4ICD induced mammary epithelial cell differentiation requires an intact Y984. (A) HER4/4ICD western blot analysis of HC11 mammary epithelial stable cell lines HC11/vector, HC11/4ICD, and HC11/4ICD-Y984F demonstrating equivalent levels of 4ICD and 4ICD-Y984F expression. Western blot of α-tubulin is included as a loading control. Each indicated HC11 stable cell line was induced to undergo differentiation in the absence of exogenous prolactin by serum starvation for two days and expression, normalized to β-actin, of the differentiation markers (B) β-casein and (C) WAP relative to HC11/vector control was determined by quantitative RT-PCR. Each quantitative RT-PCR sample was prepared in triplicate and the data represent the mean and standard error (SE) of at least three independent experiments and RNA extractions. Statistically significant differences between data sets were determined using one-way ANOVA with Bonferroni post-hoc test. Single or double asterisks indicates p<0.03 or p=0.003, respectively.