After being extracted from the frozen pulverized left ventricles or cultured cells and quantified, total proteins were prepared for western blot analysis and normalized to the matched total proteins or GAPDH according to our previous study52 (link). Briefly, separated proteins were incubated with the indicated primary antibodies overnight at 4 °C and with secondary antibodies for 60 min at room temperature. Nuclear and cytosolic protein fractions were separated using a commercial kit (Thermo Fisher Scientific) according to the manufacturer′s protocol. Proteins from cytosolic lysates were normalized to GAPDH, whereas proteins from nuclear lysates were normalized to PCNA.
Isolated total mRNA from hearts and cultured cells was reversely transcribed to complementary DNA (cDNA) with Transcriptor First Strand cDNA Synthesis Kit (Roche (Basel, Switzerland), 04896866001). Transcriptional level of target genes were normalized to Gapdh, and the primers for quantitative real-time PCR are shown in Supplementary Table3 .
Isolated total mRNA from hearts and cultured cells was reversely transcribed to complementary DNA (cDNA) with Transcriptor First Strand cDNA Synthesis Kit (Roche (Basel, Switzerland), 04896866001). Transcriptional level of target genes were normalized to Gapdh, and the primers for quantitative real-time PCR are shown in Supplementary Table
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