-NC or si-FEACR. h The expression of NAMPT protein in cardiomyocytes transfected with si-NC or si-FEACR. a, c n = 3 biological replicates; d n = 6 biological replicates; e, f n = 3 biological replicates; g n = 6 biological replicates; h n = 3 biological replicates. Data were mean ± SD, and the P-value was calculated by One-way ANOVA (g, h) or student’s t-test (d, f). The experiment technically repeats three times