fig1

Figure 1. Activation or suppression of the cWNT/β-catenin pathway in the postmitotic cardiac myocytes. (A) RT-PCR data showing the transcript levels of Ctnnb1 in the β-catenin gain-of-function (GoF) and loss-of-function (LoF) cardiac myocytes compared to wild type and Myh6-Mcm myocytes using primers designed to specifically amplify the exon 3, exon 3-4, exon 9, and exon 12-13 of the Ctnnb1 gene. (B) IGV Genome browser images of read counts of Ctnnb1 transcript levels obtained from the RNA-Seq data of wild type, Myh6-Mcm, β-catenin GoF, and LoF cardiac myocytes. (C) Western blot showing expression of the CTNNB1 protein in the cardiac myocyte protein extracts from wild type (WT), Myh6-Mcm, Myh6-Mcm:Ctnnb1^LoF, and Myh6-Mcm:Ctnnb1^GoF mice, detected using antibodies raised against N- or C-terminal region of the β-catenin protein. The lower panel represents an immunoblot showing the expression of vinculin protein, which was used as the corresponding loading control. (D) Graphs depicting quantitative data of the CTNNB1 protein levels in the wild type, Myh6-Mcm, Myh6-Mcm:Ctnnb1^LoF, and Myh6-Mcm:Ctnnb1^GoF cardiac myocyte protein extracts normalized to the vinculin protein levels (n = 3). (E) Representative immunofluorescence panels showing expression and localization of the CTNNB1 protein to the intercalated disks in the experimental groups. (F) Nuclear localization of CTNNB1 as detected by staining of thin myocardial sections with an anti CTNNB1 antibody. The number of nuclear stained positive for the CTNNB1 as well as the intensity of the fluorescence signaling were increased in the GoF mouse hearts, whereas the number of nuclei expressing CTNNB1 was not significantly changed in the LoF mouse hearts, albeit the signal intensity of the CTNNB1 was weaker. (G) Quantitative data showing the number of nuclei staining positive for the expression of CTNNB1. RT-PCR: Reverse transcription-polymerase chain reaction; Myh6-Mcm: myosin heavy chain 6-MerCreMer; RNA-Seq: RNA-Sequencing.