Exogenous ANO5 expression rescues Ano5^−/− MPC fusion. (a) Representative images of MPC fusion after ∼36 h differentiation for Ano5^+/+, Ano5^−/−, and Ano5^−/− cells rescued with 1.3 IFUs of ANO5 virus. Scale bars, 300 µm. (b) Quantitative PCR evaluation of steady-state ANO5 transcript levels, following viral infection, normalized to Gapdh. Error bars indicate SEM. (c) Quantitative PCR evaluation of steady-state, endogenous Ano6 levels normalized to Gapdh. (d) Frequency of nuclei in fibers containing less than five nuclei. Ano5^+/+ represents three independent experiments with three independently isolated cell lines (>6,500 nuclei). Ano5^−/−, ANO5 1.3 IFU, and ANO5 13 IFU represent five independent experiments, with three independently isolated cell lines (>7,000 nuclei per condition). Error bars represent SEM. Significance was determined by one-way ANOVA with Dunnett correction (***, P = 0.0003; **, P = 0.0013). (e) Cumulative frequency of nuclei per fiber size. Nuclei per fiber distributions were evaluated by fitting the data for each replicate in each condition to a single exponential. The exponential constants for each condition were statistically compared to Ano5^−/− using a one-way ANOVA with Dunnett correction (**, P = 0.0095 and *, P = 0.04). (f) Raw frequency histogram representation of data. The values on thex axis represent the center of the bins for number of nuclei per fiber. Error bars represent SEM.