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. 2023 Feb 8;11:1130058. doi: 10.3389/fcell.2023.1130058

FIGURE 3.

FIGURE 3

Harmonin overexpression leads to the decrease in Wnt signaling activity (A) Representative Western blot analysis of β-catenin (β-cat) expression in HEK293T cells transfected with harmonin a1-GFP (Harm-a1), truncated harmonin a1_R31*-GFP (Harm-R31*), or GFP (control) and β-catenin (β-cat). (B) Quantification reveals that the expression of full length harmonin a1 significantly decreases β-cat_K49/β-cat ratio (−0.2) while the expression of truncated harmonin a1-R31* did not alter the β-cat_K49/β-cat ratio when compared to control transfected HEK293T cells; 12% polyacrylamide gel. Two-tailed Student’s t-test, **p ≤ 0.01; N = 5 experiments. (C) Wnt response in transfected HEK293T cells measured by TCF/LEF luciferase activity assay. In unstimulated condition (control, left) luciferase activity is significantly decreased in Harm-a1 and Harm-R31* transfected cells compared to empty vector control. Stimulation with Wnt conditioned medium leads to an increase in luciferase activity for all, whereby control as well as Harm-R31* show significantly more cWnt signaling activity than Harm-a1 (Wnt, middle). Addition of β-cat results in the highest Wnt activity and reveals the largest difference of luciferase activity between Harm-a1 to control and Harm-R31* (β-cat, right). cWnt signaling is significantly decreased in Harm-a1 transfected cells. Two-tailed Student’s t-test, **p ≤ 0.01, **p ≤ 0.001; N = 3 experiments (three triplicates each condition).