Nuclear poly(A) binding protein 1 (PABPN1) mRNA is unstable in muscle tissuebut stable in cultured myoblasts. (A) Total RNA was collected atdifferent timepoints after injection of actinomycin D to inhibit transcriptionand PABPN1 mRNA decay was analyzed by northern blot. Time courses are shown forsamples from muscle and kidney. Peroxisome proliferator-activated receptorgamma coactivator 1α (PGC1α) and glyceraldehyde 3-phosphatedehydrogenase (GAPDH), a known unstable and stable transcript, respectively,were probed as controls (n = 3 per timepoint). To visualize PABPN1 signal inmuscle samples, the blot was exposed significantly longer than for kidneysamples. (B) Total RNA was obtained from skeletal muscle (SM) andcultured primary mouse myoblasts (Mb) and PABPN1 mRNA levels were determinedusing real-time polymerase chain reaction (PCR) and hypoxanthine-guaninephosphoribosyltransferase (HPRT) mRNA was used as an internal control. Theamount of PABPN1 mRNA relative to skeletal muscle (SM) is shown; n = 3independent samples. Data are mean ± SD; *P <0.05 vs skeletalmuscle. (C) Protein extracts were prepared from SM and Mb andimmunoblotted with anti-PABPN1 antibody. HSP90 was used as a loading control.The immunoblot is representative of at least three independent samples.(D) Total RNA was collected at different timepoints after treatmentof cultured primary mouse myoblasts with actinomycin D and PABPN1 mRNA decaywas analyzed by northern blot; c-Myc and GAPDH, known unstable and stabletranscripts in myoblasts, respectively, were probed as controls. Averages ofdensitometric measurements of northern blot bands were used to determine mRNAdecay. The image is representative of at least three independent samples.(E) The decay profile of PABPN1 mRNA in muscle, kidney and culturedmyoblasts plotted as mRNA amount relative to timepoint T = 0 h (n = 3 samplesper timepoint). Data are mean ± SD.