FIGURE 4.

Cloning and analysis of BBH1. (A) Co-immunoprecipitation (co-IP) assays were used to identify binding partners in mouse C₂C₁₂ myoblasts. Acheron binds to CASK, a previously identified partner, but not the pro-survival protein Bcl-2. It does bind to the BH3-only protein BAD, but not the related protein BAX. (B) Co-IP assays with day 18 ISM extracts demonstrate that Acheron binds to a protein that cross-reacts with an anti-mouse BAD antibody. (C) Expression of BBH1 protein across ISM development. Tubulin was used as a loading control. (D) RNA-seq quantification of BBH1 expression during development. There was no significant change (p = 0.28, n = 3 independent replicates for each stage of development). (E) The anti-BAD antibody recognizes a bacterially expressed portion of Manduca BBH1 protein in the induced (“I”) sample but not the uninduced sample (“U”).