Fig 2: The L27 domain of MPP7 cooperates with TAZ and YY1 to enhance transcription.(A) Promoters of 4 common DEGs among Mpp7, Amot, YapTaz, and Yy1 cKO (Chen et al, 2019) data sets. Putative YY1 binding sites are red blocks and TEAD binding sites, blue blocks. (B) Location and sequence of the TEAD and YY1 binding sites are indicated in the Carm1 promoter. Carm1-reporter was co-transfected with constructs in x-axis to assess transcriptional activity; (−), empty expression construct. (C) Same as in (B), except that the YY1 binding site is mutated (top). (D) Same as in (B), except that the TEAD binding site was mutated (top). (E) Co-IP between TAZ and YY1 is enhanced by fusing MPP7’s L27 domain to TAZ in 293T cells. Constructs and tagged epitopes for detection are indicated; quantification of co-IPed HA-YY1 to the right. (F) Co-IP assay to determine the relative contributions of L27 and PDZ domains of MPP7 to its interaction with YY1 in 293T cells. Constructs and tagged epitopes for detection are indicated; quantification of co-IPed HA-YY1 to the right. (G) A model for TAZ-YY1 cooperation mediated by AMOT and MPP7. S175 of AMOT is subjected to phosphorylation and regulation by actin dynamics. Data information: (B–F) n = 3 biological replicates. Error bars represent means ± SD. One-way ANOVA with Tukey’s post hoc test was performed in (B–F). Source data are available online for this figure.

Fig 3: Chromatin binding and protein association support convergence of MPP7, YY1, and TAZ/YAP.(A) ChIP-qPCR shows MPP7, YY1, and TAZ binding to Carm1 promoter (diagramed a top) at site 1 (red) harboring both TEAD and YY1 binding sites, but not at a distal site 2 (blue) without either binding site; IgG, negative control. Bar graph shows fold-enrichment (by Bio-Rad CFX Maestro); keys to the left. (B) Same as in (A) with 6 more common DEG promoters (x-axis) with YY1 and TEAD binding sites; keys and quantification as in (A). (C) Representative PLA images for MPP7 and YAP/TAZ in 3D reconstruction and single plane, countered stained with DAPI; quantification to the right. (D) Same as in (C) for MPP7 and YY1.(E) Same as in (C) for YY1 and YAP/TAZ in Con (Pax7CreERT2/+) and Mpp7 cKO. (F) Same as in (E) for CARM1-PAX7; (G) Restoring CARM1-PAX7 PLA signal by L27-TAZ but not by empty vector (EV, with IRES-mGFP, same as (−) in previous figures). Each data point in (C–G) represents PLA dots in a cell quantified by 3D imaging. Single Ab controls are shown in Appendix Fig. S8C. Data information: Scale bars = 5 µm (C–G). (A, B) n = 3 biological replicates. (C) 5 MuSCs for each group; (D) 5 MuSCs for MPP7 or YY1 and 8 MuSCs for MPP7-YY1; (E) 5 MuSCs for Con (Pax7CreERT2/+) and 7 MuSCs for Mpp7 cKO; (F) 15 MuSCs for Con and 16 MuSCs for Mpp7 cKO; (G) 11 MuSCs for EV and 19 MuSCs for L27-Taz. Error bars represent means ± SD. One-way ANOVA with Tukey’s post hoc test in (A–D). Student’s t test (two-sided) in (E–G). Source data are available online for this figure.

Fig 4: Mpp7/Amot regulatory network intersects with that of Yap/Taz.(A) H&E histology of Yap cKO and YapTaz cKO muscles at 5 dpi (Con (Pax7CreERT2/+) histology not included); quantifications of regenerated myofiber cross-sectional area to the right. (B) Venn diagram shows overlapping DEGs between the YapTaz cKO and the Mpp7 cKO. (C, D) IF images of TAZ (C) and YAP (D) in FACS-isolated and cultured Con (Pax7CreERT2/+), Mpp7 cKO, and Amot cKO MuSCs at 48 h; relative fluorescent signals (in AU) to the right. (E) Co-IP of V5-TAZ and HA-AMOT by FLAG-MPP7 expressed in 293 T cells; tagged epitopes used for IP and Western blotting are indicated; (−), empty expression construct. Quantification of relative levels of co-IPed V5-TAZ is to the right. (F) Relative TEAD-reporter (8XGTIIC-luciferase, depicted at top) activities when co-transfected with V5-Taz, Flag-Mpp7, and/or Ha-Amot constructs in 293T cells; (−), empty expression construct. (G) Relative activities of WT and TEAD-binding site mutated (MUT) Carm1-reporters co-transfected with V5-Taz or Flag-Mpp7 constructs; (−), empty expression construct. (H) Relative IF signals (AU) of CARM1 in Mpp7 cKO MuSCs transfected with empty vector (−), V5-Taz WT and V5-Taz S89A constructs (all with IRES-mGFP) (I) Relative cell fate fractions among Con (Pax7CreERT2/+), Mpp7 cKO, and Mpp7 cKO MuSCs transfected with V5-Taz S89A and V5-Taz WWm construct in single myofiber culture; Con (−) and Mpp7 cKO (−) from Fig. 2C; keys at the top. Data information: Scale bar = 50 µm in (A) and 25 µm in (C, D). (A) N = 5 mice in each group; (C, D, H) 200 cells from 2 to 3 mice in each group; (E–G) n = 3 biological experiments; (I) ≥ 150 cells in each group. Bars represent medians ± 95% CI (C, D, H) or means ± SD (A, E, F, G). Hypergeometric test was used in (B), one-way ANOVA with Tukey’s post hoc test performed in (A, E, F, G), Kruskal–Wallis test followed by Dunn’s multiple comparisons test in (C, D, H), and Chi-square test used in (I). Source data are available online for this figure.