The interaction protein of SORBS2 in myocardial tissue to find out the pathogenic mechanism of LVNC disease

Figures

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  Figure 1. Proteins interacting with SORBS2 were screened by IP experiment. (A) IP experiment screened the signal pathways that were different from SORBS2. (B) The Venn diagram shows the difference between the two groups of IgG and IP. (C) Proteins interacting with SORBS2 were screened by bioinformatics database. The main enrichment pathway of the proteins interacting with SORBS2 was screened by IP experiments.

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  Figure 2. Proteins that interact with SORBS2. (A) The proteins interacting with SORBS2 were screened according to the evaluation score and reliability analysis. We focused on YWHAQ interacting with SORBS2.

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  Figure 3. Stem cells are induced to differentiate into cardiomyocytes. (A) Human embryonic stem cells (hESC) expressing Oct4, Sox2, Nanog and SSEA. (B) hESC-derived cardiomyocytes (hESC-CMs) expressing MLC-2v, α-actinin, cardiac troponin-I (cTnI) and cardiac troponin-T (cTnT). Scale bars=25μm.

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  Figure 4. SORBS2 interacts with YWHAQ. (A) To better understand β-tubulin, ACTN2, CLTL and YWHAQ, we next performed studies in hESC-derived cardiomyocytes (hESC-CMs). Immunofluorescence revealed a clear colocalization of SORBS2 with β-tubulin, ACTN2, CLTL and YWHAQ in the hESC-CMs. The SORBS2 protein (red) colocalized with β-tubulin (green), and the merged images are shown in yellow. (B) Extracted proteins from the cardiomyocytes ware untreated (input) or precipitated by an anti-Flag or anti-Myc antibody and then separated by 4-12% SDS-PAGE. Co-immunoprecipitation (Co-IP) of the heart tissue showing SORBS2 binding with YWHAQ.

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  Figure 5. Expression of SORBS2 in hESC-CMs transfected with HBLV-h-SORBS2 was higher compared with the control groups. (A) Cardiomyocytes transfected with lentivirus. (B) Representative images of SORBS2 in hESC-CMs transfected with empty virus (control) and HBLV-h-SORBS2 (SORBS2-OE). (C) Western blotting detection of the expression levels of SORBS2 in overexpression group and control group. The protein level of SORBS2 was normalized to GAPDH for each sample. Statistical analysis of the expression of SORBS2 in hESC-CMs transfected with empty virus (control) and HBLV-h-SORBS2 (SORBS2-OE) (n =3 per group, * p < 0.05; Student's t-test). Data are shown as the mean ± SEM.

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  Figure 6. YWHAQ protein decreased in the group that overexpressed SORBS2. (A) The expression of YWHAQ inf immunoblots of hESC-derived cardiomyocytes (hESC-CMs). (B) Statistical analysis of the expression of YWHAQ in hESC-CMs transfected with HBLV-h-SORBS2 and control cells. (* *p < 0.01).