Signal transducer and activator of transcription (Stat) 5 controls the proliferation and differentiation of mammary alveolar epithelium

Functional development of mammary epithelium during pregnancy depends on prolactin signaling. However, the underlying molecular and cellular events are not fully understood. We examined the specific contributions of the prolactin receptor (PrlR) and the signal transducers and activators of transcription 5a and 5b (referred to as Stat5) in the formation and differentiation of mammary alveolar epithelium. PrlR- and Stat5-null mammary epithelia were transplanted into wild-type hosts, and pregnancy-mediated development was investigated at a histological and molecular level. Stat5-null mammary epithelium developed ducts but failed to form alveoli, and no milk protein gene expression was observed. In contrast, PrlR-null epithelium formed alveoli-like structures with small open lumina. Electron microscopy revealed undifferentiated features of organelles and a perturbation of cell–cell contacts in PrlR- and Stat5-null epithelia. Expression of NKCC1, an Na-K-Cl cotransporter characteristic for ductal epithelia, and ZO-1, a protein associated with tight junction, were maintained in the alveoli-like structures of PrlR- and Stat5-null epithelia. In contrast, the Na-Pi cotransporter Npt2b, and the gap junction component connexin 32, usually expressed in secretory epithelia, were undetectable in PrlR- and Stat5-null mice. These data demonstrate that signaling via the PrlR and Stat5 is critical for the proliferation and differentiation of mammary alveoli during pregnancy.

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The Role of Glucocorticoids in Pregnancy, Parturition, Lactation, and Nurturing in Melanocortin Receptor 2-Deficient Mice

Endocrinology ◽

10.1210/en.2010-0935 ◽

2011 ◽

Vol 152 (4) ◽

pp. 1652-1660 ◽

Cited By ~ 16

Author(s):

Dai Chida ◽

Keiko Miyoshi ◽

Tsuyoshi Sato ◽

Tetsuya Yoda ◽

Takefumi Kikusui ◽

...

Keyword(s):

Mammary Epithelium ◽

Milk Proteins ◽

Myoepithelial Cells ◽

Melanocortin Receptor ◽

Milk Protein Gene ◽

Fetal Survival ◽

Pup Retrieval ◽

Milk Protein Gene Expression ◽

In Pregnancy

Abstract Maternal glucocorticoids are critical for fetal development, but overexpression can be deleterious. Previously we established a mouse line deficient in melanocortin receptor 2 (MC2R). MC2R−/− mice have undetectable levels of corticosterone despite high levels of ACTH and defects resembling those in patients with familial glucocorticoid deficiency. Here we analyzed the role of glucocorticoids in pregnancy, parturition, lactation, and nurturing in MC2R−/− mice. MC2R−/− mice were fertile and produced normal litters when crossed with MC2R+/+ mice. However, MC2R−/− females crossed with MC2R−/− males had no live births, and approximately 20% of the embryos at d 18.5 of pregnancy were of normal body size but were dead when born. MC2R−/− pregnant females crossed with MC2R+/+ males had detectable serum corticosterone levels, suggesting the transplacental passage of corticosterone from fetus to mother. MC2R+/− pups delivered from MC2R−/− females crossed with MC2R+/+ males mice thrived poorly with MC2R−/− mothers but grew to adulthood when transferred to foster mothers after birth, suggesting that MC2R−/− females are poor mothers or cannot nurse. MC2R−/− females had normal alveoli, but penetration of mammary epithelium into fat pads and expression of milk proteins were reduced. Myoepithelial cells, which force milk out of the alveoli, were fully developed and differentiated. Pup retrieval behavior was normal in MC2R−/− mice. Exogenous corticosterone rescued expression of milk proteins in MC2R−/− mothers, and the pups of treated mothers grew to adulthood. Our results reveal the importance of glucocorticoids for fetal survival late in pregnancy, mammary gland development, and milk protein gene expression.

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Sustained activation of STAT5 is essential for chromatin remodeling and maintenance of mammary-specific function

The Journal of Cell Biology ◽

10.1083/jcb.200807021 ◽

2009 ◽

Vol 184 (1) ◽

pp. 57-66 ◽

Cited By ~ 75

Author(s):

Ren Xu ◽

Celeste M. Nelson ◽

John L. Muschler ◽

Mandana Veiseh ◽

Barbara K. Vonderhaar ◽

...

Keyword(s):

Prolactin Receptor ◽

Functional Differentiation ◽

Specific Gene ◽

Tissue Specific ◽

Signal Transducers ◽

Prolactin Signaling ◽

Transcription Factor Activation ◽

Mammary Epithelia ◽

Necessary And Sufficient ◽

Sustained Activation

Epithelial cells, once dissociated and placed in two-dimensional (2D) cultures, rapidly lose tissue-specific functions. We showed previously that in addition to prolactin, signaling by laminin-111 was necessary to restore functional differentiation of mammary epithelia. Here, we elucidate two additional aspects of laminin-111 action. We show that in 2D cultures, the prolactin receptor is basolaterally localized and physically segregated from its apically placed ligand. Detachment of the cells exposes the receptor to ligation by prolactin leading to signal transducers and activators of transcription protein 5 (STAT5) activation, but only transiently and not sufficiently for induction of milk protein expression. We show that laminin-111 reorganizes mammary cells into polarized acini, allowing both the exposure of the prolactin receptor and sustained activation of STAT5. The use of constitutively active STAT5 constructs showed that the latter is necessary and sufficient for chromatin reorganization and β-casein transcription. These results underscore the crucial role of continuous laminin signaling and polarized tissue architecture in maintenance of transcription factor activation, chromatin organization, and tissue-specific gene expression.

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Inactivation of Stat5 in Mouse Mammary Epithelium during Pregnancy Reveals Distinct Functions in Cell Proliferation, Survival, and Differentiation

Molecular and Cellular Biology ◽

10.1128/mcb.24.18.8037-8047.2004 ◽

2004 ◽

Vol 24 (18) ◽

pp. 8037-8047 ◽

Cited By ~ 352

Author(s):

Yongzhi Cui ◽

Greg Riedlinger ◽

Keiko Miyoshi ◽

Wei Tang ◽

Cuiling Li ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Death ◽

Mammary Gland ◽

Mammary Epithelium ◽

Mouse Mammary Gland ◽

Signal Transducers ◽

Proliferation And Differentiation ◽

Transgenic Lines ◽

Mammary Gland Morphogenesis ◽

Gland Morphogenesis

ABSTRACT This study explored the functions of the signal transducers and activators of transcription 5a and 5b (referred to as Stat5 here) during different stages of mouse mammary gland development by using conditional gene inactivation. Mammary gland morphogenesis includes cell specification, proliferation and differentiation during pregnancy, cell survival and maintenance of differentiation throughout lactation, and cell death during involution. Stat5 is activated by prolactin, and its presence is mandatory for the proliferation and differentiation of mammary epithelium during pregnancy. To address the question of whether Stat5 is also necessary for the maintenance and survival of the differentiated epithelium, the two genes were deleted at different time points. The 110-kb Stat5 locus in the mouse was bracketed with loxP sites, and its deletion was accomplished by using two Cre-expressing transgenic lines. Loss of Stat5 prior to pregnancy prevented epithelial proliferation and differentiation. Deletion of Stat5 during pregnancy, after mammary epithelium had entered Stat5-mediated differentiation, resulted in premature cell death, indicating that at this stage epithelial cell proliferation, differentiation, and survival require Stat5.

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Epithelial Xbp1 Is Required for Cellular Proliferation and Differentiation during Mammary Gland Development

Molecular and Cellular Biology ◽

10.1128/mcb.00136-15 ◽

2015 ◽

Vol 35 (9) ◽

pp. 1543-1556 ◽

Cited By ~ 19

Author(s):

Daisuke Hasegawa ◽

Veronica Calvo ◽

Alvaro Avivar-Valderas ◽

Abigale Lade ◽

Hsin-I Chou ◽

...

Keyword(s):

Mammary Gland ◽

Mammary Gland Development ◽

Cellular Proliferation ◽

Prolactin Receptor ◽

Mammary Epithelium ◽

Branching Morphogenesis ◽

Direct Function ◽

Proliferation And Differentiation ◽

The Unfolded Protein Response ◽

Gland Development

Xbp1, a key mediator of the unfolded protein response (UPR), is activated by IRE1α-mediated splicing, which results in a frameshift to encode a protein with transcriptional activity. However, the direct function of Xbp1 in epithelial cells during mammary gland development is unknown. Here we report that the loss of Xbp1 in the mammary epithelium through targeted deletion leads to poor branching morphogenesis, impaired terminal end bud formation, and spontaneous stromal fibrosis during the adult virgin period. Additionally, epithelial Xbp1 deletion induces endoplasmic reticulum (ER) stress in the epithelium and dramatically inhibits epithelial proliferation and differentiation during lactation. The synthesis of milk and its major components, α/β-casein and whey acidic protein (WAP), is significantly reduced due to decreased prolactin receptor (Prlr) and ErbB4 expression in Xbp1-deficient mammary epithelium. Reduction of Prlr and ErbB4 expression and their diminished availability at the cell surface lead to reduced phosphorylated Stat5, an essential regulator of cell proliferation and differentiation during lactation. As a result, lactating mammary glands in these mice produce less milk protein, leading to poor pup growth and postnatal death. These findings suggest that the loss of Xbp1 induces a terminal UPR which blocks proliferation and differentiation during mammary gland development.

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Hormone-dependent milk protein gene expression in bovine mammary explants from biopsies at different stages of pregnancy

Journal of Dairy Research ◽

10.1017/s0022029904000068 ◽

2004 ◽

Vol 71 (2) ◽

pp. 135-140 ◽

Cited By ~ 14

Author(s):

Paul A Sheehy ◽

James J Della-Vedova ◽

Kevin R Nicholas ◽

Peter C Wynn

Keyword(s):

Gene Expression ◽

Milk Protein ◽

Protein Gene ◽

Bovine Milk ◽

Mammary Tissue ◽

Endocrine Regulation ◽

Surgical Biopsy ◽

Milk Protein Gene ◽

Milk Protein Gene Expression

A method for the collection of mammary biopsies developed previously was refined and used to study the endocrine regulation of bovine milk protein gene expression. Our surgical biopsy method used real-time ultrasound imaging and epidural analgesia to enable recovery of a sufficient quantity of mammary tissue from late-pregnant dairy cows for explant culture in vitro. The time of biopsy was critical for prolactin-dependent induction of milk protein gene expression in mammary explants, as only mammary tissue from cows nearing 30 d prepartum was hormone-responsive. This suggests that during the later stages of pregnancy a change in the responsiveness of milk protein gene expression to endocrine stimuli occurred in preparation for lactation. This may relate to the diminution of a putative population of undifferentiated cells that were still responsive to prolactin. Alternatively, the metabolic activity of the tissue had increased to the level whereby the response of the tissue was no longer assessable using this model in vitro.

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Hormonal control of milk protein gene expression during late pregnancy and post partum in the tammar wallaby

Livestock Production Science ◽

10.1016/0301-6226(93)90221-3 ◽

1993 ◽

Vol 35 (1-2) ◽

pp. 195-196

Keyword(s):

Gene Expression ◽

Milk Protein ◽

Protein Gene ◽

Post Partum ◽

Late Pregnancy ◽

Tammar Wallaby ◽

Hormonal Control ◽

Milk Protein Gene ◽

Milk Protein Gene Expression

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Synergistic Actions of Glucocorticoid and Prolactin in Murine Milk-Protein Gene Expression

Actions of Prolactin on Molecular Processes ◽

10.1201/9781351069458-7 ◽

2018 ◽

pp. 121-135 ◽

Cited By ~ 1

Author(s):

Mihir R. Banerjee ◽

Ravi S. Menon

Keyword(s):

Gene Expression ◽

Milk Protein ◽

Protein Gene ◽

Milk Protein Gene ◽

Milk Protein Gene Expression

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Transforming growth factor beta3 induces cell death during the first stage of mammary gland involution

Development ◽

10.1242/dev.127.14.3107 ◽

2000 ◽

Vol 127 (14) ◽

pp. 3107-3118 ◽

Cited By ~ 2

Author(s):

A.V. Nguyen ◽

J.W. Pollard

Keyword(s):

Cell Death ◽

Mammary Gland ◽

Transforming Growth Factor ◽

Alveolar Epithelium ◽

Mammary Epithelium ◽

Significant Inhibition ◽

Mammary Tissue ◽

Lactogenic Hormone ◽

Null Mutant Mice ◽

Transforming Growth Factor Beta3

Involution of the mammary gland following weaning is divided into two distinct phases. Initially, milk stasis results in the induction of local factors that cause apoptosis in the alveolar epithelium. Secondly after a prolonged absence of suckling, the consequent decline in circulating lactogenic hormone concentrations initiates remodeling of the mammary gland to the virgin-like state. We have shown that immediately following weaning TGFbeta3 mRNA and protein is rapidly induced in the mammary epithelium and that this precedes the onset of apoptosis. Unilateral inhibition of suckling and hormonal reconstitution experiments showed that TGFbeta3 induction is regulated by milk stasis and not by the circulating hormonal concentration. Directed expression of TGFbeta3 in the alveolar epithelium of lactating mice using a beta-lactoglobulin promoter mobilized SMAD4 translocation to the nucleus and caused apoptosis of these cells, but not tissue remodeling. Transplantation of neonatal mammary tissue derived from TGFbeta3 null mutant mice into syngenic hosts resulted in a significant inhibition of cell death compared to wild-type mice upon milk stasis. These results provide direct evidence that TGFbeta3 is a local mammary factor induced by milk stasis that causes apoptosis in the mammary gland epithelium during involution.

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