Activation of beta -catenin signaling in differentiated mammary secretory cells induces transdifferentiation into epidermis and squamous metaplasias - PubMed (original) (raw)

Activation of beta -catenin signaling in differentiated mammary secretory cells induces transdifferentiation into epidermis and squamous metaplasias

Keiko Miyoshi et al. Proc Natl Acad Sci U S A. 2002.

Abstract

Mammary anlagen are formed in the embryo as a derivative of the epidermis, a process that is controlled by Lef-1 and therefore possibly by beta-catenin. To investigate the role of beta-catenin signaling in mammary alveolar epithelium, we have stabilized endogenous beta-catenin in differentiating alveolar epithelium through the deletion of exon 3 (amino acids 5-80) of the beta-catenin gene. This task was accomplished in mice carrying a floxed beta-catenin gene and a Cre transgene under control of the mammary-specific whey acidic protein (WAP) gene promoter or the mouse mammary tumor virus-long terminal repeat (MMTV-LTR). Stabilized beta-catenin was obtained during the first pregnancy, and its presence resulted in the dedifferentiation of alveolar epithelium followed by a transdifferentiation into epidermal and pilar structures. Extensive squamous metaplasia, but no adenocarcinomas, developed upon beta-catenin activation during pregnancy and persisted throughout involution. These data demonstrate that the activation of beta-catenin signaling induces a program that results in loss of mammary epithelial cell differentiation and induction of epidermal structures.

PubMed Disclaimer

Figures

Figure 1

Western blot analyses of wild-type mice and mice expressing stabilized β-catenin (Catnb+/Δex3). β-catenin, Stat5a, and actin protein levels during wild-type mammary gland development (Left) and_Catnb_+/Δex3 mice (Right). The arrowheads point to the wild-type (Upper) and truncated (Lower) β-catenin forms, respectively. (Right) The lower bars (#1 and #2) indicate tissue derived from the two respective time points of the same mouse. All mice were mated at 12 wk of age. V, virgin; P, pregnancy; L, lactation; I, involution; V 4w, virgin 4-wk-old; V 10w, virgin 10-wk-old; P5, P16, and P19, pregnancy day 5, 16, and 19; L1 and L12, lactation day 1 and day 12; I2 and I4, involution day 2 and 4; cont., control; and Δβ-cat,Catnb+/Δex3:_WAPCre_mice.

Figure 2

Transdifferentiation of mammary alveolar epithelia to epidermal structures. (A) At pregnancy day 15, the gland was heterogeneously differentiated. Evidence of normal alveolar structures (B) and hyperproliferative epidermal cysts containing ghost cells (C). (D) At lactation day 1, the transdifferentiated lesions were more widespread. (E) Evidence of apparently normal secreting alveoli and (F) hyperproliferative epidermal cysts containing ghost cells. (G) Ten weeks after involution, we preferentially detected squamous metaplasias. (H) Cells were consistently hyperplastic and the center of the nodule was keratinized. (I) Evidence of the epidermal appearance of these structures.

Figure 3

Keratin 1 (K1) and 5 (K5) are expressed in mammary epithelium transdifferentiated into epidermal structures. (A and_A_′) Immunofluorescent staining for K1 (red) and K5 (red) in wild-type epidermis. (B_–_F and_B_′–F_′) Immunofluorescent staining for K1 (red) and K5 (red) and β-catenin (green) in_Catnb+/Δex3 WAPcre mice at pregnancy day 15. K1 was detected in the spinous layer only (A) and K5 in the spinous and basal layers (_A_′). In normal ductal and alveolar structures, no K1 (B and C) was expressed and K5 was found in myoepithelial cells (_B_′ and _C_′). In nodules, β-catenin started to accumulate and cells in the center started to express K1 (D). K5 was only found in the outer layers (_D_′ and_E_′). K1 was detected in the pilar part (E). The nodule next to normal mammary epithelium expressed K1 but not K5 (F and _F_′). K5 was detected in myoepithelial cells (_F_′).

Figure 4

Loss of Npt2b and NKCC1 expression and cytoplasmic accumulation of β-catenin in_Catnb_+/Δex3 WAPcre mice at lactation day 1. (A_–_C) Immunofluorescent staining by using Npt2b (red) and β-catenin (green) primary Abs. (D_–_F) Immunofluorescent staining using NKCC1 (red) and β-catenin (green) primary Abs. (A) Wild-type alveoli showing apical localization of Npt2b (white arrowhead) and basolateral localization of β-catenin. (B) An alveolus showing loss of apical Npt2b (white arrow) adjacent to alveoli exhibiting apical Npt2b similar to wild-type. (C) Cytoplasmic accumulation of β-catenin (white circle) and loss of detectable apical Npt2b. (D) Wild-type alveoli exhibiting colocalization (orange) of NKCC1 and β-catenin in the basolateral membrane. (E) Loss of alveolar NKCC1 expression (white circle) but normal membranous expression of β-catenin. (F) Loss of alveolar NKCC1 expression (white arrowhead) and cytoplasmic accumulation of β-catenin.

Figure 5

Progression of transdifferentiation in_Catnb_+/Δex3 WAPcre mice. (A) Area of apparently normal alveoli with normal membranous localization of α- (red) and β- (green) catenin. The structures appear orange. (B) Stage I: Normal membranous localization of α-catenin, accompanied by local accumulation of β-catenin (arrowhead). (C) Stage II: Normal membranous localization of α-catenin and the formation of cyst-like structures that exhibit accumulation of β-catenin (arrowheads). (D) Stage III: Cysts exhibiting membranous localization of α- and β-catenin (arrowhead), accompanied by the accumulation of ghost cells (GC) in the lumen (nonspecific green staining). (E) Stage IV: Complete loss of membranous localization of α- and β-catenin (arrowheads), and accumulation of ghost cells in the lumen (GC). (F) Stage V: Loss of α-catenin expression, lack of detectable membranous β-catenin expression (arrowheads), and keratin-like deposition in the lumen (KE). Tissue from day 15 of pregnancy (A_–_E); involuted tissue (F).

Figure 6

Stabilization of β-catenin through MMTV-Cre-induced deletion of exon 3 causes metaplasia. (A and B) Hematoxylin/eosin stained section of mammary tissue from a 5-mo-old_Catnb_+/Δex3:MMTVCre mouse after one pregnancy. (C_–_E) Immunofluorescent staining for K1 (red; C), K5 (red;D), and NKCC1 (red, E). The green color represents β-catenin staining. The phenotype was similar to that observed in_Catnb_+/Δex3:WAPCre mice. K1 was detected in the inside of nodules, and K5 was detected in the outer and inner layer of the nodules in_Catnb_+/Δex3:MMTVCre. NKCC1 was partially lost in areas with accumulated β-catenin (arrowhead).

Figure 7

Schematic representation of the differences observed between stabilization of β-catenin through the deletion of exon 3 (loss of amino acids 5–80) in ductal and alveolar epithelium by using MMTV-Cre mice and in alveolar epithelium during pregnancy by using WAP-Cre mice (A), and expression of stabilized β-catenin (loss of amino acids 1–89) by using an MMTV-based transgene (B; ref. 10). (A) Stages of transdifferentiation. Wild-type: Alveoli exhibit basolateral α- and β-catenin staining. Stage I: Local cytoplasmic accumulation of β-catenin. Stage II: Increased local cytoplasmic accumulation of β-catenin, normal membranous localization of α-catenin and the formation of epidermal cysts. Stage III: Accumulation of ghost cells in the lumen of the epidermal cysts accompanied by membranous localization of α- and β-catenin. Stage IV: Complete loss of α- and β-catenin accompanied by increased accumulation of ghost cells in the lumen. Stage V: No definitive α- and β-catenin staining, appearance of keratin-like deposits in the lumen. (B) Stages of differentiation and transformation of mammary epithelium in mice expressing stabilized β-catenin under control of the MMTV-LTR. This diagram is based on published data (10). Precocious alveolar budding occurred during early (4 wk) virgin development. By 12 wk, premature differentiation of alveoli was apparent as assessed by histological analyses and milk protein gene expression. Virgin females (6–8 mo old) presented with adenocarcinomas. α-catenin, red; β-catenin, green; keratin-like deposits, orange.

References

1. McCrea P D, Turck C W, Gumbiner B. Science. 1991;254:1359–1361. -PubMed
1. Roose J, Huls G, van Beest M, Moerer P, van der Horn K, Goldschmeding R, Logtenberg T, Clevers H. Science. 1999;285:1923–1926. -PubMed
1. Hovanes K, Li T W, Munguia J E, Truong T, Milovanovic T, Lawrence Marsh J, Holcombe R F, Waterman M L. Nat Genet. 2001;28:53–57. -PubMed
1. Haegel H, Larue L, Ohsugi M, Fedorov L, Herrenknecht K, Kemler R. Development (Cambridge, UK) 1995;121:3529–3537. -PubMed
1. Huelsken J, Vogel R, Brinkmann V, Erdmann B, Birchmeier C, Birchmeier W. J Cell Biol. 2000;148:567–578. -PMC -PubMed

Activation of beta -catenin signaling in differentiated mammary secretory cells induces transdifferentiation into epidermis and squamous metaplasias - PubMed (original) (raw)