Targeted inactivation of the EGF and amphiregulin genes reveals distinct roles for EGF receptor ligands in mouse mammary gland development

Targeted mice lacking functional EGF or amphiregulin (AR) were derived and bred to the TGF alpha-knockout to generate mice lacking various combination s of the three ligands, In contrast to EGF receptor (EGFR) knockout mice, t riple null mice lacking half of the EGFR ligand family were healthy and fer tile, indicative of overlapping or compensatory functions among EGF family members. Nevertheless, pups born to triple null dams frequently died or wer e runted, suggesting a mammary gland defect. Comparison of individual and c ombinatorial knockouts established that specific loss of AR severely stunte d ductal outgrowth during puberty, consistent with dramatic expression of A R transcripts in normal developing ducts. Surprisingly, loss of all three l igands did not significantly affect cellular proliferation, apoptosis, or E RK activation within terminal end buds. Following pregnancy, most AR single null females, but few triple null females could nurse their young, reveali ng collaborative roles for EGF and TGF alpha in mammopoiesis and lactogenes is. In triple null glands, alveoli were poorly organized and differentiated , and milk protein gene expression was decreased. Additionally, Stat5a acti vation was frequently reduced in AR single and combinatorial nulls in assoc iation with impaired lactation, Collectively, our results provide genetic c onfirmation of a requirement for EGFR signaling throughout the development of the mouse mammary gland, and reveal stage-dependent activities for diffe rent EGFR ligands, Finally, the additional loss of growth factors from pups nursed by triple null dams further worsened their survival and growth, est ablishing functions for both maternal- and neonatal-derived growth factors.