Abstract
BackgroundDuring pregnancy, as the mammary gland prepares for synthesis and delivery of milk to newborns, a luminal mammary epithelial cell (MEC) subpopulation proliferates rapidly in response to systemic hormonal cues that activate STAT5A. While the receptor tyrosine kinase ErbB4 is required for STAT5A activation in MECs during pregnancy, it is unclear how ErbB3, a heterodimeric partner of ErbB4 and activator of phosphatidyl inositol-3 kinase (PI3K) signaling, contributes to lactogenic expansion of the mammary gland.MethodsWe assessed mRNA expression levels by expression microarray of mouse mammary glands harvested throughout pregnancy and lactation. To study the role of ErbB3 in mammary gland lactogenesis, we used transgenic mice expressing WAP-driven Cre recombinase to generate a mouse model in which conditional ErbB3 ablation occurred specifically in alveolar mammary epithelial cells (aMECs).ResultsProfiling of RNA from mouse MECs isolated throughout pregnancy revealed robust Erbb3 induction during mid-to-late pregnancy, a time point when aMECs proliferate rapidly and undergo differentiation to support milk production. Litters nursed by ErbB3KO dams weighed significantly less when compared to litters nursed by ErbB3WT dams. Further analysis revealed substantially reduced epithelial content, decreased aMEC proliferation, and increased aMEC cell death during late pregnancy. Consistent with the potent ability of ErbB3 to activate cell survival through the PI3K/Akt pathway, we found impaired Akt phosphorylation in ErbB3KO samples, as well as impaired expression of STAT5A, a master regulator of lactogenesis. Constitutively active Akt rescued cell survival in ErbB3-depleted aMECs, but failed to restore STAT5A expression or activity. Interestingly, defects in growth and survival of ErbB3KO aMECs as well as Akt phosphorylation, STAT5A activity, and expression of milk-encoding genes observed in ErbB3KO MECs progressively improved between late pregnancy and lactation day 5. We found a compensatory upregulation of ErbB4 activity in ErbB3KO mammary glands. Enforced ErbB4 expression alleviated the consequences of ErbB3 ablation in aMECs, while combined ablation of both ErbB3 and ErbB4 exaggerated the phenotype.ConclusionsThese studies demonstrate that ErbB3, like ErbB4, enhances lactogenic expansion and differentiation of the mammary gland during pregnancy, through activation of Akt and STAT5A, two targets crucial for lactation.
Highlights
During pregnancy, as the mammary gland prepares for synthesis and delivery of milk to newborns, a luminal mammary epithelial cell (MEC) subpopulation proliferates rapidly in response to systemic hormonal cues that activate signal transducer and activator of transcription 5A (STAT5A)
We assessed Erbb3 mRNA levels in expression microarray dUTP nick end-labeling (TUNEL) analysis, revealing datasets derived from mouse mammary glands harvested markedly increased terminal dUTP nick end-labeling (TUNEL)+ cells in ErbB3FL/FL × WAPi-Cre (ErbB3KO) mammary at distinct developmental time points of pregnancy and glands at 16.5 d.p.c. and lactation day 1 (L1), no differences in the F1 lactation [52] (Fig. 1a), revealing relatively low Erbb3 dur- percentage of TUNEL+ MECs were seen by L5 (Fig. 2b; ing early pregnancy (1–7 days post coitus (d.p.c.)) but ap- Additional file 1: Figure S4)
To in- may contribute to the plasticity that exists between vestigate the impact of ErbB3 ablation from Alveolar mammary epithelial cell (aMEC), we crossed mice harboring floxed Erbb3 alleles (Erbb3FL/FL)
Summary
As the mammary gland prepares for synthesis and delivery of milk to newborns, a luminal mammary epithelial cell (MEC) subpopulation proliferates rapidly in response to systemic hormonal cues that activate STAT5A. As the mammary gland prepares for synthesis and delivery of milk to newborns, an apically located (i.e., luminal) MEC subpopulation proliferates rapidly in response to systemic hormonal cues such as progesterone and prolactin (PRL), as well as locally derived cues such as neuregulin (NRG, known as heregulin). At parturition, these milk-producing MECs, referred to as alveolar MECs (aMECs), undergo terminal differentiation for milk synthesis and delivery [8]. Engineered mouse models (GEMMs) confirmed that the PRL/PRLR/Jak2/STAT5A signaling axis is crucial for lactogenesis [17,18,19]
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